Controlling Pericellular Oxygen Tension in Cell Culture Reveals Distinct Breast Cancer Responses to Low Oxygen Tensions.

In oxygen (O2)-controlled cell culture, an indispensable tool in biological research, it is presumed that the incubator setpoint equals the O2 tension experienced by cells (i.e., pericellular O2). However, it is discovered that physioxic (5% O2) and hypoxic (1% O2) setpoints regularly induce anoxic (0% O2) pericellular tensions in both adherent and suspension cell cultures. Electron transport chain inhibition ablates this effect, indicating that cellular O2 consumption is the driving factor. RNA-seq analysis revealed that primary human hepatocytes cultured in physioxia experience ischemia-reperfusion injury due to cellular O2 consumption. A reaction-diffusion model is developed to predict pericellular O2 tension a priori, demonstrating that the effect of cellular O2 consumption has the greatest impact in smaller volume culture vessels. By controlling pericellular O2 tension in cell culture, it is found that hypoxia vs. anoxia induce distinct breast cancer transcriptomic and translational responses, including modulation of the hypoxia-inducible factor (HIF) pathway and metabolic reprogramming. Collectively, these findings indicate that breast cancer cells respond non-monotonically to low O2, suggesting that anoxic cell culture is not suitable for modeling hypoxia. Furthermore, it is shown that controlling atmospheric O2 tension in cell culture incubators is insufficient to regulate O2 in cell culture, thus introducing the concept of pericellular O2-controlled cell culture.

HIF-stabilizing biomaterials: from hypoxia-mimicking to hypoxia-inducing.

Recent advances in our understanding of hypoxia and hypoxia-mediated mechanisms shed light on the critical implications of the hypoxic stress on cellular behavior. However, tools emulating hypoxic conditions (i.e., low oxygen tensions) for research are limited and often suffer from major shortcomings, such as lack of reliability and off-target effects, and they usually fail to recapitulate the complexity of the tissue microenvironment. Fortunately, the field of biomaterials is constantly evolving and has a central role to play in the development of new technologies for conducting hypoxia-related research in several aspects of biomedical research, including tissue engineering, cancer modeling, and modern drug screening. In this perspective, we provide an overview of several strategies that have been investigated in the design and implementation of biomaterials for simulating or inducing hypoxic conditions-a prerequisite in the stabilization of hypoxia-inducible factor (HIF), a master regulator of the cellular responses to low oxygen. To this end, we discuss various advanced biomaterials, from those that integrate hypoxia-mimetic agents to artificially induce hypoxia-like responses, to those that deplete oxygen and consequently create either transient (<1 day) or sustained (>1 day) hypoxic conditions. We also aim to highlight the advantages and limitations of these emerging biomaterials for biomedical applications, with an emphasis on cancer research.

Controlling pericellular oxygen tension in cell culture reveals distinct breast cancer responses to low oxygen tensions.

Oxygen (O2) tension plays a key role in tissue function and pathophysiology. O2-controlled cell culture, in which the O2 concentration in an incubator's gas phase is controlled, is an indispensable tool to study the role of O2 in vivo. For this technique, it is presumed that the incubator setpoint is equal to the O2 tension that cells experience (i.e., pericellular O2). We discovered that physioxic (5% O2) and hypoxic (1% O2) setpoints regularly induce anoxic (0.0% O2) pericellular tensions in both adherent and suspension cell cultures. Electron transport chain inhibition ablates this effect, indicating that cellular O2 consumption is the driving factor. RNA-seq revealed that primary human hepatocytes cultured in physioxia experience ischemia-reperfusion injury due to anoxic exposure followed by rapid reoxygenation. To better understand the relationship between incubator gas phase and pericellular O2 tensions, we developed a reaction-diffusion model that predicts pericellular O2 tension a priori. This model revealed that the effect of cellular O2 consumption is greatest in smaller volume culture vessels (e.g., 96-well plate). By controlling pericellular O2 tension in cell culture, we discovered that MCF7 cells have stronger glycolytic and glutamine metabolism responses in anoxia vs. hypoxia. MCF7 also expressed higher levels of HIF2A, CD73, NDUFA4L2, etc. and lower levels of HIF1A, CA9, VEGFA, etc. in response to hypoxia vs. anoxia. Proteomics revealed that 4T1 cells had an upregulated epithelial-to-mesenchymal transition (EMT) response and downregulated reactive oxygen species (ROS) management, glycolysis, and fatty acid metabolism pathways in hypoxia vs. anoxia. Collectively, these results reveal that breast cancer cells respond non-monotonically to low O2, suggesting that anoxic cell culture is not suitable to model hypoxia. We demonstrate that controlling atmospheric O2 tension in cell culture incubators is insufficient to control O2 in cell culture and introduce the concept of pericellular O2-controlled cell culture.

Hypoxia-inducing cryogels uncover key cancer-immune cell interactions in an oxygen-deficient tumor microenvironment.

Hypoxia is a major factor shaping the immune landscape, and several cancer models have been developed to emulate hypoxic tumors. However, to date, they still have several limitations, such as the lack of reproducibility, inadequate biophysical cues, limited immune cell infiltration, and poor oxygen (O2) control, leading to non-pathophysiological tumor responses. Therefore, it is essential to develop better cancer models that mimic key features of the tumor extracellular matrix and recreate tumor-associated hypoxia while allowing cell infiltration and cancer-immune cell interactions. Herein, hypoxia-inducing cryogels (HICs) have been engineered using hyaluronic acid (HA) to fabricate three-dimensional microtissues and model a hypoxic tumor microenvironment. Specifically, tumor cell-laden HICs have been designed to deplete O2 locally and induce long-standing hypoxia. HICs promoted changes in hypoxia-responsive gene expression and phenotype, a metabolic adaptation to anaerobic glycolysis, and chemotherapy resistance. Additionally, HIC-supported tumor models induced dendritic cell (DC) inhibition, revealing a phenotypic change in the plasmacytoid DC (pDC) subset and an impaired conventional DC (cDC) response in hypoxia. Lastly, our HIC-based melanoma model induced CD8+ T cell inhibition, a condition associated with the downregulation of pro-inflammatory cytokine secretion, increased expression of immunomodulatory factors, and decreased degranulation and cytotoxic capacity of T cells. Overall, these data suggest that HICs can be used as a tool to model solid-like tumor microenvironments and has great potential to deepen our understanding of cancer-immune cell relationship in low O2 conditions and may pave the way for developing more effective therapies.

704/DNA vaccines leverage cytoplasmic DNA stimulation to promote anti-HIV neutralizing antibody production in mice and strong immune response against alpha-fetoprotein in non-human primates.

Genetic immunization is an attractive approach for prophylactic and therapeutic vaccination using synthetic vectors to deliver antigen-encoding nucleic acids. Recently, DNA delivered by a physical means or RNA by liposomes consisting of four different lipids demonstrated good protection in human phase III clinical trials and received Drugs Controller General of India and US FDA approval to protect against COVID-19, respectively. However, the development of a system allowing for efficient and simple delivery of nucleic acids while improving immune response priming has the potential to unleash the full therapeutic potential of genetic immunization. DNA-based gene therapies and vaccines have the potential for rapid development, as exemplified by the recent approval of Collategene, a gene therapy to treat human critical limb ischemia, and ZyCoV, a DNA vaccine delivered by spring-powered jet injector to protect against SARS-CoV2 infection. Recently, we reported amphiphilic block copolymer 704 as a promising synthetic vector for DNA vaccination in various models of human diseases. This vector allows dose sparing of antigen-encoding plasmid DNA. Here, we report the capacity of 704-mediated HIV and anti-hepatocellular carcinoma DNA vaccines to induce the production of specific antibodies against gp120 HIV envelope proteins in mice and against alpha-fetoprotein antigen in non-human primates, respectively. An investigation of the underlying mechanisms showed that 704-mediated vaccination did trigger a strong immune response by (1) allowing a direct DNA delivery into the cytosol, (2) promoting an intracytoplasmic DNA sensing leading to both interferon and NF-κB cascade stimulation, and (3) inducing antigen expression by muscle cells and presentation by antigen-presenting cells, leading to the induction of a robust adaptive response. Overall, our findings suggest that the 704-mediated DNA vaccination platform is an attractive method to develop both prophylactic and therapeutic vaccines.

Engineering injectable, biocompatible, and highly elastic bioadhesive cryogels.

The extracellular matrix (ECM), an integral component of all organs, is inherently tissue adhesive and plays a pivotal role in tissue regeneration and remodeling. However, man-made three-dimensional (3D) biomaterials that are designed to mimic ECMs do not intrinsically adhere to moisture-rich environments and often lack an open macroporous architecture required for facilitating cellularization and integration with the host tissue post-implantation. Furthermore, most of these constructs usually entail invasive surgeries and potentially a risk of infection. To address these challenges, we recently engineered biomimetic and macroporous cryogel scaffolds that are syringe injectable while exhibiting unique physical properties, including strong bioadhesive properties to tissues and organs. These biomimetic catechol-containing cryogels were prepared from naturally-derived polymers such as gelatin and hyaluronic acid and were functionalized with mussel-inspired dopamine (DOPA) to impart bioadhesive properties. We found that using glutathione as an antioxidant and incorporating DOPA into cryogels via a PEG spacer arm led to the highest tissue adhesion and improved physical properties overall, whereas DOPA-free cryogels were weakly tissue adhesive. As shown by qualitative and quantitative adhesion tests, DOPA-containing cryogels were able to adhere strongly to several animal tissues and organs such as the heart, small intestine, lung, kidney, and skin. Furthermore, these unoxidized (i.e., browning-free) and bioadhesive cryogels showed negligible cytotoxicity toward murine fibroblasts and prevented the ex vivo activation of primary bone marrow-derived dendritic cells. Finally, in vivo data suggested good tissue integration and a minimal host inflammatory response when subcutaneously injected in rats. Collectively, these minimally invasive, browning-free, and strongly bioadhesive mussel-inspired cryogels show great promise for various biomedical applications, potentially in wound healing, tissue engineering, and regenerative medicine.

Hypoxia-inducing cryogels uncover key cancer-immune cell interactions in an oxygen-deficient tumor microenvironment.

Hypoxia, an important feature of solid tumors, is a major factor shaping the immune landscape, and several cancer models have been developed to emulate hypoxic tumors. However, to date, they still have several limitations, such as the lack of reproducibility, inadequate biophysical cues, limited immune cell infiltration, and poor oxygen (O 2 ) control, leading to non-pathophysiological tumor responses. As a result, it is essential to develop new and improved cancer models that mimic key features of the tumor extracellular matrix and recreate tumor-associated hypoxia while allowing cell infiltration and cancer-immune cell interactions. Herein, hypoxia-inducing cryogels (HICs) have been engineered using hyaluronic acid (HA) as macroporous scaffolds to fabricate three-dimensional microtissues and model a hypoxic tumor microenvironment. Specifically, tumor cell-laden HICs have been designed to deplete O 2 locally and induce long-standing hypoxia. This state of low oxygen tension, leading to HIF-1α stabilization in tumor cells, resulted in changes in hypoxia-responsive gene expression and phenotype, a metabolic adaptation to anaerobic glycolysis, and chemotherapy resistance. Additionally, HIC-supported tumor models induced dendritic cell (DC) inhibition, revealing a phenotypic change in plasmacytoid B220 + DC (pDC) subset and an impaired conventional B220 - DC (cDC) response in hypoxia. Lastly, our HIC-based melanoma model induced CD8+ T cell inhibition, a condition associated with the downregulation of pro-inflammatory cytokine secretion, increased expression of immunomodulatory factors, and decreased degranulation and cytotoxic capacity of T cells. Overall, these data suggest that HICs can be used as a tool to model solid-like tumor microenvironments and identify a phenotypic transition from cDC to pDC in hypoxia and the key contribution of HA in retaining cDC phenotype and inducing their hypoxia-mediated immunosuppression. This technology has great potential to deepen our understanding of the complex relationships between cancer and immune cells in low O 2 conditions and may pave the way for developing more effective therapies.

Biomaterial-assisted local oxygenation safeguards the prostimulatory phenotype and functions of human dendritic cells in hypoxia.

Dendritic cells (DCs), professional antigen-presenting cells, function as sentinels of the immune system. DCs initiate and fine-tune adaptive immune responses by presenting antigenic peptides to B and T lymphocytes to mount an effective immune response against cancer and pathogens. However, hypoxia, a condition characterized by low oxygen (O2) tension in different tissues, significantly impacts DC functions, including antigen uptake, activation and maturation, migration, as well as T-cell priming and proliferation. In this study, we employed O2-releasing biomaterials (O2-cryogels) to study the effect of localized O2 supply on human DC phenotype and functions. Our results indicate that O2-cryogels effectively mitigate DC exposure to hypoxia under hypoxic conditions. Additionally, O2-cryogels counteract hypoxia-induced inhibition of antigen uptake and migratory activity in DCs through O2 release and hyaluronic acid (HA) mediated mechanisms. Furthermore, O2-cryogels preserve and restore DC maturation and co-stimulation markers, including HLA-DR, CD86, and CD40, along with the secretion of proinflammatory cytokines in hypoxic conditions. Finally, our findings demonstrate that the supplemental O2 released from the cryogels preserves DC-mediated T-cell priming, ultimately leading to the activation and proliferation of allogeneic CD3+ T cells. This work emphasizes the potential of local oxygenation as a powerful immunomodulatory agent to improve DC activation and functions in hypoxia, offering new approaches for cancer and infectious disease treatments.

Engineering hyaluronic acid-based cryogels for CD44-mediated breast tumor reconstruction.

Breast cancer is a major health concern worldwide and is the leading cause of cancer-related death among American women. Traditional therapies, such as surgery, chemotherapy, and radiotherapy, are usually ineffective. Furthermore, cancer recurrence following targeted therapy often results from acquired drug resistance. Therefore, more realistic tumor models than monolayer cell culture for drug screening and discovery in an in vitro setting would facilitate the development of new therapeutic strategies. Toward this goal, we first developed a simple, rapid, low-cost, and high-throughput method for generating uniform multi-cellular tumor spheroids (MCTS) with controllable size. Next, biomimetic cryogel scaffolds fabricated from hyaluronic acid (HA) were utilized as a platform to reconstruct breast tumor microtissues with aspects of the complex tumor microenvironment in three dimensions. Finally, we investigated the interactions between the HA-based cryogels and CD44-positive breast tumor cells, individually or as MCTS. We found that incorporating the adhesive RGD peptide in cryogels led to the formation of a monolayer of tumor cells on the polymer walls, whereas MCTS cultured on RGD-free HA cryogels resulted in the growth of large and dense microtumors, more similar to native tumor masses. As a result, the MCTS-laden HA cryogel system induced a highly aggressive and chemotherapy drug-resistant tumor model. RGD-free HA-based cryogels represent an effective starting point for designing tumor models for preclinical research, therapeutic drug screening, and early cancer diagnosis.

Biomaterials and Oxygen Join Forces to Shape the Immune Response and Boost COVID-19 Vaccines.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to an unprecedented global health crisis, resulting in a critical need for effective vaccines that generate protective antibodies. Protein subunit vaccines represent a promising approach but often lack the immunogenicity required for strong immune stimulation. To overcome this challenge, it is first demonstrated that advanced biomaterials can be leveraged to boost the effectiveness of SARS-CoV-2 protein subunit vaccines. Additionally, it is reported that oxygen is a powerful immunological co-adjuvant and has an ability to further potentiate vaccine potency. In preclinical studies, mice immunized with an oxygen-generating coronavirus disease 2019 (COVID-19) cryogel-based vaccine (O2-CryogelVAX) exhibit a robust Th1 and Th2 immune response, leading to a sustained production of highly effective neutralizing antibodies against the virus. Even with a single immunization, O2-CryogelVAX achieves high antibody titers within 21 days, and both binding and neutralizing antibody levels are further increased after a second dose. Engineering a potent vaccine system that generates sufficient neutralizing antibodies after one dose is a preferred strategy amid vaccine shortage. The data suggest that this platform is a promising technology to reinforce vaccine-driven immunostimulation and is applicable to current and emerging infectious diseases.

Injectable Lignin-co-Gelatin Cryogels with Antioxidant and Antibacterial Properties for Biomedical Applications.

For several biomedical applications, it is essential to develop novel bioactive materials. Such biomaterials could potentially improve wound healing, prevent infections, or be used in immunoengineering. For example, bioactive materials that reduce oxidative stress without relying on antibiotics and other drugs could be beneficial. Hydrogel-based biomaterials, especially those derived from natural polymers, have been regarded as one of the most promising scaffolds for biomedical research. These multifunctional scaffolds can exhibit high water adsorption capacity, biocompatibility, and biomechanical properties that can match native tissues. Cryogels are a special type of hydrogels in which polymers are cross-linked around ice crystals. As a result, cryogels exhibit unique physical features, including a macroporous and interconnected network, flexibility, shape-memory properties, and syringe injectability. Herein, we developed a multifunctional, i.e., antibacterial, antioxidant, and injectable cryogel by combining lignin with gelatin. The cryogel with 0.2% lignin showed a compressive modulus of 25 kPa and a compressive stress of 140 kPa at 80% strain, which is, respectively, 1.8 and 7 times higher than those of the pure gelatin cryogels. Meanwhile, such a cryogel formulation could completely recover its shape after compression up to 90% and was needle-injectable. Additionally, the lignin-co-gelatin cryogel with 0.1-0.2 lignin showed 8-10 mm of inhibition zone against the most common surgical site infection-associated pathogenic bacteria. Furthermore, lignin-co-gelatin cryogel was found to scavenge free radicals and have good cytocompatibility, and the cryogels with up to 0.2% lignin minimally activate naïve mouse bone marrow-derived dendritic cells. Overall, the current approach shows great promise for the design of bioresource-based multifunctional cryogels for a wide range of biomedical applications.

Harnessing biomaterials for therapeutic strategies against COVID-19.

With the emergence of the coronavirus disease 2019 (COVID-19), the world is experiencing a profound human health crisis. The number of infections and deaths due to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to increase every minute, pinpointing major shortcomings in our ability to prevent viral outbreaks. Although several COVID-19 vaccines have been recently approved for emergency use, therapeutic options remain limited, and their long-term potency has yet to be validated. Biomaterials science has a pivotal role to play in pushing the boundaries of emerging technologies for antiviral research and treatment. In this perspective, we discuss how biomaterials can be harnessed to develop accurate COVID-19 infection models, enhance antiviral drug delivery, foster new antiviral strategies, and boost vaccine efficacy. These efforts will not only contribute to stop or mitigate the current pandemic but will also provide unorthodox platforms to understand, prevent, and protect us from future viral outbreaks.

Oxygen-Generating Cryogels Restore T Cell Mediated Cytotoxicity in Hypoxic Tumors.

Solid tumors are protected from antitumor immune responses due to their hypoxic microenvironments. Weakening hypoxia-driven immunosuppression by hyperoxic breathing of 60% oxygen has shown to be effective in unleashing antitumor immune cells against solid tumors. However, efficacy of systemic oxygenation is limited against solid tumors outside of lungs and has been associated with unwanted side effects. As a result, it is essential to develop targeted oxygenation alternatives to weaken tumor hypoxia as novel approaches to restore immune responses against cancer. Herein, we report on injectable oxygen-generating cryogels (O2-cryogels) to reverse tumor-induced hypoxia. These macroporous biomaterials were designed to locally deliver oxygen, inhibit the expression of hypoxia-inducible genes in hypoxic melanoma cells, and reduce the accumulation of immunosuppressive extracellular adenosine. Our data show that O2-cryogels enhance T cell-mediated secretion of cytotoxic proteins, restoring the killing ability of tumor-specific CTLs, both in vitro and in vivo. In summary, O2-cryogels provide a unique and safe platform to supply oxygen as a co-adjuvant in hypoxic tumors and have the potential to improve cancer immunotherapies.

Hyaluronic Acid-Based Shape-Memory Cryogel Scaffolds for Focal Cartilage Defect Repair.

Traumatic joint injuries can result in significant cartilage defects, which can greatly increase the risk of osteoarthritis development. Due to the limited self-healing capacity of avascular cartilage, tissue engineering approaches are required for filling defects and promoting cartilage regeneration. Current approaches utilize invasive surgical procedures for extraction and implantation of autologous chondrocytes; therefore, injectable biomaterials have gained interest to minimize the risk of infection as well as patient pain and discomfort. In this study, we engineered biomimetic, hyaluronic acid (HA)-based cryogel scaffolds that possess shape-memory properties as they contract and regain their shape after syringe injection to noninvasively fill cartilage defects. The cryogels, fabricated with HA and glycidyl methacrylate at -20°C, resulted in an elastic, macroporous, and highly interconnected network that provided a conducive microenvironment for chondrocytes to remain viable and metabolically active after injection through a syringe needle. Chondrocytes seeded within cryogels and cultured for 15 days exhibited enhanced cell proliferation, metabolism, and production of cartilage extracellular matrix glycosaminoglycans compared with HA-based hydrogels. Furthermore, immunohistochemical staining revealed production of collagen type II from chondrocyte-seeded cryogels, indicating the maintenance of cell phenotype. These results demonstrate the potential of chondrocyte-seeded, HA-based, injectable cryogel scaffolds to promote regeneration of cartilage tissue for nonsurgically invasive defect repair. Impact statement Hyaluronic acid-based shape-memory cryogels provide a conducive microenvironment for chondrocyte adhesion, proliferation, and matrix biosynthesis for use in repair of cartilage defects. Due to their sponge-like elastic properties, cryogels can fully recover their original shape back after injection while not impacting metabolism or viability of encapsulated cells. Clinically, they provide an opportunity for filling focal cartilage defects by using a single, minimally invasive injection of a cell encapsulating biocompatible three-dimensional scaffold that can return to its original structure to fit the defect geometry and enable matrix regeneration.

Needle-injectable microcomposite cryogel scaffolds with antimicrobial properties.

Porous three-dimensional hydrogel scaffolds have an exquisite ability to promote tissue repair. However, because of their high water content and invasive nature during surgical implantation, hydrogels are at an increased risk of bacterial infection. Recently, we have developed elastic biomimetic cryogels, an advanced type of polymeric hydrogel, that are syringe-deliverable through hypodermic needles. These needle-injectable cryogels have unique properties, including large and interconnected pores, mechanical robustness, and shape-memory. Like hydrogels, cryogels are also susceptible to colonization by microbial pathogens. To that end, our minimally invasive cryogels have been engineered to address this challenge. Specifically, we hybridized the cryogels with calcium peroxide microparticles to controllably produce bactericidal hydrogen peroxide. Our novel microcomposite cryogels exhibit antimicrobial properties and inhibit antibiotic-resistant bacteria (MRSA and Pseudomonas aeruginosa), the most common cause of biomaterial implant failure in modern medicine. Moreover, the cryogels showed negligible cytotoxicity toward murine fibroblasts and prevented activation of primary bone marrow-derived dendritic cells ex vivo. Finally, in vivo data suggested tissue integration, biodegradation, and minimal host inflammatory responses when the antimicrobial cryogels, even when purposely contaminated with bacteria, were subcutaneously injected in mice. Collectively, these needle-injectable microcomposite cryogels show great promise for biomedical applications, especially in tissue engineering and regenerative medicine.

Injectable Cryogels for Biomedical Applications.

To prevent postoperative complications, there has been a substantial interest in designing syringe-injectable hydrogels. To date, cryogels remain the only viable option for preformed and large-scale hydrogels to be delivered through a conventional needle-syringe injection. Cryogels, a type of hydrogel with exceptional features, are fabricated at subzero temperatures. This process typically results in a biomaterial with a unique macroporous network, shape-memory properties, and exceptional flexibility allowing syringe injectability. These advanced biomaterials have been used for a number of biomedical applications, including tissue engineering, drug delivery, and more recently, immunotherapy. This review summarizes the recent progress on the design of injectable cryogels, their current limitations, and strategies to further improve their properties for translatability into the clinic.

A bioinspired, photostable UV-filter that protects mammalian cells against UV-induced cellular damage.

While commercially available suncare products are effective at absorbing ultraviolet (UV)-light, recent studies indicate systemic toxicities associated with many traditional chemical and physical UV-filters. We demonstrate the application of xanthommatin, a biochrome present in arthropods and cephalopods, as an alternative chemical UV-filter that is cytocompatible while maintaining its photostability and photoprotective properties.

Lipidic Aminoglycoside Derivatives: A New Class of Immunomodulators Inducing a Potent Innate Immune Stimulation.

Development of simple and fully characterized immunomodulatory molecules is an active area of research to enhance current immunotherapies. Monophosphoryl lipid A (MPL), a nontoxic lipidic derivative from bacteria, is the first and currently only adjuvant approved in humans. However, its capacity to induce a potent response against weak immunogenic tumoral-associated antigens remains limited. Herein, a new generation of lipidic immunomodulators to conduct a structure-activity relationship study to determine the minimal structural elements conferring immunomodulatory properties is introduced. Two lead molecules characterized by a short succinyl linker between two oleyl chains and a polar headgroup consisting of either naturally occurring tobramycin (DOST) or kanamycin (DOSK) are identified. These two lipoaminoglycosides self-assemble in very small vesicles. In a wide variety of cells including 3D human cell culture, DOST and DOSK induce the upregulation of proinflammatory cytokines and interferon-inducible proteins in a dose and time-dependent manner via a caveolae-dependent proinflammatory mechanism and phosphatidylinositol phospholipase C activation. Furthermore, after intratumoral administration, these lipoaminoglycosides induce an efficient immune response leading to significant antitumor activity in a mouse breast cancer model. Altogether, these findings indicate that DOST and DOSK are two groundbreaking synthetic lipid immunostimulators that can be used as adjuvants to enhance current immunotherapeutic treatments.

Autoclavable and Injectable Cryogels for Biomedical Applications.

Prior to any clinical application, terminal sterilization of biomaterials is a critical process imposed by the Food and Drug Administration. Of all the methods available for sterilization, high-pressure steam sterilization such as autoclaving is the most widely used. While autoclave sterilization minimizes pathogen contamination, it can dramatically impact both structural and biological properties of biomaterials. It has recently been reported that injectable cryogels with shape memory properties hold great promises as 3D macroporous biomimetic scaffolds for biomedical applications including tissue engineering. In this study, the impact of autoclave sterilization on properties of a series of cryogels is measured. Unlike conventional hydrogels, cryogels made of natural polymers demonstrate a strong resilience to autoclave sterilization. This process does not alter either their macrostructural or unique physical properties including syringe injectability. The scaffolds' bioactive sites are preserved and autoclaved cryogels retain their excellent cytological compatibility post-autoclaving. Furthermore, autoclaved cryogels do not trigger a notable activation of primary murine bone marrow-derived dendritic cells suggesting a minimal risk for biomaterial-induced inflammation, which is further confirmed by an in vivo histologic analysis. In summary, these results further demonstrate the huge potential of cryogels in the biomedical field and their capacity to be translated into clinical applications.

Latest Progress in Electrospun Nanofibers for Wound Healing Applications.

Electrospinning is a versatile technique used to create native tissue-like fibrous scaffolds. Recently, it has gained a large amount of attention for generation of bioactive dressing materials suitable for treatment of both chronic and acute wounds. In this Review, we focus on the latest advances made in the application of electrospun scaffolds for bioactive wound healing. We first provide a brief overview of the wound healing process and electrospinning approaches. We then discuss fabrication of scaffolds made from natural and synthetic polymers via electrospinning for effective wound treatment and management. Natural polymers used for wound healing included in our Review cover protein based polymers such as collagen, gelatin, and silk and polysaccharide based polymers such as chitosan, hyaluronic acid, and alginate. In addition, we discuss aliphatic polyesters, super hydrophilic polymers, and polyurethanes as some of the most commonly used synthetic polymers for wound healing and wound dressing applications. Next, we review multifunctional and "smart" scaffolds developed by electrospinning based approaches. We place an emphasis on how flexibility of the electrospinning process enables production of advanced scaffolds such as core-shell fibrous scaffolds, multilayer scaffolds, and surface modified scaffolds. Taken together, it is clear that electrospinning is an emerging technology that provides a unique opportunity for engineering more effective wound dressing, management, and care products.