Biology subjects

Contag, C. H.

Publications and source records attributed to Contag, C. H..

8 recordsLinked to original sources

bioRxiv2022

Polylactide Degradation Activates Immune Cells by Metabolic Reprogramming

Polylactide (PLA) is the most widely utilized biopolymer in medicine. However, chronic inflammation and excessive fibrosis resulting from its degradation remain significant obstacles to extended clinical use. Immune cell activation has been correlated to the acidity of breakdown products, yet methods to neutralize the pH have not significantly reduced adverse responses. Using a bioenergetic model, we observed delayed cellular changes that were not apparent in the short-term. Amorphous and semi-crystalline PLA degradation products, including monomeric L-lactic acid, mechanistically remodel metabolism in cells leading to a reactive immune microenvironment characterized by elevated proinflammatory cytokines. Selective inhibition of metabolic reprogramming and altered bioenergetics both reduce these undesirable high cytokine levels and stimulate anti-inflammatory signals. Our results present a new biocompatibility paradigm by identifying metabolism as a target for immunomodulation to increase tolerance to biomaterials, ensuring safe clinical application of PLA-based implants for soft- and hard-tissue regeneration, and advancing nanomedicine and drug delivery.

bioengineering↗

bioRxiv2022

Magnetothermal control of temperature-sensitive repressors in superparamagnetic iron nanoparticle-coated Bacillus subtilis

Superparamagnetic iron oxide nanoparticles (SPIONs) are used as contrast agents in magnetic resonance imaging (MRI) and magnetic particle imaging (MPI) and resulting images can be used to guide magnetothermal heating. Alternating magnetic fields (AMF) cause local temperature increases in regions with SPIONs, and we investigated the ability of magnetic hyperthermia to regulate temperature-sensitive repressors (TSRs) of bacterial transcription. The TSR, TlpA39, was derived from a Gram-negative bacterium, and used here for thermal control of reporter gene expression in Gram-positive, Bacillus subtilis. In vitro heating of B. subtilis with TlpA39 controlling bacterial luciferase expression, resulted in a 14.6-fold (12-hour; h) and 1.8-fold (1-h) increase in reporter transcripts with a 9.0-fold (12-h) and 11.1-fold (1-h) increase in bioluminescence. To develop magnetothermal control, B. subtilis cells were coated with three SPION variations. Electron microscopy coupled with energy dispersive X-ray spectroscopy revealed an external association with, and retention of, SPIONs on B. subtilis. Furthermore, using long duration AMF we demonstrated magnetothermal induction of the TSRs in SPION-coated B. subtilis with a maximum of 4.6-fold increases in bioluminescence. After intramuscular injections of SPION-coated B. subtilis, histology revealed that SPIONs remained in the same locations as the bacteria. For in vivo studies, 1-h of AMF is the maximum exposure due to anesthesia constraints. Both in vitro and in vivo, there was no change in bioluminescence after 1-h of AMF treatment. Pairing TSRs with magnetothermal energy using SPIONs for localized heating with AMF can lead to transcriptional control that expands options for targeted bacteriotherapies.

bioengineering↗

bioRxiv2022

Gla-domain mediated targeting of externalized phosphatidylserine for intracellular delivery

Phosphatidylserine (PS) is a negatively charged phospholipid normally localized to the inner leaflet of the plasma membrane of cells but is externalized onto the cell surface during apoptosis as well as in malignant and infected cells. Consequently, PS may comprise an important molecular target in diagnostics, imaging and targeted delivery of therapeutic agents. While an array of PS binding-molecules exist, their utility has been limited by their inability to recognize PS and internalize diagnostic or therapeutic payloads. We describe the generation, isolation, characterization, and utility of a PS binding motif comprised of a carboxylated glutamic acid (GLA) residue domain, that both recognizes cell surface-exposed PS and is internalized into these cells after binding. Internalization is independent of the traditional endosomal-lysosomal pathway, directly entering the cytosol of the target cell in a rapid and energy-independent fashion. We demonstrate that this PS recognition extends to extracellular vesicles and stem cells and that GLA-domain conjugated probes can be detected upon intravenous administration in animal models of infectious disease and cancer. GLA domain binding and internalization offers new opportunities for targeting specific cells for imaging and delivery of therapeutics.

cell biology↗

bioRxiv2022

Harnessing insect olfactory neural circuits for noninvasive detection of human cancer

There is overwhelming evidence that metabolic processes are altered in cancer cells and these changes are manifested in the volatile organic compound (VOC) composition of exhaled breath. Here, we take a novel approach of an insect olfactory neural circuit-based VOC sensor for cancer detection. We combined an in vivo antennae-attached insect brain with an electrophysiology platform and employed biological neural computation rules of antennal lobe circuitry for data analysis to achieve our goals. Our results demonstrate that three different human oral cancers can be robustly distinguished from each other and from a non-cancer oral cell line by analyzing individual cell culture VOC composition-evoked olfactory neural responses in the insect antennal lobe. By evaluating cancer vs. non-cancer VOC-evoked population neural responses, we show that olfactory neurons response-based classification of oral cancer is sensitive and reliable. Moreover, this brain-based cancer detection approach is very fast (detection time ~ 250 ms). We also demonstrate that this cancer detection technique is effective across changing chemical environments mimicking natural conditions. Our brain-based cancer detection system comprises a novel VOC sensing methodology that will spur the development of more forward engineering technologies for noninvasive detection of cancer.

bioengineering↗

bioRxiv2021

Magnetic particle imaging of magnetotactic bacteria as living contrast agents is improved by altering magnetosome structures

Iron nanoparticles used as imaging contrast agents can help differentiate between normal and diseased tissue, or track cell movement and localize pathologies. Magnetic particle imaging (MPI) is an imaging modality that uses the magnetic properties of iron nanoparticles to provide specific, quantitative and sensitive imaging data. MPI signals depend on the size, structure and composition of the nanoparticles; MPI-tailored nanoparticles have been developed by modifying these properties. Magnetotactic bacteria produce magnetosomes which mimic synthetic nanoparticles, and thus comprise a living contrast agent in which nanoparticle formation can be modified by mutating genes. Specifically, genes that encode proteins critical to magnetosome formation and regulation, such as mamJ which helps with filament turnover. Deletion of mamJ in Magnetospirillum gryphiswaldense, MSR-1 led to clustered magnetosomes instead of the typical linear chains. Here we examined the effects of this magnetosome structure and revealed improved MPI signal and resolution from clustered magnetosomes compared to linear chains. Bioluminescent MSR-1 with the mamJ deletion were injected intravenously into tumor-bearing and healthy mice and imaged using both in vivo bioluminescence imaging (BLI) and MPI. BLI revealed the location and viability of bacteria which was used to validate localization of MPI signals. BLI identified the viability of MSR-1 for 24 hours and MPI detected iron in the liver and in multiple tumors. Development of living contrast agents offers new opportunities for imaging and therapy by using multimodality imaging to track the location and viability of the therapy and the resulting biological effects.

bioengineering↗

bioRxiv2021

A New Bioink For Improved 3D Bioprinting Of Bone-Like Constructs

Bone tissue loss can occur due to disease, trauma or following surgery, in each case treatment involving the use of bone grafts or biomaterials is usually required. Recent development of three-dimensional (3D) bioprinting (3DBP) has enabled the printing of customized bone substitutes. Bioinks used for bone 3DBP employ various particulate phases such as ceramic and bioactive glass particles embedded in the bioink creating a composite. When composite bioinks are used for 3DBP based on extrusion, particles are heterogeneously distributed causing damage to cells due to stresses created during flow in the matrix of the composite. Therefore, the objective of this study was to develop cell-friendly osteopromotive bioink mitigating the risk of cell damage due to the flow of particles. Towards this end, we have linked organic and inorganic components, gelatin methacryloyl (GelMA) and Ag-doped bioactive glass (Ag-BaG), to produce a hybrid material, GelMA-Ag-BaG (GAB). The distribution of the elements present in the Ag-BaG in the resulting hybrid GAB structure was examined. Rheological properties of the resulting hydrogel and its printability, as well as the degree of swelling and degradation over time, were also evaluated. GAB was compared to GelMA alone and GelMA-Ag-BaG nanocomposites. Results showed the superiority of the hybrid GAB bioink in terms of homogenous distribution of the elements in the structure, rheological properties, printability, and degradation profiles. Accordingly, this new bioink represents a major advance for bone 3DBP.

bioengineering↗

bioRxiv2021

Tracking the fates of iron-labeled tumor cells in vivo using Magnetic Particle Imaging

The use of imaging to detect and monitor the movement and accumulation of cells in living subjects can provide significant insights that can improve our understanding of metastasis and guide therapeutic development. For cell tracking using Magnetic Resonance Imaging (MRI), cells are labeled with iron oxides and the effects of the iron on water provides contrast. However, due to low specificity and difficulties in quantification with MRI, other modalities and approaches need to be developed. Magnetic Particle Imaging (MPI) is an emerging imaging technique which directly detects magnetic iron, allowing for a specific, quantitative and sensitive readout. Here, we use MPI to image iron-labeled tumor cells longitudinally, from implantation and growth at a primary site to movement to distant anatomic sites. In vivo bioluminescent imaging (BLI) was used to localize tumor metastases and computed tomography (CT) allowed for correlation of these signals to anatomic locations. These three imaging modalities provide information on immune escape and metastasis of iron-labeled, and unlabeled, tumor cells, and the accumulation of cell-free iron contrast over time. We identified iron signals by MPI and tumor cells via BLI, and correlated these positive contrast images with CT scans to reveal the anatomic sites with cancer cells; histologic analysis confirmed the presence of iron-labeled tumor cells in the tissues, suggesting that the metastatic cells retained enough iron for MPI detection. The use of multi-modality cell tracking reveals the movement, accumulation and fates of labeled cells that will be helpful understanding cancer progression and guiding the development of targeted therapies.

biophysics↗

bioRxiv2021

Engineered endosymbionts capable of directing mammalian cell gene expression

Modular methods for directing mammalian gene expression would enable advances in tissue regeneration, enhance cell-based therapeutics and improve modulation of immune responses. To address this challenge, engineered endosymbionts (EES) that escape endosomal destruction, reside in the cytoplasm of mammalian cells, and secrete proteins that are transported to the nucleus to control host cell gene expression were developed. Microscopy confirmed that EES escape phagosomes, replicate within the cytoplasm, and can secrete reporter proteins into the cytoplasm that were then transported to the nucleus. Synthetic operons encoding the mammalian transcription factors, Stat-1 and Klf6 or Klf4 and Gata-3 were recombined into the EES genome. Using controlled induction, these EES were shown to direct gene expression in J774A.1 macrophage/monocyte cells and modulate the host cell fates. Expressing mammalian transcription factors from engineered intracellular bacteria as endosymbionts comprises a new tool for directing host cell gene expression for therapeutic and research purposes. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=194 SRC="FIGDIR/small/463266v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@2b5935org.highwire.dtl.DTLVardef@1edd8edorg.highwire.dtl.DTLVardef@3cb5b9org.highwire.dtl.DTLVardef@1be9b5f_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗