Biology subjects

T. Annamalai, R.

Publications and source records attributed to T. Annamalai, R..

6 recordsLinked to original sources

bioRxiv2024

Substrate Curvature Influences Cytoskeletal Rearrangement and Modulates Macrophage Phenotype

Inflammation serves as a critical defense mechanism against pathogens and tissue damage but can lead to chronic diseases, such as cardiovascular disease and diabetes, when dysregulated. Macrophages play a pivotal role in orchestrating inflammatory responses, transitioning from pro-inflammatory M1 to anti-inflammatory M2 phenotypes to resolve inflammation and promote tissue repair. Current approaches to modulate macrophage phenotype predominantly rely on biochemical cues, which may induce systemic side effects. Given the mechanosensitivity of macrophages, this study investigates biophysical cues, specifically substrate curvature, as a localized strategy to regulate macrophage phenotype and minimize systemic repercussions. We hypothesized that substrate curvature influences macrophage immunophenotype by modulating F-actin polymerization. To test this hypothesis, we fabricated spherical microgels with tunable curvatures and characterized their biophysical properties. Our findings indicate that macrophages adhere to microgel surfaces irrespective of curvature, but the curvature significantly alters F-actin dynamics. Furthermore, manipulating cytoskeletal dynamics via selective actin inhibition partially reversed curvature-induced changes in macrophage phenotype. These results underscore the pivotal role of substrate curvature in modulating macrophage behavior and immunophenotype. Overall, our study demonstrates that substrate curvature significantly influences macrophage cytoskeletal dynamics and resulting immunophenotype. This simple approach can be utilized as a localized immunomodulatory treatment for inflammatory diseases.

bioengineering↗

bioRxiv2023

Carbonized Tetracycline: a new class of nanomaterial with tuneable antioxidant, reduced cytotoxicity, immunomodulatory, and osteogenic properties

Tetracycline (TET), a broad-spectrum antibiotic, also possesses different non-antibiotic activities such as inhibition of metalloproteinase (MMP), anti-inflammatory, antioxidant, high bone affinity, etc. However, the comparatively low efficacy of these non-antibiotic properties along with adverse effects such as hyperpigmentation, phototoxicity, long-term skeletal retention, etc. have not helped their broad utilization similar to their use as an antibiotic. In a unique attempt to improve the non-antibiotic properties while reducing the adverse effects, we converted the TET to nano-carbons through partial carbonization. After sorting out two water-dispersible C-TETs (C-TETHT - hydrothermal and C-TETHP - hot plate) based on their improved antioxidant activity, they have been characterized through a host of analytical techniques that showed distinct differences in morphology, size, shape, and surface functionality. Excitingly, the C-TETHT and C-TETHP have shown differential biological activity in a dosage and time-dependent manner in terms of cytotoxicity, immunomodulation, and osteogenic activity that was found to be associated with their carbonized parameters. Overall, the carbonized nano-drugs, C-TETHT and C-TETHP have presented substantial early promises on their non-antibiotic properties that could be further explored to develop into some effective therapeutics.

bioengineering↗

bioRxiv2023

Magnetic nanocomplexes coupled with an external magnetic field modulate macrophage phenotype: a non-invasive strategy for bone regeneration.

Chronic inflammation is a major cause for the pathogenesis of musculoskeletal diseases such as fragility fracture, and nonunion. Studies have shown that modulating the immune phenotype of macrophages from proinflammatory to prohealing mode can heal recalcitrant bone defects. Current therapeutic strategies predominantly apply biochemical cues, which often lack target specificity and controlling their release kinetics in vivo is challenging spatially and temporally. We show a magnetic iron-oxide nanocomplexes (MNC)-based strategy to resolve chronic inflammation in the context of promoting fracture healing. MNC internalized pro-inflammatory macrophages, when coupled with an external magnetic field, exert an intracellular magnetic force on the cytoskeleton, which promotes a prohealing phenotype switch. Mechanistically, the intracellular magnetic force perturbs actin polymerization, thereby significantly reducing nuclear to cytoplasm redistribution of MRTF-A and HDAC3, major drivers of inflammatory and osteogenic gene expressions. This significantly reduces Nos2 gene expression and subsequently downregulates the inflammatory response, as confirmed by quantitative PCR analysis. These findings are a proof of concept to develop MNC-based resolution-centric therapeutic intervention to direct macrophage phenotype and function towards healing and can be translated either to supplement or replace the currently used anti-inflammatory therapies for fracture healing.

bioengineering↗

bioRxiv2022

Biomechanical Stimulation of Muscles Influences Bone Phenotype by Modulating Myokine Secretion

Diabetes is a chronic metabolic disorder that affects 422 million people worldwide and can lead to diabetic myopathy and bone diseases. The etiology of musculoskeletal complications in diabetes and the interplay between the muscular and osseous systems are poorly understood. Exercise training promises to prevent diabetic myopathy and diabetic bone disease and offer protective effects on muscle and bone. Although the muscle-bone interaction is largely biomechanical, the muscle secretome, specifically the myokines, has significant implications for bone biology. Here, we have developed an in vitro model to elucidate the effects of mechanical strain on myokine secretion and its impact on bone metabolism decoupled from physical stimuli. We developed modular bone constructs using crosslinked gelatin hydrogels which facilitated osteogenic differentiation of osteoprogenitor cells. Then muscle constructs were made from fibrin hydrogel, which enabled myoblast differentiation and formed mature myotubes. We investigated the myokine expression by the muscle constructs under strain regimens replicating endurance (END) and high-intensity interval training (HIIT) in hyperglycemic conditions. In monocultures, both regimens induced higher expression of Il15 and Igf1, while END supported more myoblasts differentiation and myotube maturation than HIIT. When cocultured with bone constructs, the HIIT regimen increased Glut4 expression in muscle contructs that END supporting higher glucose uptake. Likewise, the muscle constructs under the HIIT regimen promoted a healthier and matured bone phenotype than END. Interestingly, under static conditions, myostatin (Mstn) expression was significantly downregulated in muscle constructs cocultured with bone constructs compared to monocultures. Our in vivo analysis of the role of myostatin on bone structure and function also showed that myostatin knockout (GDF8-/-) enhanced muscle mass and moderately influenced bone phenotype in adult mice. Together, our in vitro coculture system allowed orthogonal manipulation of mechanical strain on muscle constructs while facilitating biochemical crosstalk between bone and muscle constructs. Such systems can provide an individualized microenvironment and allow decoupled biomechanical manipulation, which is unachievable using traditional models. In the long-term, these in-vitro systems will help identify molecular targets and develop engineered therapies for diabetic bone disease.

bioengineering↗

bioRxiv2021

Injectable Nanoporous Microgels Generate Vascularized Constructs and Support Bone Regeneration in Critical-sized Defects

Large and aberrant bone fractures require ossification and concomitant vascularization for proper healing. Evidence indicates that osteogenesis and vessel growth are coupled in bone fractures. Although the synergistic role of endothelial cells has been recognized, vascularizing large bone grafts remains a challenge and has apprehended the clinical translation of engineered bone constructs. Here, we describe a facile method to fabricate vascularized constructs using chitosan and gelatin-based microgels that promote osteogenesis of human mesenchymal stromal cells (MSC) while supporting endothelial sprouting and network formation. The microgels are enzymatically degradable and had a high hydration rate with a volume swelling ratio of ~560% and a polymer density of ~430 mg/cm3, which is comparable to that of native skeletal tissues. AFM indentation of the surface showed an average Youngs modulus of 189 kPa, falling in a range that is conducive to both osteogenesis and vasculogenesis. The osteogenic microgel containing chitosan, gelatin, and hydroxyapatite, mimicking the bone matrix, supported robust attachment, proliferation, and differentiation of MSC. On the other hand, the vasculogenic microgels containing only gelatin, enriched endothelial phenotype and enabled vascular networks formation when embedded in 3D matrices. Combining the two types of microgels created a hybrid construct that sustained the functions of both osteogenic and vasculogenic microgels and enhanced one another. Using a murine model, we also show that the osteogenic microgels regenerate bone in a critical-sized defect with >95% defect closure by week 12. These multifunctional microgels can be administered minimally invasively and can conformally fill large bone defects. This work lays the foundation to establish principles of designing multiphasic scaffolds with tissue-specific biophysical and biochemical properties for regenerating vascularized and interfacial tissues.

bioengineering↗

bioRxiv2019

Bioresponsive Microspheres for On-demand Delivery of Anti-inflammatory Cytokines for Osteoarthritis

Despite innovations in surgical interventions, treatment of cartilage injury in osteoarthritic joints remains a challenge due to concomitant inflammation. Obstructing a single dominant inflammatory cytokine have shown remarkable clinical benefits in rheumatoid arthritis, and similar strategies are being suggested to target inflammatory pathways in osteoarthritis (OA). Here we describe the utility of gelatin microspheres that are responsive to arthritic flares, resulting in on-demand, and spatiotemporally controlled release of anti-inflammatory cytokines for cartilage preservation and repair. These microspheres had net negative charge potential to sequester cationic anti-inflammatory cytokines, and the magnitude of the negative charge potential increased with increase in crosslinking density. The enzymatic degradation of the microcarriers was concentration dependent. Release of anti-inflammatory cytokines from the loaded microspheres was directly correlated with the degradation of the gelatin matrix. Exposure of the IL-4 and IL-13 loaded microspheres reduced the inflammation of chondrocytes up to 80%. Hence, the delivery of these microspheres in an osteoarthritic joint can attenuate the stimulation of chondrocytes to secrete catabolic factors including proteinases and nitric oxide. The microsphere format also allows for minimally invasive delivery and is less susceptible to mechanically-induced drug release and are conformant to the intra-articular space. Consequently, bioresponsive microspheres are an effective tool for OA prevention and treatment.

bioengineering↗