Bioengineered Cell Niche for Skeletal Muscle Regeneration

Skeletal muscles can self-repair minor strains, lacerations, and contusions; however, in cases of volumetric muscle lossand muscle degenerative diseases, tissue fails to regenerate. Current cell-based therapies, such as myoblast transplantation, have significant drawbacks of low survival rates and engraftment efficacy, mainly due to the absence of supportive cell microenvironment. Scaffolds that mimic the natural cell microenvironment provide a robust platform to support cell adhesion, migration, proliferation, and differentiation. Electrospinning is a versatile technology platform used for fabricating the fiber scaffold that mimics the extracellular matrix. Thus, we aim to reconstitute the cell microenvironment through development of aligned fiber scaffolds by electrospinning as oriented muscle fibers create natural microenvironment of myogenic cells. In particular, aligned fiber scaffolds will be optimized in term of mechanical properties and fiber diameters as fiber curvature and mechanical stiffness provide significant physical cues for myogenic cell behaviors. Here, we fabricated and characterized electrospun polyester fiber scaffolds with different diameters from micro-scale to nano-scale. The mechanical properties of the fabricated nanofibers were found to be in the range of contractile muscles as evidenced from atomic force microscopy measurements. With these scaffolds, C2C12 myoblasts were seeded and analyzed for the initial attachment. It was shown that aligned fibers with varying diameters resulted in different responses in cell attachment, indicating the role of cell topography sensing in cell-biomaterial interactions. Current ongoing studies focus on long-term in vitro culture of scaffolds in a custom-made muscle bioreactor emulating the contraction/relaxation of skeletal muscle tissue

Harnessing Notch Signaling for Biomaterial Scaffold-based Bone Regeneration

Bone fracture has recently become prevalent, especially with an increasingly aging population. Current bone grafts procedures, including autografts and allografts, are hindered by multiple factors, such as limited supplies and inconsistent bone healing. Scaffold-based bone tissue engineering emerges as a prospective strategy to aid in bone regeneration through delivery of growth factors such as bone morphogenic proteins (BMPs). However, the use of BMPs suffers from several drawbacks such as protein instability and immunogenicity. Therefore, there exists a great need for the development of novel therapies to promote bone healing. Notch signaling, a pathway critical for cell-fate determination has been shown to regulate osteogenesis, which suggests the potential of targeting Notch to enhance bone repair. The long-term goal of this work is to develop biomaterial-based regenerative technologies to induce bone regeneration by fine-tuning Notch signaling. In this study, a three-dimensional (3D) porous scaffold system was fabricated from biodegradable poly(lactide-co-glycolide) (PLGA) to mimic structural and mechanical properties of native bone using a microsphere sintering technology. In vitro studies were conducted to evaluate the effects of notch inhibition via a γ-secretase inhibitor DAPT on osteoblast responses. When the DAPT was added during the 2D culture on tissue culture polystyrene (TCPS), the osteoblast mineral deposition was significantly enhanced. Intriguingly, the enhancement was more pronounced on the 3D PLGA scaffolds, which may be attributed to the increased cell-cell contact in the 3D culture environment. Current efforts are focused on scaffold-based modulation of Notch signaling with both quantitative and temporal precision for enhanced osteogenesis

Peripheral Neuropathy and Hindlimb Paralysis in a Mouse Model of Adipocyte-Specific Knockout of Lkb1

Brown adipose tissues (BAT) burn lipids to generate heat through uncoupled respiration, thus representing a powerful target to counteract lipid accumulation and obesity. The tumor suppressor liver kinase b1 (Lkb1) is a key regulator of cellular energy metabolism; and adipocyte-specific knockout of Lkb1 (Ad-Lkb1 KO) leads to the expansion of BAT, improvements in systemic metabolism and resistance to obesity in young mice. Here we report the unexpected finding that the Ad-Lkb1 KO mice develop hindlimb paralysis at mid-age. Gene expression analyses indicate that Lkb1 KO upregulates the expression of inflammatory cytokines in interscapular BAT and epineurial brown adipocytes surrounding the sciatic nerve. This is followed by peripheral neuropathy characterized by infiltration of macrophages into the sciatic nerve, axon degeneration, reduced nerve conductance, and hindlimb paralysis. Mechanistically, Lkb1 KO reduces AMPK phosphorylation and amplifies mammalian target-of-rapamycin (mTOR)-dependent inflammatory signaling specifically in BAT but not WAT. Importantly, pharmacological or genetic inhibition of mTOR ameliorates inflammation and prevents paralysis. These results demonstrate that BAT inflammation is linked to peripheral neuropathy