Nanoengineered Osteoinductive and Elastomeric Scaffolds for Bone Tissue Engineering

Synthesis and fabrication of porous and elastomeric nanocomposite scaffolds from biodegradable poly(glycerol sebacate) (PGS) and osteoinductive nanosilicates is reported. Nanosilicates are mineral-based two-dimensional (2D) nanomaterials with high surface area which reinforced PGS network. The addition of nanosilicates to PGS resulted in mechanically stiff and elastomeric nanocomposites. The degradation rate and mechanical stiffness of nanocomposite network could be modulated by addition of nanosilicates. Nanocomposite scaffolds supported cell adhesion, spreading, and proliferation and promoted osteogenic differentiation of preosteoblasts. The addition of nanosilicates to PGS scaffolds increased alkaline phosphatase (ALP) activity and production of matrix mineralization. In vivo studies demonstrated biocompatibility and biodegradability of nanocomposite scaffolds. Overall, the combination of elasticity and tailorable stiffness, tunable degradation profiles, and the osteoinductive capability of the scaffolds offer a promising approach for bone tissue engineering

Alleviating Cell Lysate-Induced Inhibition to Enable RT-PCR from Single Cells in Picoliter-Volume Double Emulsion Droplets

Microfluidic droplet assays enable single-cell polymerase chain reaction (PCR) and sequencing analyses at unprecedented scales, with most methods encapsulating cells within nanoliter-sized single emulsion droplets (water-in-oil). Encapsulating cells within picoliter double emulsion (DE) (water-in-oil-in-water) allows sorting droplets with commercially available fluorescence-activated cell sorter (FACS) machines, making it possible to isolate single cells based on phenotypes of interest for downstream analyses. However, sorting DE droplets with standard cytometers requires small droplets that can pass FACS nozzles. This poses challenges for molecular biology, as prior reports suggest that reverse transcription (RT) and PCR amplification cannot proceed efficiently at volumes below 1 nL due to cell lysate-induced inhibition. To overcome this limitation, we used a plate-based RT-PCR assay designed to mimic reactions in picoliter droplets to systematically quantify and ameliorate the inhibition. We find that RT-PCR is blocked by lysate-induced cleavage of nucleic acid probes and primers, which can be efficiently alleviated through heat lysis. We further show that the magnitude of inhibition depends on the cell type, but that RT-PCR can proceed in low-picoscale reaction volumes for most mouse and human cell lines tested. Finally, we demonstrate one-step RT-PCR from single cells in 20 pL DE droplets with fluorescence quantifiable via FACS. These results open up new avenues for improving picoscale droplet RT-PCR reactions and expanding microfluidic droplet-based single-cell analysis technologies

Dapagliflozin effects on lung fluid volumes in patients with heart failure and reduced ejection fraction: Results from the DEFINE‐HF trial

Sodium‐glucose cotransporter‐2 (SGLT2) inhibitors have been shown to reduce the risk of cardiovascular death or worsening heart failure (HF), and improve symptom burden, physical function and quality of life in patients with HF and reduced ejection fraction. The mechanisms of the HF benefits of SGLT2 inhibitors, however, remain unclear. In this substudy of the DEFINE‐HF trial, patients randomized to dapagliflozin or placebo had lung fluid volumes (LFVs) measured by remote dieletric sensing at baseline and after 12 weeks of therapy. A significantly greater proportion of dapagliflozin‐treated patients (as compared with placebo) experienced improvement in LFVs and fewer dapagliflozin‐treated patients had no change or deterioration in LFVs after 12 weeks of treatment. To our knowledge, this is the first study to suggest a direct effect of dapagliflozin (or any SGLT2 inhibitor) on more effective “decongestion”, contributing in a meaningful way to the ongoing debate regarding the mechanisms of SGLT2 inhibitor HF benefits.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/167750/1/dom14352.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/167750/2/dom14352_am.pd