Matrix glycation modulated cell behavior to induce inflammation equivalent to that produced by incubation with P. gingivalis LPS. Periodontal inflammation also led to matrix glycation, thus demonstrating a definite interaction between diabetes and periodontitis.

Regeneration of human heart muscle after injury is extremely limited and an unmet clinical need.Despite extensive research, there are no methods for the reproducible generation of clinical quality stem-cell-derived cardiovascular progenitors (CVPs). Here, we identified laminin-221 (LN-221) as the main cardiac laminin, which was produced here as human recombinant protein, and showed that LN-221 promotes differentiation of pluripotent hESCs towards cardiomyocyte lineage and downregulates genes associated with pluripotency and teratoma development. We developed a chemically defined, xeno-free laminin-based cardiomyocyte differentiation protocol to reproducibly generate CVPs that form human muscle in vivo. We assessed the reproducibility of the differentiation protocol using timecourse bulk RNA sequencing developed from two different hESC lines. Single-cell RNA sequencing of CVPs derived from two hESC lines further showed high reproducibility and identified three main progenitor subpopulations. These CVPs were transplanted into myocardial infarction mice, where heart function was measured by echocardiogram and human heart muscle bundle formation was identified histologically. This method may provide clinical quality cells for use in regenerative cardiology.

Background: Ischemic heart disease is a huge global burden where patients often have irreversibly damaged heart muscle. State-of-the-art technology using stem cell-derived products for cellular therapy could potentially replace damaged heart muscle for regenerative cardiology. Methods and Results: Pluripotent human embryonic stem cells (hESCs) were differentiated on a laminin LN521+221 matrix to cardiovascular progenitors (CVPs). Global transcriptome analyses at multiple time points by single-cell RNA-sequencing demonstrated high reproducibility (R2 > 0.95) between two hESCs lines. We identified several CVP signature genes as quality batch control parameters which are highly specific to our CVPs as compared to canonical cardiac progenitor genes. A total of 200 million CVPs were injected into the infarcted region caused by permanent ligation of the coronary arteries of 10 immunosuppressed pigs and maintained for 4- and 12-weeks post transplantation. The transplanted cells engrafted and proliferated in the infarcted area as indicated by IVIS imaging, histology staining and spatial transcriptomic analysis. Spatial transcriptomic analysis at 1 week following transplantation showed that the infarcted region expressed human genes in the same area as immunohistology sections. Heart function was analyzed by magnetic resonance imaging (MRI) and computerized tomography (CT). Functional studies revealed overall improvement in left ventricular ejection fraction by 21.35 +/- 3.3 %, which was accompanied by significant improvements in ventricular wall thickness and wall motion, as well as a reduction in infarction size after CVP transplantation as compared to medium control pigs (P < 0.05). Immunohistology analysis revealed maturation of the CVPs to cardiomyocytes (CMs) where the human grafts aligned with host tissue forming end-to-end connections typical for heart muscle. Electrophysiology analyses revealed electric continuity between injected and host tissue CMs. Episodes of ventricular tachyarrhythmia (VT) over a period of 25 days developed in four pigs, one pig had persistent VT, while the rest remained in normal sinus rhythm. All ten pigs survived the experiment without any VT-related death. Conclusions: We report a highly reproducible, chemically defined and fully humanized differentiation method of hESCs for the generation of potent CVPs. This method may pave the way for lasting stem cell therapy of myocardial infarction (MI) in humans.