Monocyte adhesion to and transmigration through endothelium following cardiopulmonary bypass shearing is mediated by IL-8 signaling.

Introduction: The use of cardiopulmonary bypass (CPB) can induce sterile systemic inflammation that contributes to morbidity and mortality, especially in children. Patients have been found to have increased expression of cytokines and transmigration of leukocytes during and after CPB. Previous work has demonstrated that the supraphysiologic shear stresses existing during CPB are sufficient to induce proinflammatory behavior in non-adherent monocytes.

Engineered myeloid precursors differentiate into osteoclasts and resorb heterotopic ossification in mice.

Introduction: Heterotopic ossification (HO) occurs following orthopedic trauma, spinal cord injuries, brain trauma and limb amputations. Once symptomatic, HO causes pain, limited mobility and decreased quality of life. Current treatments are limited and have significant complications with high recurrence rates, underscoring the need for improved therapeutic interventions.

Differentiation and Characterization of Osteoclasts from Human Induced Pluripotent Stem Cells.

This protocol details the propagation and passaging of human iPSCs and their differentiation into osteoclasts. First, iPSCs are dissociated into a single-cell suspension for further use in embryoid body induction. Following mesodermal induction, embryoid bodies undergo hematopoietic differentiation, producing a floating hematopoietic cell population.

Grand Challenges at the Interface of Engineering and Medicine.

Over the past two decades Biomedical Engineering has emerged as a major discipline that bridges societal needs of human health care with the development of novel technologies. Every medical institution is now equipped at varying degrees of sophistication with the ability to monitor human health in both non-invasive and invasive modes. The multiple scales at which human physiology can be interrogated provide a profound perspective on health and disease.

Comparison of osteoclast differentiation protocols from human induced pluripotent stem cells of different tissue origins.

Background: Ever since their discovery, induced pluripotent stem cells (iPSCs) have been extensively differentiated into a large variety of cell types. However, a limited amount of work has been dedicated to differentiating iPSCs into osteoclasts. While several differentiation protocols have been published, it remains unclear which protocols or differentiation methods are preferable regarding the differentiation of osteoclasts.

Comparison of osteoclast differentiation protocols from human induced pluripotent stem cells of different tissue origins.

Background: Ever since their discovery, induced pluripotent stem cells (iPSCs) have been extensively differentiated into a large variety of cell types. However, a limited amount of work has been dedicated to differentiating iPSCs into osteoclasts. While several differentiation protocols have been published, it remains unclear which protocols or differentiation methods are preferrable regarding the differentiation of osteoclasts.

Dissecting specific Wnt components governing osteogenic differentiation potential by human periodontal ligament stem cells through interleukin-6.

Periodontal ligament stem cells (PDLSCs) play a significant role on periodontal tissue and alveolar bone homeostasis. During inflammation, interleukin (IL)-6 serves as one of key cytokine players controlling tissue reaction as well as alveolar bone tissue remodeling. It is believed that periodontal tissue inflammation causes periodontium degradation, especially alveolar bone.

Role of Runx2 in Calcific Aortic Valve Disease in Mouse Models.

Calcific aortic valve disease is common in the aging population and is characterized by the histological changes of the aortic valves including extracellular matrix remodeling, osteochondrogenic differentiation, and calcification. Combined, these changes lead to aortic sclerosis, aortic stenosis (AS), and eventually to heart failure. Runt-related transcription factor 2 (Runx2) is a transcription factor highly expressed in the calcified aortic valves.

Engineered osteoclasts resorb necrotic alveolar bone in anti-RANKL antibody-treated mice.

Medication-related osteonecrosis of the jaw (MRONJ) is a serious side effect of antiresorptive medications such as denosumab (humanized anti-RANKL antibody), yet its pathophysiology remains elusive. It has been posited that inhibition of osteoclastic bone resorption leads to the pathological sequelae of dead bone accumulation, impaired new bone formation, and poor wound healing in MRONJ, but this hypothesis has not been definitively tested. We previously engineered myeloid precursors with a conditional receptor activator of nuclear factor kappa-Β intracellular domain (iRANK cells), which differentiate into osteoclasts in response to a chemical inducer of dimerization (CID) independently of RANKL.

Adapter Protein RapGEF1 Is Required for ERK1/2 Signaling in Response to Elevated Phosphate in Vascular Smooth Muscle Cells.

The sodium-dependent phosphate transporter, SLC20A1, is required for elevated inorganic phosphate (Pi) induced vascular smooth muscle cell (VSMC) matrix mineralization and phenotype transdifferentiation. Recently, elevated Pi was shown to induce ERK1/2 phosphorylation through SLC20A1 by Pi uptake-independent functions in VSMCs, suggesting a cell signaling response to elevated Pi. Previous studies identified Rap1 guanine nucleotide exchange factor (RapGEF1) as an SLC20A1-interacting protein and RapGEF1 promotes ERK1/2 phosphorylation through Rap1 activation.

Slc20a1/Pit1 and Slc20a2/Pit2 are essential for normal skeletal myofiber function and survival.

Low blood phosphate (Pi) reduces muscle function in hypophosphatemic disorders. Which Pi transporters are required and whether hormonal changes due to hypophosphatemia contribute to muscle function is unknown. To address these questions we generated a series of conditional knockout mice lacking one or both house-keeping Pi transporters Pit1 and Pit2 in skeletal muscle (sm), using the postnatally expressed human skeletal actin-cre.

PiT-2, a type III sodium-dependent phosphate transporter, protects against vascular calcification in mice with chronic kidney disease fed a high-phosphate diet.

PiT-2, a type III sodium-dependent phosphate transporter, is a causative gene for the brain arteriolar calcification in people with familial basal ganglion calcification. Here we examined the effect of PiT-2 haploinsufficiency on vascular calcification in uremic mice using wild-type and global PiT-2 heterozygous knockout mice. PiT-2 haploinsufficiency enhanced the development of vascular calcification in mice with chronic kidney disease fed a high-phosphate diet.

Increased Calcific Aortic Valve Disease in response to a diabetogenic, procalcific diet in the LDLrApoB mouse model.

Objective: Calcific aortic valve disease (CAVD) is a major cause of aortic stenosis (AS) and cardiac insufficiency. Patients with type II diabetes mellitus (T2DM) are at heightened risk for CAVD, and their valves have greater calcification than nondiabetic valves. No drugs to prevent or treat CAVD exist, and animal models that might help identify therapeutic targets are sorely lacking.

Role of smooth muscle cells in vascular calcification: implications in atherosclerosis and arterial stiffness.

Vascular calcification is associated with a significant increase in all-cause mortality and atherosclerotic plaque rupture. Calcification has been determined to be an active process driven in part by vascular smooth muscle cell (VSMC) transdifferentiation within the vascular wall. Historically, VSMC phenotype switching has been viewed as binary, with the cells able to adopt a physiological contractile phenotype or an alternate 'synthetic' phenotype in response to injury.

Loss of PiT-2 results in abnormal bone development and decreased bone mineral density and length in mice.

Normal bone mineralization requires phosphate oversaturation in bone matrix vesicles, as well as normal regulation of phosphate metabolism via the interplay among bone, intestine, and kidney. In turn, derangement of phosphate metabolism greatly affects bone function and structure. The type III sodium-dependent phosphate transporters, PiT-1 and PiT-2, are believed to be important in tissue phosphate metabolism and physiological bone formation, but their requirement and molecular roles in bone remain poorly investigated.

Osteoclast precursors do not express CD68: results from CD68 promoter-driven RANK transgenic mice.

Macrophages and osteoclasts are thought to derive from CD68 lineage marker-positive common myeloid precursors. We used the CD68 promoter to drive an inducible receptor activator of NF-κB (iRANK) construct that selectively activates RANK signaling in myeloid cells in vivo. The cytoplasmic portion of RANK was fused to a mutant FK506 binding domain, which selectively binds the chemical inducer of dimerization AP20187 and initiates signaling.

Vascular calcification in CKD-MBD: Roles for phosphate, FGF23, and Klotho.

Vascular calcification (VC) is highly prevalent in aging, diabetes mellitus, and chronic kidney disease (CKD). VC is a strong predictor of cardiovascular morbidity and mortality in the CKD population. Complex pathological mechanisms are involved in the development of VC, including osteochondrogenic differentiation and apoptosis of vascular smooth muscle cells, instability and release of extracellular vesicles loaded calcium and phosphate, and elastin degradation.

Runx2 deletion in smooth muscle cells inhibits vascular osteochondrogenesis and calcification but not atherosclerotic lesion formation.

Aims: Vascular smooth muscle cells (SMCs) are major precursors contributing to osteochondrogenesis and calcification in atherosclerosis. Runt-related transcription factor-2 (Runx2) has been found essential for SMC differentiation to an osteochondrogenic phenotype and subsequent calcification in vitro. A recent study using a conditional targeting allele that produced a truncated Runx2 protein in SMCs of ApoE-/- mice showed reduced vascular calcification, likely occurring via reduction of receptor activator of nuclear factor-κB ligand (RANKL), macrophage infiltration, and atherosclerotic lesion formation.

Osteopontin protects against high phosphate-induced nephrocalcinosis and vascular calcification.

Pathologic calcification is a significant cause of increased morbidity and mortality in patients with chronic kidney disease. The precise mechanisms of ectopic calcification are not fully elucidated, but it is known to be caused by an imbalance of procalcific and anticalcific factors. In the chronic kidney disease population, an elevated phosphate burden is both highly prevalent and a known risk factor for ectopic calcification.

Slc20a2 deficiency results in fetal growth restriction and placental calcification associated with thickened basement membranes and novel CD13 and lamininα1 expressing cells.

The essential nutrient phosphorus must be taken up by the mammalian embryo during gestation. The mechanism(s) and key proteins responsible for maternal to fetal phosphate transport have not been identified. Established parameters for placental phosphate transport match those of the type III phosphate transporters, Slc20a1 and Slc20a2.

SLC20A2 Deficiency in Mice Leads to Elevated Phosphate Levels in Cerbrospinal Fluid and Glymphatic Pathway-Associated Arteriolar Calcification, and Recapitulates Human Idiopathic Basal Ganglia Calcification.

Idiopathic basal ganglia calcification is a brain calcification disorder that has been genetically linked to autosomal dominant mutations in the sodium-dependent phosphate co-transporter, SLC20A2. The mechanisms whereby deficiency of Slc20a2 leads to basal ganglion calcification are unknown. In the mouse brain, we found that Slc20a2 was expressed in tissues that produce and/or regulate cerebrospinal fluid, including choroid plexus, ependyma and arteriolar smooth muscle cells.

Runx2 Expression in Smooth Muscle Cells Is Required for Arterial Medial Calcification in Mice.

Arterial medial calcification (AMC) is a hallmark of aging, diabetes, and chronic kidney disease. Smooth muscle cell (SMC) transition to an osteogenic phenotype is a common feature of AMC, and is preceded by expression of runt-related transcription factor 2 (Runx2), a master regulator of bone development. Whether SMC-specific Runx2 expression is required for osteogenic phenotype change and AMC remains unknown.