A Bioinspired Astrocyte-Derived Coating Promotes the In Vitro Proliferation of Human Neural Stem Cells While Maintaining Their Stemness.

The repair of neuronal tissue is a challenging process due to the limited proliferative capacity of neurons. Neural stem cells (NSCs) can aid in the regeneration process of neural tissue due to their high proliferation potential and capacity to differentiate into neurons. The therapeutic potential of these cells can only be achieved if sufficient cells are obtained without losing their differentiation potential. Toward this end, an astrocyte-derived coating (HAc) was evaluated as a promising substrate to promote the proliferation of NSCs. Mass spectroscopy and scanning electron microscopy were used to characterize the HAc. The proliferation rate and the expression of stemness and differentiation markers in NSCs cultured on the HAc were evaluated and compared to the responses of these cells to commonly used coating materials including Poly-L-Ornithine (PLO), and a Human Induced Pluripotent Stem Cell (HiPSC)-based coating. The use of the HAc promotes the in vitro cell growth of NSCs. The expression of the stemness markers Sox2 and Nestin, and the differentiation marker DCX in the HAc group was akin to the expression of these markers in the controls. In summary, HAc supported the proliferation of NSCs while maintaining their stemness and neural differentiation potential.

Circulating miRNAs associated with bone mineral density in healthy adult baboons.

MicroRNAs (miRNAs) regulate gene expression post-transcriptionally and circulate in the blood, making them attractive biomarkers of disease state for tissues like bone that are challenging to interrogate directly. Here, we report on five miRNAs-miR-197-3p, miR-320a, miR-320b, miR-331-5p, and miR-423-5p-associated with bone mineral density (BMD) in 147 healthy adult baboons. These baboons ranged in age from 15 to 25 years (45-75 human equivalent years) and 65% were female with a broad range of BMD values including a minority of osteopenic animals. miRNAs were generated via RNA sequencing from buffy coats collected at necropsy and areal BMD (aBMD) measured postmortem via dual-energy X-ray absorptiometry (DXA) of the lumbar vertebrae. Differential expression analysis controlled for the underlying pedigree structure of these animals to account for genetic variation which may drive miRNA abundance and aBMD values. While many of these miRNAs have been associated with the risk of osteoporosis in humans, this finding is of interest because the cohort represents a model of normal aging and bone metabolism rather than a disease cohort. The replication of miRNA associations with osteoporosis or other bone metabolic disorders in animals with healthy aBMD suggests an overlap in normal variation and disease states. We suggest that these miRNAs are involved in the regulation of cellular proliferation, apoptosis, and protein composition in the extracellular matrix throughout life; and age-related dysregulation of these systems may lead to disease. These miRNAs may be early indicators of progression to disease in advance of clinically detectible osteoporosis.

Extracellular matrix derived from chondrocytes promotes rapid expansion of human primary chondrocytes in vitro with reduced dedifferentiation.

A significant expansion of autologous chondrocytes in vitro is required for cell-based cartilage repair. However, the in vitro expansion of chondrocytes under standard culture conditions inevitably leads to the dedifferentiation of chondrocytes and contributes to suboptimal clinical outcomes. To address this challenge, we focused our efforts on developing an improved in vitro expansion protocol, which shortens the expansion time with decreased dedifferentiation. It is known that the tissue microenvironment plays a critical role in regulating the cellular functions of resident cells and provides guidance in tissue-specific regeneration. We hypothesized that chondrocyte extracellular matrix (ECM) mimics a native microenvironment and that it may support chondrocyte expansion in vitro. To test this hypothesis, we prepared decellularized ECMs from allogeneic human articular chondrocytes (HAC) (AC-ECM) and bone marrow stromal cells (BM-ECM) and studied their effects on the in vitro expansion of primary HAC. The differential composition and physical properties of these two ECMs were revealed by mass spectrometry and atomic force microscopy. Compared with standard tissue culture polystyrene (TCP) or BM-ECM, HAC cultured on AC-ECM proliferated faster and maintained the highest ratio of COL2A1/COL1A1. Furthermore, a pellet culture study demonstrated that cells expanded on AC-ECM produced a more cartilage-like ECM than cells expanded on BM-ECM or TCP. This is the first report on modulating chondrocyte expansion and dedifferentiation using cell type-specific ECM and on identifying AC-ECM as a preferred substrate for in vitro expansion of HAC cell-based therapies. STATEMENT OF SIGNIFICANCE: To reduce the dedifferentiation of chondrocytes during in vitro expansion, cell type-specific extracellular matrix (ECM), which mimics a native microenvironment, was prepared from human articular chondrocytes (AC-ECM) or bone marrow stromal cells (BM-ECM). As demonstrated by mass spectrometry and atomic force microscopy, AC-ECM and BM-ECM have differential ECM compositions and physical characteristics. Human articular chondrocytes (HAC) expanded faster and maintained a better chondrocyte phenotype on AC-ECM than on BM-ECM or a standard culture surface. AC-ECM has potential to be developed for expanding HAC for cell-based therapies.