Facilitated recruitment of mesenchymal stromal cells by bone marrow concentrate and platelet rich plasma.

Biologics containing growth factors are frequently used to enhance healing after musculoskeletal injuries. One mechanism of action is thought to be though the ability of biologics to induce homing and migration of endogenous mesenchymal stromal cells (MSCs) to a target tissue. However, the ability of biologics to stimulate chemotaxis (directed migration of cells) and chemokinesis (increase rate of cell migration) of MSCs is unknown.The aim of this study was to directly compare the ability of biologics including platelet rich plasma (PRP) and bone marrow concentrate (BMC) to induce MSC migration. The hypothesis was that leukocyte-low platelet rich plasma (Llo PRP) would induce migration to a greater extent than leukocyte-high platelet rich plasma (Lhi PRP) or BMC.Bone marrow-derived MSCs were isolated from 8 horses. Migration of MSCs toward a biologic (BMC, Llo PRP, and Lhi PRP) or the positive control platelet derived growth factor (PDGF) was continuously traced and measured for 24hrs using time-lapse microscopy and a microfluidics device. Cell migration, chemotaxis and chemokinesis were determined by measurements of displacement, number of cells migrated, and cell flux.All biologics resulted in a significantly greater percentage of MSCs migrated compared to the positive control (PDGF). MSCs migrated further toward BMC compared to Llo PRP. Cell migration, measured as cell flux, was greater toward BMC and Lhi PRP than Llo PRP.The biologics BMC and Lhi PRP elicit greater chemotaxis and chemokinesis of MSCs than Llo PRP. However, all biologics recruited the same number of MSCs suggesting that differences in other regenerative effects, such as growth factor concentration, between biologics should be strongly considered when choosing a biologic for treatment of musculoskeletal injuries. The results of this study have the potential to reduce the need, risks, and costs associated with MSC culture and delivery

Facilitated recruitment of mesenchymal stromal cells by bone marrow concentrate and platelet rich plasma

<div><p>Background</p><p>Biologics containing growth factors are frequently used to enhance healing after musculoskeletal injuries. One mechanism of action is thought to be though the ability of biologics to induce homing and migration of endogenous mesenchymal stromal cells (MSCs) to a target tissue. However, the ability of biologics to stimulate chemotaxis (directed migration of cells) and chemokinesis (increase rate of cell migration) of MSCs is unknown.</p><p>Hypothesis/Purpose</p><p>The aim of this study was to directly compare the ability of biologics including platelet rich plasma (PRP) and bone marrow concentrate (BMC) to induce MSC migration. The hypothesis was that leukocyte-low platelet rich plasma (L^lo PRP) would induce migration to a greater extent than leukocyte-high platelet rich plasma (L^hi PRP) or BMC.</p><p>Methods</p><p>Bone marrow-derived MSCs were isolated from 8 horses. Migration of MSCs toward a biologic (BMC, L^lo PRP, and L^hi PRP) or the positive control platelet derived growth factor (PDGF) was continuously traced and measured for 24hrs using time-lapse microscopy and a microfluidics device. Cell migration, chemotaxis and chemokinesis were determined by measurements of displacement, number of cells migrated, and cell flux.</p><p>Results</p><p>All biologics resulted in a significantly greater percentage of MSCs migrated compared to the positive control (PDGF). MSCs migrated further toward BMC compared to L^lo PRP. Cell migration, measured as cell flux, was greater toward BMC and L^hi PRP than L^lo PRP.</p><p>Conclusion</p><p>The biologics BMC and L^hi PRP elicit greater chemotaxis and chemokinesis of MSCs than L^lo PRP. However, all biologics recruited the same number of MSCs suggesting that differences in other regenerative effects, such as growth factor concentration, between biologics should be strongly considered when choosing a biologic for treatment of musculoskeletal injuries. The results of this study have the potential to reduce the need, risks, and costs associated with MSC culture and delivery.</p></div

Cell flux toward a putative chemoattractant.

<p>PDGF, BMC, L^hi PRP resulted in significantly more cell flux than L^lo PRP (bars). PDGF, BMC, L^lo PRP, L^hi PRP resulted in significantly greater cell flux compared to the NC (asterisks). Data are represented as median with maximum and minimum values; n = 8. NC = neutral control, PDGF = platelet derived growth factor, BMA = bone marrow aspirate, BMC = bone marrow aspirate concentrate, L^lo PRP = leukocyte low platelet rich plasma, L^hi PRP = leukocyte high platelet rich plasma. Significance was determined by a Kruskal-Wallis followed by Wilcoxon multiple comparison post-hoc test. A p-value < 0.01 was considered significant.</p

Facilitated recruitment of mesenchymal stromal cells by bone marrow concentrate and platelet rich plasma - Fig 4

<p>Cell migration images within the microfluidics μ- slide chemotaxis device at (A) time = 0hrs, and (B) time = 24hrs. Cells show preferential migration toward leukocyte high platelet rich plasma (L^hi PRP) compared to the neutral control (NC).</p

The μ-slide chemotaxis device.

<p>The area between ports A and B is the observation area where MSCs are seeded. The trapezoid shaped area between ports A, B, C, and D is the reservoir well where putative chemoattractants are placed. The trapezoid shaped area between ports A, B, E, and F is the reservoir well where the neutral control (stem cell media with 10% fetal bovine serum) is placed.</p

Displacement of cells toward a putative chemoattractant.

<p>BMC and PDGF resulted in greater displacement in comparison to L^lo PRP (horizontal bars). PDGF resulted in greater displacement compared to the NC (asterisk). Data are represented as median with maximum and minimum values; n = 8. NC = neutral control, PDGF = platelet derived growth factor, BMA = bone marrow aspirate, BMC = bone marrow concentrate, L^lo PRP = leukocyte low platelet rich plasma, L^hi PRP = leukocyte high platelet rich plasma. Significance was determined by a Kruskal-Wallis followed by Wilcoxon multiple comparison post-hoc test. A p-value < 0.01 was considered significant.</p

Percentage of cells migrated toward a biologic.

<p>All putative chemoattractants resulted in significantly greater percentage of cells migrated (asterisks) than the NC. Data are represented as median with maximum and minimum values; n = 8. NC = neutral control, PDGF = platelet derived growth factor, BMA = bone marrow aspirate, BMC = bone marrow aspirate concentrate, L^lo PRP = leukocyte low platelet rich plasma, L^hi PRP = leukocyte high platelet rich plasma. Significance was determined by a Kruskal-Wallis followed by Wilcoxon multiple comparison post-hoc test. A p-value < 0.01 was considered significant.</p

A microRNA miR-34a-Regulated Bimodal Switch Targets Notch in Colon Cancer Stem Cells

SummarymicroRNAs regulate developmental cell-fate decisions, tissue homeostasis, and oncogenesis in distinct ways relative to proteins. Here, we show that the tumor suppressor microRNA miR-34a is a cell-fate determinant in early-stage dividing colon cancer stem cells (CCSCs). In pair-cell assays, miR-34a distributes at high levels in differentiating progeny, whereas low levels of miR-34a demarcate self-renewing CCSCs. Moreover, miR-34a loss of function and gain of function alter the balance between self-renewal versus differentiation both in vitro and in vivo. Mechanistically, miR-34a sequesters Notch1 mRNA to generate a sharp threshold response where a bimodal Notch signal specifies the choice between self-renewal and differentiation. In contrast, the canonical cell-fate determinant Numb regulates Notch levels in a continuously graded manner. Altogether, our findings highlight a unique microRNA-regulated mechanism that converts noisy input into a toggle switch for robust cell-fate decisions in CCSCs