Inhibition of fibroblast proliferation in cardiac myocyte cultures by surface microtopography

Abstract

Cardiac myocyte cultures usually require pharmacological intervention to prevent overproliferation of contaminating nonmyocytes. Our aim is to prevent excessive fibroblast cell proliferation without the use of cytostatins. We have produced a silicone surface with 10-microm vertical projections that we term "pegs," to which over 80% of rat neonatal cardiac fibroblasts attach within 48 h after plating. There was a 50% decrease in cell proliferation by 5 days of culture compared with flat membranes (P < 0.001) and a concomitant 60% decrease (P < 0.01) in cyclin D1 protein levels, suggesting a G1/S1 cell cycle arrest due to microtopography. Inhibition of Rho kinase with 5 or 20 microM Y-27632 reduced attachment of fibroblasts to the pegs by over 50% (P < 0.001), suggesting that this signaling pathway plays an important role in the process. Using mobile and immobile 10-microm polystyrene spheres, we show that reactive forces are important for inhibiting fibroblast cell proliferation, because mobile spheres failed to reduce cell proliferation. In primary myocyte cultures, pegs also inhibit fibroblast proliferation in the absence of cytostatins. The ratio of aminopropeptide of collagen protein from fibroblasts to myosin from myocytes was significantly reduced in cultures from pegged surfaces (P < 0.01), suggesting an increase in the proportion of myocytes on the pegged surfaces. Connexin43 protein expression was also increased, suggesting improved myocyte-myocyte interaction in the presence of pegs. We conclude that this microtextured culture system is useful for preventing proliferation of fibroblasts in myocyte cultures and may ultimately be useful for tissue engineering applications in vivo.

Microtopographical Cues in 3D Attenuate Fibrotic Phenotype and Extracellular Matrix Deposition: Implications for Tissue Regeneration

“…Micropegs on a 2D surface decrease proliferation of fibroblasts compared to flat surfaces. 4 In addition, this was correlated with a significant decrease in Cyclin D1 expression, demonstrating that cell proliferation was affected at the level of G1/S cell cycle transition. 4 Further, this effect was dependent on local micropeg-cell interactions and was regulated by contractile mechanics.…”

“…In the last several years, mechanical, topographical, and geometrical cues have been shown to modulate cellular morphology and behavior in twodimensional (2D) and three-dimensional (3D) systems. [4][5][6][7][8][9][10][11][12] However, the potential to control fibroblast behavior and ECM synthesis using discrete microtopographical cues has not been fully explored. This knowledge could provide a better understanding of the role of topography in matrix remodeling, with the potential to enhance regenerative therapies.…”

Nanopattern-induced changes in morphology and motility of smooth muscle cells

“…Knowledge of these interactions is crucial to the understanding of many fundamental biological questions and to the design of medical devices. Tissue engineering is an example where control of these interactions is essential to the creation of functional engineered-tissues [1][2][3][4][5][6]. The performance of many implantable medical devices is also dependent on the desired device-tissue response [7].…”

Actin dynamics is rapidly regulated by the PTEN and PIP2 signaling pathways leading to myocyte hypertrophy

“…To confirm the PTEN inhibition study, a specific PTEN inhibitor SF1670 originally used in neutrophils (15) and was confirmed here in cardiomyocytes. ANG II increased PIP 2 , which was significantly reduced by SF1670 treatment while also dramatically increasing PIP 3 (Fig. 3, A and B).…”