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Frederick Grinnell

Publications and source records attributed to Frederick Grinnell.

18 recordsLinked to original sources
PubMed2006

P21-activated kinase 1: convergence point in PDGF- and LPA-stimulated collagen matrix contraction by human fibroblasts.

Fibroblast three-dimensional collagen matrix culture provides a tissue-like model that can be used to analyze cell form and function. The physiological agonists platelet-derived growth factor (PDGF) and lysophosphatidic acid (LPA) both stimulate human fibroblasts to contract floating collagen matrices. In this study, we show that the PDGF and LPA signaling pathways required for matrix contraction converge on p21-activated kinase 1 (PAK1) and its downstream effector cofilin1 and that contraction depends on cellular ruffling activity, rather than on the protrusion and retraction of cellular dendritic extensions. We also show that, depending on the agonist, different Rho effectors cooperate with PAK1 to regulate matrix contraction, Rho kinase in the case of PDGF and mDia1 in the case of LPA. These findings establish a unified framework for understanding the cell signaling pathways involved in fibroblast contraction of floating collagen matrices.

Adaptor Proteins, Signal Transducing↗
PubMed2005

Nested collagen matrices: a new model to study migration of human fibroblast populations in three dimensions.

Fibroblast-3D collagen matrix culture provides a model system to analyze cell physiology under conditions that more closely resemble tissue than conventional 2D cell culture. Previous work has focused primarily on remodeling and contraction of collagen matrices by fibroblasts, and there has been little research on migration of cell populations within the matrix. Here, we introduce a nested collagen matrix model to analyze migration of fibroblasts in 3D collagen matrices. Nested collagen matrices were prepared by embedding contracted cell-containing matrices (also called dermal equivalents) inside cell-free matrices; migration occurred from the former to the latter. Control experiments with human dermal fragments in place of dermal equivalents confirmed the reliability of the model. Human fibroblast migration in nested collagen matrices occurred after a lag phase of 8-16 h, and cells migrating out of the inner matrices were bipolar with leading dendritic extensions. Migration was myosin II, Rho kinase and metalloproteinase-dependent but did not require plasma fibronectin. Platelet-derived growth factor but not lysophosphatidic acid or serum stimulated cell migration, although all three of these physiological agonists promote matrix remodeling and contraction. The nested collagen matrix model is a relatively easy, rapid and quantitative method to measure migration of cell populations. Our studies using this model demonstrate important differences between regulation of fibroblast migration and remodeling in collagen matrices.

Cell Movement↗
PubMed2005

Cell-matrix entanglement and mechanical anchorage of fibroblasts in three-dimensional collagen matrices.

Fibroblast-3D collagen matrix culture provides a physiologically relevant model to study cell-matrix interactions. In tissues, fibroblasts are phagocytic cells, and in culture, they have been shown to ingest both fibronectin and collagen-coated latex particles. Compared with cells on collagen-coated coverslips, phagocytosis of fibronectin-coated beads by fibroblasts in collagen matrices was found to be reduced. This decrease could not be explained by integrin reorganization, tight binding of fibronectin beads to the collagen matrix, or differences in overall bead binding to the cells. Rather, entanglement of cellular dendritic extensions with collagen fibrils seemed to interfere with the ability of the extensions to interact with the beads. Moreover, once these extensions became entangled in the matrix, cells developed an integrin-independent component of adhesion. We suggest that cell-matrix entanglement represents a novel mechanism of cell anchorage that uniquely depends on the three-dimensional character of the matrix.

Cell Culture Techniques↗
PubMed2003

Different molecular motors mediate platelet-derived growth factor and lysophosphatidic acid-stimulated floating collagen matrix contraction.

Fibroblast-collagen matrix contraction has been used as a model system to study how cells organize connective tissue. Previous work showed that lysophosphatidic acid (LPA)-stimulated floating collagen matrix contraction is independent of Rho kinase, whereas platelet-derived growth factor (PDGF)-stimulated contraction is Rho kinase-dependent. The current studies were carried out to learn more about the molecular motors responsible for LPA- and PDGF-stimulated contraction. We found that neither PDGF nor LPA-dependent contractile mechanisms require myosin II regulatory light chain kinase or increased phosphorylation of myosin II regulatory light chain (measured as diphosphorylation). Low concentrations of the specific myosin II inhibitor blebbistatin blocked PDGF-stimulated matrix contraction and LPA-stimulated retraction of fibroblast dendritic extensions but not LPA-stimulated matrix contraction. These data suggest that PDGF- and LPA-stimulated floating matrix contraction utilize myosin II-dependent and -independent mechanisms, respectively. LPA-dependent, Rho kinase-independent force generation also was detected during fibroblast spreading on collagen-coated coverslips.

Cells, Cultured↗
PubMed2003

LPA-stimulated fibroblast contraction of floating collagen matrices does not require Rho kinase activity or retraction of fibroblast extensions.

Fibroblasts synthesize, organize, and maintain connective tissues during development and in response to injury and fibrotic disease. These morphogenetic processes depend on cell-matrix remodeling, which has been investigated using cells cultured in three-dimensional collagen matrices. The current studies were carried out to test the role of Rho kinase activity and retraction of fibroblast extensions on the matrix remodeling process. We found that remodeling (contraction) of floating collagen matrices stimulated by lysophosphatidic acid (LPA) did not require Rho kinase activity or retraction of fibroblast extensions. On the other hand, LPA-stimulated contraction of restrained matrices became Rho kinase dependent after the matrices were allowed to develop mechanical loading for 2-4 h, suggesting that the remodeling process itself was able to feed back to modulate cell behavior in an iterative process. Modulation was specific for LPA since fibroblast-collagen matrix contraction stimulated by platelet-derived growth factor was Rho kinase dependent before or after mechanical loading developed.

Antineoplastic Agents↗
PubMed2003

Fibroblast quiescence in floating collagen matrices: decrease in serum activation of MEK and Raf but not Ras.

Fibroblasts synthesize, organize, and maintain connective tissues during development and in response to injury and fibrotic disease. Studies on cells in three-dimensional collagen matrices have shown that fibroblasts switch between proliferative and quiescence phenotypes, depending upon whether matrices are attached or floating during matrix remodeling. Previous work showed that cell signaling through the ERK pathway was decreased in fibroblasts in floating matrices. In the current research, we extend the previous findings to show that serum stimulation of fibroblasts in floating matrices does not result in ERK translocation to the nucleus. In addition, there was decreased serum activation of upstream members of the ERK signaling pathway, MEK and Raf, even though Ras became GTP loaded. The findings suggest that quiescence of fibroblasts in floating collagen matrices may result from a defect in Ras coupling to its downstream effectors.

Cell Culture Techniques↗
PubMed2003

Fibroblast biology in three-dimensional collagen matrices.

Research on fibroblast biology in three-dimensional collagen matrices offers new opportunities to understand the reciprocal and adaptive interactions that occur between cells and surrounding matrix in a tissue-like environment. Such interactions are integral to the regulation of connective tissue morphogenesis and dynamics that characterizes tissue homeostasis and wound repair. During fibroblast-collagen matrix remodeling, mechanical signals from the remodeled matrix feed back to modulate cell behavior in an iterative process. As mechanical loading (tension) within the matrix increases, the mechanisms used by cells to remodel the matrix change. Fibroblasts in matrices that are under tension or relaxed respond differently to growth factor stimulation, and switching between mechanically loaded and unloaded conditions influences whether cells acquire proliferative/biosynthetic active or quiescent/resting phenotypes.

Animals↗
PubMed2003

Dendritic fibroblasts in three-dimensional collagen matrices.

Cell motility determines form and function of multicellular organisms. Most studies on fibroblast motility have been carried out using cells on the surfaces of culture dishes. In situ, however, the environment for fibroblasts is the three-dimensional extracellular matrix. In the current research, we studied the morphology and motility of human fibroblasts embedded in floating collagen matrices at a cell density below that required for global matrix remodeling (i.e., contraction). Under these conditions, cells were observed to project and retract a dendritic network of extensions. These extensions contained microtubule cores with actin concentrated at the tips resembling growth cones. Platelet-derived growth factor promoted formation of the network; lysophosphatidic acid stimulated its retraction in a Rho and Rho kinase-dependent manner. The dendritic network also supported metabolic coupling between cells. We suggest that the dendritic network provides a mechanism by which fibroblasts explore and become interconnected to each other in three-dimensional space.

Actins↗
PubMed2002

Transforming growth factor beta stimulates fibroblast-collagen matrix contraction by different mechanisms in mechanically loaded and unloaded matrices.

Studies were carried out to test the idea that transforming growth factor beta (TGFbeta) stimulated fibroblast contraction of collagen matrices by different mechanisms depending on mechanical loading on the cells and matrix. Under mechanically unloaded conditions (floating matrices), TGFbeta stimulated contraction directly as an agonist and indirectly by preactivating cells to express the myofibroblast phenotype. Increased contraction of floating matrices by preactivated cells appeared to result in part from an autocrine mechanism. Under mechanically loaded conditions (stressed matrices), TGFbeta had no direct agonist effect on contraction. Fibroblasts preactivated to become myofibroblasts showed increased ability to transfer tension to stressed matrices, and tension persisted even after the cells' actin cytoskeleton was disrupted. Our findings are consistent with the idea that fibroblasts activated to become myofibroblasts in vitro have increased contractile activity and indicate that multiple mechanisms that differ depending on mechanical loading on the cells and matrix are involved.

Actins↗
PubMed2002

Modulation of fibroblast morphology and adhesion during collagen matrix remodeling.

When fibroblasts are placed within a three-dimensional collagen matrix, cell locomotion results in translocation of the flexible collagen fibrils of the matrix, a remodeling process that has been implicated in matrix morphogenesis during development and wound repair. In the current experiments, we studied formation and maturation of cell-matrix interactions under conditions in which we could distinguish local from global matrix remodeling. Local remodeling was measured by the movement of collagen-embedded beads towards the cells. Global remodeling was measured by matrix contraction. Our observations show that no direct relationship occurs between protrusion and retraction of cell extensions and collagen matrix remodeling. As fibroblasts globally remodel the collagen matrix, however, their overall morphology changes from dendritic to stellate/bipolar, and cell-matrix interactions mature from punctate to focal adhesion organization. The less well organized sites of cell-matrix interaction are sufficient for translocating collagen fibrils, and focal adhesions only form after a high degree of global remodeling occurs in the presence of growth factors. Rho kinase activity is required for maturation of fibroblast morphology and formation of focal adhesions but not for translocation of collagen fibrils.

Actinin↗
PubMed2002

Bioethical pluralism and complementarity.

This essay presents complementarity as a novel feature of bioethical pluralism. First introduced by Neils Bohr in conjunction with quantum physics, complementarity in bioethics occurs when different perspectives account for equally important features of a situation but are mutually exclusive. Unlike conventional approaches to bioethical pluralism, which attempt in one fashion or another to isolate and choose between different perspectives, complementarity accepts all perspectives. As a result, complementarity results in a state of holistic, dynamic tension, rather than one that yields singular or final moral judgments.

Bioethics↗