Biomedical subjects

Jan Lammerding

Publications and source records attributed to Jan Lammerding.

10 recordsLinked to original sources

PubMed2006

Lamins A and C but not lamin B1 regulate nuclear mechanics.

Mutations in the nuclear envelope proteins lamins A and C cause a broad variety of human diseases, including Emery-Dreifuss muscular dystrophy, dilated cardiomyopathy, and Hutchinson-Gilford progeria syndrome. Cells lacking lamins A and C have reduced nuclear stiffness and increased nuclear fragility, leading to increased cell death under mechanical strain and suggesting a potential mechanism for disease. Here, we investigated the contribution of major lamin subtypes (lamins A, C, and B1) to nuclear mechanics by analyzing nuclear shape, nuclear dynamics over time, nuclear deformations under strain, and cell viability under prolonged mechanical stimulation in cells lacking both lamins A and C, cells lacking only lamin A (i.e. "lamin C-only" cells), cells lacking wild-type lamin B1, and wild-type cells. Lamin A/C-deficient cells exhibited increased numbers of misshapen nuclei and had severely reduced nuclear stiffness and decreased cell viability under strain. Lamin C-only cells had slightly abnormal nuclear shape and mildly reduced nuclear stiffness but no decrease in cell viability under strain. Interestingly, lamin B1-deficient cells exhibited normal nuclear mechanics despite having a significantly increased frequency of nuclear blebs. Our study indicates that lamins A and C are important contributors to the mechanical stiffness of nuclei, whereas lamin B1 contributes to nuclear integrity but not stiffness.

Animals↗

PubMed2006

Prelamin A and lamin A appear to be dispensable in the nuclear lamina.

Lamin A and lamin C, both products of Lmna, are key components of the nuclear lamina. In the mouse, a deficiency in both lamin A and lamin C leads to slow growth, muscle weakness, and death by 6 weeks of age. Fibroblasts deficient in lamins A and C contain misshapen and structurally weakened nuclei, and emerin is mislocalized away from the nuclear envelope. The physiologic rationale for the existence of the 2 different Lmna products lamin A and lamin C is unclear, although several reports have suggested that lamin A may have particularly important functions, for example in the targeting of emerin and lamin C to the nuclear envelope. Here we report the development of lamin C-only mice (Lmna(LCO/LCO)), which produce lamin C but no lamin A or prelamin A (the precursor to lamin A). Lmna(LCO/LCO) mice were entirely healthy, and Lmna(LCO/LCO) cells displayed normal emerin targeting and exhibited only very minimal alterations in nuclear shape and nuclear deformability. Thus, at least in the mouse, prelamin A and lamin A appear to be dispensable. Nevertheless, an accumulation of farnesyl-prelamin A (as occurs with a deficiency in the prelamin A processing enzyme Zmpste24) caused dramatically misshapen nuclei and progeria-like disease phenotypes. The apparent dispensability of prelamin A suggested that lamin A-related progeroid syndromes might be treated with impunity by reducing prelamin A synthesis. Remarkably, the presence of a single Lmna(LCO) allele eliminated the nuclear shape abnormalities and progeria-like disease phenotypes in Zmpste24-/- mice. Moreover, treating Zmpste24-/- cells with a prelamin A-specific antisense oligonucleotide reduced prelamin A levels and significantly reduced the frequency of misshapen nuclei. These studies suggest a new therapeutic strategy for treating progeria and other lamin A diseases.

Animals↗

PubMed2005

Focal adhesion kinase signaling regulates cardiogenesis of embryonic stem cells.

The signaling steps that induce cardiac differentiation in embryonic stem (ES) cells are incompletely understood. We examined the effect of adhesion signaling including Src and focal adhesion kinase (FAK) on cardiogenesis in mouse ES cells using alpha-myosin heavy chain promoter-driven enhanced green fluorescent protein or luciferase as reporters. Cardiac transcription factors including Nkx2.5 and Tbx5 mRNA were first expressed at day 4 in hanging drop embryoid bodies, and adhesion of embryoid bodies to surfaces at or before that day strongly inhibited differentiation of ES cells to cardiomyocytes. Since adhesion signaling could suppress cardiogenesis through Src kinases, embryoid bodies were exposed to the small molecule PP2, known as a Src family kinase inhibitor. PP2 during embryoid body adhesion dramatically increased cardiomyocyte differentiation and decreased mRNA expression of neuronal cellular adhesion molecule and alpha-fetoprotein, neuroectodermal, and endodermal markers, respectively. Surprisingly, although there was an interaction between Src and FAK in cardiogenesis, the procardiogenic effect of PP2 appeared incompletely explained by Src kinase inhibition, since another Src family kinase inhibitor, SU6656, failed to induce cardiogenesis. Instead, PP2 specifically inhibited adhesion-induced FAK phosphorylation. In ES cells stably expressing FAK-related nonkinase, which functions as a dominant negative FAK, cell migration from embryoid bodies was inhibited, whereas alpha-myosin heavy chain expression and myosin-stained cardiomyocytes were increased, suggesting that reducing cell motility may contribute to cardiogenesis. These data indicate that FAK is a key regulator of cardiogenesis in mouse ES cells and that FAK signaling within embryoid bodies can direct stem cell lineage commitment.

Animals↗

PubMed2005

Abnormal nuclear shape and impaired mechanotransduction in emerin-deficient cells.

Emery-Dreifuss muscular dystrophy can be caused by mutations in the nuclear envelope proteins lamin A/C and emerin. We recently demonstrated that A-type lamin-deficient cells have impaired nuclear mechanics and altered mechanotransduction, suggesting two potential disease mechanisms (Lammerding, J., P.C. Schulze, T. Takahashi, S. Kozlov, T. Sullivan, R.D. Kamm, C.L. Stewart, and R.T. Lee. 2004. J. Clin. Invest. 113:370-378). Here, we examined the function of emerin on nuclear mechanics and strain-induced signaling. Emerin-deficient mouse embryo fibroblasts have abnormal nuclear shape, but in contrast to A-type lamin-deficient cells, exhibit nuclear deformations comparable to wild-type cells in cellular strain experiments, and the integrity of emerin-deficient nuclear envelopes appeared normal in a nuclear microinjection assay. Interestingly, expression of mechanosensitive genes in response to mechanical strain was impaired in emerin-deficient cells, and prolonged mechanical stimulation increased apoptosis in emerin-deficient cells. Thus, emerin-deficient mouse embryo fibroblasts have apparently normal nuclear mechanics but impaired expression of mechanosensitive genes in response to strain, suggesting that emerin mutations may act through altered transcriptional regulation and not by increasing nuclear fragility.

Animals↗

PubMed2005

The nuclear membrane and mechanotransduction: impaired nuclear mechanics and mechanotransduction in lamin A/C deficient cells.

Mutations in the lamin A/C gene cause a variety of human diseases including Emery-Dreifuss muscular dystrophy, dilated cardiomyopathy and Hutchinson-Gilford progeria syndrome. The tissue specific effects of lamin mutations are unclear, in part because the function of lamin A/C is incompletely defined, but the many muscle specific phenotypes suggest that defective lamin A/C could increase cellular mechanical sensitivity. Lamin A/C deficient fibroblasts were subjected to mechanical strain to measure nuclear mechanical properties and strain-induced signalling. We found that lamin A/C deficient fibroblasts are characterized by impaired nuclear mechanics and mechanotransduction, reflected by increased nuclear deformations, increased nuclear fragility, attenuated expression of mechanosensitive genes, and impaired transcriptional activation, leading to impaired viability of mechanically strained cells. Lamins and other nuclear envelope proteins can thus affect several levels of the mechanotransduction cascade, altering nuclear and cytoskeletal mechanics as well as playing an important role in mechanically activated gene regulation. Individual mutations in the lamin A/C gene could potentially selectively interfere with any of these functions, explaining the tissue-specific effects observed in the laminopathies.

Animals↗

PubMed2004

Lamin A/C deficiency causes defective nuclear mechanics and mechanotransduction.

Mutations in the lamin A/C gene (LMNA) cause a variety of human diseases including Emery-Dreifuss muscular dystrophy, dilated cardiomyopathy, and Hutchinson-Gilford progeria syndrome. The tissue-specific effects of lamin mutations are unclear, in part because the function of lamin A/C is incompletely defined, but the many muscle-specific phenotypes suggest that defective lamin A/C could increase cellular mechanical sensitivity. To investigate the role of lamin A/C in mechanotransduction, we subjected lamin A/C-deficient mouse embryo fibroblasts to mechanical strain and measured nuclear mechanical properties and strain-induced signaling. We found that Lmna-/- cells have increased nuclear deformation, defective mechanotransduction, and impaired viability under mechanical strain. NF-kappaB-regulated transcription in response to mechanical or cytokine stimulation was attenuated in Lmna-/- cells despite increased transcription factor binding. Lamin A/C deficiency is thus associated with both defective nuclear mechanics and impaired mechanically activated gene transcription. These findings suggest that the tissue-specific effects of lamin A/C mutations observed in the laminopathies may arise from varying degrees of impaired nuclear mechanics and transcriptional activation.

Animals↗

PubMed2004

Mechanotransduction in cardiac myocytes.

Cardiac myocytes react to diverse mechanical demands with a multitude of transient and long-term responses to normalize the cellular mechanical environment. Several stretch-activated signaling pathways have been identified, most prominently guanine nucleotide binding proteins (G-proteins), mitogen-activated protein kinases (MAPK), Janus-associated kinase/signal transducers and activators of transcription (JAK/STAT), protein kinase C (PKC), calcineurin, intracellular calcium regulation, and several autocrine and paracrine factors. Multiple levels of crosstalk exist between pathways. The cellular response to changes in the mechanical environment can lead to cardiac myocyte hypertrophy, cellular growth that can be accompanied by pathological myocyte dysfunction, and tissue fibrosis. Several candidates for the primary mechanosensor in cardiac myocytes have been identified, ranging from stretch-activated ion channels in the membrane to yet-unknown mechanosensitive mechanisms in the nucleus. New and refined experimental techniques will exploit advances in molecular biology and biological imaging to study mechanotransduction in isolated cells and genetically engineered mice to explore the function of individual proteins.

Animals↗

PubMed2003

Tetraspanin CD151 regulates alpha6beta1 integrin adhesion strengthening.

The tetraspanin CD151 molecule associates specifically with laminin-binding integrins, including alpha6beta1. To probe strength of alpha6beta1-dependent adhesion to laminin-1, defined forces (0-1.5 nN) were applied to magnetic laminin-coated microbeads bound to NIH 3T3 cells. For NIH 3T3 cells bearing wild-type CD151, adhesion strengthening was observed, as bead detachment became more difficult over time. In contrast, mutant CD151 (with the C-terminal region replaced) showed impaired adhesion strengthening. Static cell adhesion to laminin-1, and detachment of beads coated with fibronectin or anti-alpha6 antibody were all unaffected by CD151 mutation. Hence, CD151 plays a key role in selectively strengthening alpha6beta1 integrin-mediated adhesion to laminin-1.

3T3 Cells↗

PubMed2003

A three-dimensional viscoelastic model for cell deformation with experimental verification.

A three-dimensional viscoelastic finite element model is developed for cell micromanipulation by magnetocytometry. The model provides a robust tool for analysis of detailed strain/stress fields induced in the cell monolayer produced by forcing one microbead attached atop a single cell or cell monolayer on a basal substrate. Both the membrane/cortex and the cytoskeleton are modeled as Maxwell viscoelastic materials, but the structural effect of the membrane/cortex was found to be negligible on the timescales corresponding to magnetocytometry. Numerical predictions are validated against experiments performed on NIH 3T3 fibroblasts and previous experimental work. The system proved to be linear with respect to cytoskeleton mechanical properties and bead forcing. Stress and strain patterns were highly localized, suggesting that the effects of magnetocytometry are confined to a region extending <10 microm from the bead. Modulation of cell height has little effect on the results, provided the monolayer is >5 micro m thick. NIH 3T3 fibroblasts exhibited a viscoelastic timescale of approximately 1 s and a shear modulus of approximately 1000 Pa.

Animals↗

PubMed2002

Signaling pathways that influence extracellular remodeling.

BACKGROUND: Remodeling of myocardial tissue requires a rearrangement of cells and extracellular matrix to form the new geometry through processes that are incompletely defined. Exploring new pathways beyond neurohormonal inhibition is essential for developing new therapies for the growing epidemic of heart failure. METHODS: One strategy relies on the discovery that progressive ventricular dilation requires matrix metalloproteinases, a family of enzymes that degrade components of matrix but may also participate in activation or release of signaling molecules. Here we will briefly review evidence that matrix metalloproteinase inhibition represents a potential strategy for preventing heart failure. CONCLUSION: Future applications for understanding cell-matrix interactions, including discovering new pathways with proteomics, will also be discussed. Finally, we propose that defining matrix-remodeling events at the level of the membrane and matrix receptors will be essential, and that new bioengineering tools will provide us with the necessary methods.

Extracellular Matrix↗