Fibrillar force generation by fibroblasts depends on formin.

Fibroblasts in the extra-cellular matrix (ECM) often adopt a predominantly one-dimensional fibrillar geometry by virtue of their adhesion to the fibrils in the ECM. How much forces such fibrillar fibroblasts exert and how they respond to the extended stiffness of their micro-environment comprising of other ECM components and cells are not clear. We use fibroblasts adherent on fibronectin lines micropatterned onto soft polyacrylamide gels as an in vitro experimental model that maintains fibrillar cell morphology while still letting the cell mechanically interact with a continuous micro-environment of specified stiffness. We find that the exerted traction, quantified as the strain energy or the maximum exerted traction stress, is not a function of cell length. Both the strain energy and the maximum traction stress exerted by fibrillar cells are similar for low (13 kPa) or high (45 kPa) micro-environmental stiffness. Furthermore, we find that fibrillar fibroblasts exhibit prominent linear actin structures. Accordingly, inhibition of the formin family of nucleators strongly decreases the exerted traction forces. Interestingly, fibrillar cell migration is, however, not affected under formin inhibition. Our results suggest that fibrillar cell migration in such soft microenvironments is not dependent on high cellular force exertion in the absence of other topological constraints.

Structural Dynamics of the Fibrous Basis of Reparative Regenerate during Spontaneous Healing of the Skin Wound.

Structural dynamics of the fibrous basis of the reparative regenerate during spontaneous skin wound healing comprises multiple stages, it successively transforms from one organization level to another more complex level, forms a multilevel 3D structure including molecular, supramolecular, fibrillar, fiber, and tissue elements. The formed reparative regenerate is integrated with the preserved skin, together they have common fibrous basis consisting of three parts that are different in organization of fibrous structures: atypical (central), tissue organospecific (peripheral), and transitional.

Interactions between fibroblastic reticular cells and B cells promote mesenteric lymph node lymphangiogenesis.

Lymphatic growth (lymphangiogenesis) within lymph nodes functions to promote dendritic cell entry and effector lymphocyte egress in response to infection or inflammation. Here we demonstrate a crucial role for lymphotoxin-beta receptor (LTßR) signaling to fibroblastic reticular cells (FRCs) by lymphotoxin-expressing B cells in driving mesenteric lymph node lymphangiogenesis following helminth infection. LTßR ligation on fibroblastic reticular cells leads to the production of B-cell-activating factor (BAFF), which synergized with interleukin-4 (IL-4) to promote the production of the lymphangiogenic factors, vascular endothelial growth factors (VEGF)-A and VEGF-C, by B cells. In addition, the BAFF-IL-4 synergy augments expression of lymphotoxin by antigen-activated B cells, promoting further B cell-fibroblastic reticular cell interactions. These results underlie the importance of lymphotoxin-dependent B cell-FRC cross talk in driving the expansion of lymphatic networks that function to promote and maintain immune responsiveness.The growth of lymph nodes in response to infection requires lymphangiogenesis. Dubey et al. show that the mesenteric lymph node lymphangiogenesis upon helminth infection depends on the signaling loop between the B and fibroblastic reticular cells (FRCs), whereby the FRCs respond to lymphotoxin secreted by B cells by releasing B cell activating factor.

Influence of the fibroblastic reticular network on cell-cell interactions in lymphoid organs.

Secondary lymphoid organs (SLO), such as lymph nodes and the spleen, display a complex micro-architecture. In the T cell zone the micro-architecture is provided by a network of fibroblastic reticular cells (FRC) and their filaments. The FRC network is thought to enhance the interaction between immune cells and their cognate antigen. However, the effect of the FRC network on cell interaction cannot be quantified to date because of limitations in immunological methodology. We use computational models to study the influence of different densities of FRC networks on the probability that two cells meet. We developed a 3D cellular automaton model to simulate cell movements and interactions along the FRC network inside lymphatic tissue. We show that the FRC network density has only a small effect on the probability of a cell to come into contact with a static or motile target. However, damage caused by a disruption of the FRC network is greatest at FRC densities corresponding to densities observed in the spleen of naïve mice. Our analysis suggests that the FRC network as a guiding structure for moving T cells has only a minor effect on the probability to find a corresponding dendritic cell. We propose alternative hypotheses by which the FRC network might influence the functionality of immune responses in a more significant way.

Bone marrow fibrosis: pathophysiology and clinical significance of increased bone marrow stromal fibres.

In bone marrow biopsies, stromal structural fibres are detected by reticulin and trichrome stains, routine stains performed on bone marrow biopsy specimens in diagnostic laboratories. Increased reticulin staining (reticulin fibrosis) is associated with many benign and malignant conditions while increased trichrome staining (collagen fibrosis) is particularly prominent in late stages of severe myeloproliferative diseases or following tumour metastasis to the bone marrow. Recent evidence has shown that the amount of bone marrow reticulin staining often exhibits no correlation to disease severity, while the presence of type 1 collagen, as detected by trichrome staining, is often associated with more severe disease and a poorer prognosis. It was originally thought that increases in bone marrow stromal fibres themselves contributed to the haematopoietic abnormalities seen in certain diseases, but recent studies suggest that these increases are a result of underlying cellular abnormalities rather than a cause. A growing body of evidence suggests that increased deposition of bone marrow stromal fibres is mediated by transforming growth factor-beta and other factors elaborated by megakaryocytes, but it is likely that other cells, cytokines and growth factors are also involved. This suggests new avenues for investigation into the pathogenesis of various disorders associated with increased bone marrow stromal fibres.

Tissue-specific function of lymph node fibroblastic reticulum cells.

OBJECTIVE: We present the first characterization of the cytokine expression pattern of lymph node fibroblastic reticulum cells (FRC), which are the stromal cells responsible for maintaining the highly structured nodal reticular fiber framework. METHODS: Microarray expression profiles of cultured nodal FRC and dermal fibroblasts (DF) were compared as well as their response to TNF, IL-4, IL-6 and IL-13, cytokines responsible for intranodal stromal activation. RESULTS: Hierarchical clustering of FRC and DF short-term culture samples revealed genes that were differentially expressed in FRC and DF. Identified differently regulated genes were confirmed by RNase protection analysis, PCR or immunohistochemistry. At earlier culture time points, FRC showed higher levels of several chemokines, including CCL2/MCP-1, and cytokines, e.g. IL-6, whereas several genes related to the production of extracellular matrix and angiogenesis were preferentially expressed in early DF cultures. By 60 days in culture, FRC and DF showed similar expression patterns consistent with homogenization of specialized stromal subsets. FRC and DF showed nearly identical transcriptional responses to exogenous TNF stimulation. CONCLUSIONS: Cultured FRC showed an overall transcriptional profile similar to cultured DF, including parallel responsiveness to TNF, but with differences in the expression of chemotactic chemokines, which reflect their biological roles.

An anatomic and biomechanic study of the wrist extensor retinaculum septa and tendon compartments.

PURPOSE: The anatomy of the extensor retinaculum of the wrist has been described previously; the purpose of this study was to describe the specific anatomy of the septal attachments on the radius and to investigate the mechanical strength of each septal attachment on the radius and each of the 6 compartments of the extensor retinaculum. METHODS: Thirty-four wrists from 24 fresh-frozen and 10 embalmed cadavers were used. First, anatomic measurements of the individual extensor retinaculum septums were performed with calipers and a 3-dimensional digitizer. Next each extensor retinaculum septum was excised as a bone-retinaculum-bone autograft and was tested in tension to failure with a materials testing machine. Finally the 6 extensor retinaculum compartments were tested to failure. RESULTS: Septum 1/2 had the largest radial surface area and septum 3/4 had the smallest. Septum 1/2 also was found to have the highest failure strength at 51.3 +/- 15.3 N. In compartment testing, compartments 1 and 2 had the highest overall resistance to failure and compartment 5 had the lowest. Compartment 6, which was thought to be the weakest because of clinically observed subluxation of the extensor carpi ulnaris tendon, had stronger failure data than expected. CONCLUSIONS: This study offers detailed analysis of the extensor retinaculum compartments and 3-dimensional anatomy of the septal attachments. Clinically this study lends insight to the strength of bone-retinaculum-bone autografts and the etiology of extensor carpi ulnaris subluxation.

Cadherin-11 induces rheumatoid arthritis fibroblast-like synoviocytes to form lining layers in vitro.

The synovial lining of diarthrodial joints is composed of a condensed network of synoviocytes that form an intact layer via cell-to-cell contacts with significant intercellular matrix spaces. However, the molecular basis for synovial lining formation and its structural integrity has not been previously defined. In this study, using a three-dimensional fibroblast-like synoviocyte in vitro organ culture system, we provide evidence that cadherin-11 expressed in fibroblast-like synoviocytes plays a determining role in establishing the synovial lining layer. Fibroblast-like synoviocytes that were grown in three-dimensional matrices demonstrated formation of a lining structure at the interface between the matrix and the fluid phase. Treatment of fibroblast-like synoviocyte organ cultures with a cadherin-11-Fc fusion protein efficiently abrogated lining layer organization. Moreover, because E-cadherin-expressing fibroblasts failed to organize a lining layer structure at the tissue boundary, this effect appears to be a distinct characteristic of fibroblasts expressing cadherin-11. We found that cadherin-11 mediated fibroblast-like synoviocyte cell-to-cell adhesion via formation of adherens junctions that were linked to and remodeled the actin cytoskeleton. Together, these studies implicate cadherin-11 in synovial tissue and lining layer formation and provide an in vitro system to model fibroblast-like synoviocyte behavior and function in organizing the synovial tissue.

The conduit system transports soluble antigens from the afferent lymph to resident dendritic cells in the T cell area of the lymph node.

Resident dendritic cells (DC) within the T cell area of the lymph node take up soluble antigens that enter via the afferent lymphatics before antigen carrying DC arrive from the periphery. The reticular network within the lymph node is a conduit system forming the infrastructure for the fast delivery of soluble substances from the afferent lymph to the lumen of high endothelial venules (HEVs). Using high-resolution light microscopy and 3D reconstruction, we show here that these conduits are unique basement membrane-like structures ensheathed by fibroblastic reticular cells with occasional resident DC embedded within this cell layer. Conduit-associated DC are capable of taking up and processing soluble antigens transported within the conduits, whereas immigrated mature DC occur remote from the reticular fibers. The conduit system is, therefore, not a closed compartment that shuttles substances through the lymph node but represents the morphological equivalent to the filtering function of the lymph node.

A model cochlear partition involving longitudinal elasticity.

This paper addresses the issue of longitudinal stiffness within the cochlea. A one-dimensional model of the cochlear partition is presented in which the resonant sections are coupled by longitudinal elastic elements. These elements functionally represent the aggregate mechanical effect of the connective tissue that spans the length of the organ of Corti. With the plate-like morphology of the cochlear partition in mind, the contribution of longitudinal elasticity to partition dynamics is appreciable, though weak and nonlinear. If the elasticity is considered Hookian then the nonlinearity takes a cubic form. Numerical solutions are presented that demonstrate the compressive nature of the partial differential nonlinear equations and their ability to produce realistic cubic distortion product otoacoustic emissions. Within the framework of this model, some speculations can be made regarding the dynamical function of the phalangeal processes, the sharpness of active cochlear mechanics, and the propogation of pathology along the partition.

Relevance and dynamics of myelofibrosis regarding hematopoietic reconstitution after allogeneic bone marrow transplantation in chronic myelogenous leukemia--a single center experience on 160 patients.

A retrospective single center study was performed on 516 trephine biopsies derived from 160 patients with stable phase Ph+-CML and allogeneic BMT. Following morphometric quantification of reticulin-collagen fibers we tried to elucidate (1) the dynamics of bone marrow fibrosis in the post-transplant period; and (2) the influence of manifest myelofibrosis on relevant engraftment parameters. An evaluation of fiber density at standardized endpoints after BMT was carried out on a selected cohort of 124 patients (399 biopsy specimens). A manifest myelofibrosis (more than a three-fold increase compared to the normal fiber content) before BMT was found in 26% of our patients. Concentrating on bone marrow areas with reconstituting hematopoiesis, several findings emerged. Pretransplant myelofibrosis was associated with an initial regression following BMT, but insidiously recurred in the areas of regenerating hematopoiesis or developed in a few patients without increased pregraft fibers during the post-transplant period (mean observation time more than 4 months). Severe acute GVHD (grades III and IV) was significantly correlated with a greater amount of reticulin fibers in the early post-transplant period (9 to 30 days after BMT). Regarding engraftment parameters, a significant delay was detectable in the time to achieve transfusion independence for the patients with manifest myelofibrosis compared to those without pre-transplant fiber increase.

Reticulin-M, an antianaphylactic peptide secreted by reticulo-endothelial system.

I. Moldovan (1923) demonstrated in the blood ultrafiltrate from anaphylactically prepared guinea pigs, which believed to anaphylactic shock, or which were previously injected with coloidal substances as India ink to stimulate the RES, an antianaphylactic principle named Reticulin-M(R). Authors extracted R from organs rich in RES with acetone, established their, peptidic nature and tested it on anaphylactic prepared guinea pig uterine horns in vitro. The peptides were fractioned by high voltage paper electrophoresis. The fractions were cut up in four groups, eluted and tested. The second basic group was identified as active and concentrated by acetone, recipitation. In conclusion several techniques were used to obtain, to test, to isolate and to concentrate R, a natural antianaphylactic peptidic factor, which may be a cytokine.

The differential diagnosis of polycythaemia--a bone marrow study (the bone marrow in polycythaemia).

Bone marrow sections from posterior iliac crest aspiration and/or trephine specimens have been examined in 39 patients with true polycythaemia, along with a variety of other clinical and laboratory data. The emphasis has been on objective assessment of cellularity and megakaryocyte concentration in a prospective four year study. In patients with untreated primary polycythaemia mean cellularity was 87.0% and 86.4% for aspirate and trephine specimens compared with 55.5% and 48.7% for secondary cases and 48.3% and 45.5% for controls. Eighty per cent of primary polycythemia patients had moderate to marked megakaryocytic hyperplasia. We conclude that, in the presence of an elevated red cell volume, marrow cellularity of greater than 75%, particularly when accompanied by megakaryocytic hyperplasia, of iliac crest aspirate or trephine specimens is sufficient per se to establish a diagnosis of polycythaemia rubra vera.

Functionality of muscle proteins in gelation mechanisms of structured meat products.

Recent advances in muscle biology concerning the discoveries of a large variety of proteins have been described in this review. The existence of polymorphism in several muscle proteins is now well established. Various isoforms of myosin not only account for the difference in physiological functions and biochemical activity of different fiber types or muscles, but also seem to differ in functional properties in food systems. The functionality of various muscle proteins, especially myosin and actin in the gelation process in modal systems which simulate structured meat products, is discussed at length. Besides, the role of different subunits and subfragments of myosin molecule in the gelation mechanism, and the various factors affecting heat-induced gelation of actomyosin in modal systems are also highlighted. Finally, the areas which need further investigation in this discipline have been suggested.