Reorganization Of Collagen Fibers Research Articles

Myringoplasty and Tympanostomy Tube Placement

Myringoplasty with tympanostomy tube placement may be performed for patients with a variety of middle ear conditions. Often, eustachian tube dysfunction causes otitis media, tympanic membrane perforation, or conductive hearing loss. In the present case, myringoplasty was performed using the CO2 laser, providing reorganization of collagen fibers and improved compliance of the tympanic membrane. Given the ongoing eustachian tube dysfunction, a pressure equalization tube was placed to prevent recurrent retraction and atelectasis of the ear drum.

MMPs Regulate Neuronal Substance P After a Painful Equibiaxial Stretch in a Co-Culture Collagen Gel Model Simulating Injury of an Innervated Ligament

Chronic joint pain is a major health problem that can result from abnormal loading of the innervated ligamentous capsule that surrounds synovial joints. The matrix metalloproteinases-1 (MMP-1) and MMP-9 are hypothesized pain mediators from stretch-induced injuries since they increase in pathologic joint tissues and are implicated in biomechanical and nociceptive pathways that underlay painful joint injuries. There is also emerging evidence that MMP-1 and MMP-9 have mechanistic interactions with the nociceptive neuropeptide substance P. Yet, how a ligament stretch induces painful responses during sub-failure loading and whether MMP-1 or MMP-9 modulates nociception via substance P are unknown. We used a neuron–fibroblast co-culture collagen gel model of the capsular ligament to test whether a sub-failure equibiaxial stretch above the magnitude for initiating nociceptive responses in neurons also regulates MMP-1 and MMP-9. Pre-stretch treatment with the MMP inhibitor ilomastat also tested whether inhibiting MMPs attenuates the stretch-induced nociceptive responses. Because of the role of MMPs in collagen remodeling, collagen microstructural kinematics were measured in all tests. Co-culture gels were incubated for one week in either normal conditions, with five days of ilomastat treatment, or with five days of a vehicle control solution before a planar equibiaxial stretch that imposed strains at magnitudes that induce pain in vivo and increase nociceptive modulators in vitro. Force, displacement, and strain were measured, and polarized light imaging captured collagen fiber kinematics during loading. At 24 h after stretch, immunolabeling quantified substance P, MMP-1, and MMP-9 protein expression. The same sub-failure equibiaxial stretch was imposed on all co-cultures, inducing a significant re-organization of collagen fibers (p ≤ 0.031) indicative of fiber realignment. Stretch induces a doubling of substance P expression in normal and vehicle-treated co-cultures (p = 0.038) that is prevented with ilomastat treatment (p = 0.114). Although MMP-1 and MMP-9 expression are unaffected by the stretch in all co-culture groups, ilomastat treatment abolishes the correlative relationships between MMP-1 and substance P (p = 0.002; R2 = 0.13) and between MMP-1 and MMP-9 (p = 0.007; R2 = 0.11) that are detected without an inhibitor. Collectively, these findings implicate MMPs in a painful ligamentous injury and contribute to a growing body of work linking MMPs to nociceptive-related signaling pathways and/or pain.

Comparison of the systemic action of electroacupuncture and laserpuncture in experimental injuries on the back skin of Wistar rats (Rattus novergicus).

To evaluate the systemic action of electroacupuncture and laserpuncture in the repair of excisional cutaneous injuries on the back of adult female Wistar rats. Ninety animals were divided into three experimental groups: C-control; E-treated with electroacupuncture [acupoints: Feishu (BL13), Geshu (BL17), Zusanli (ST36)]; L-treated with laserpuncture (same acupoints) and euthanized on the 7th, 14th and 21st days for tissue removal and preparation for histomorphometric and biochemical (dosages, zymography and Western blotting) analysis. The number of fibroblasts in the E and L groups presented higher values than the C on the 14th and 21st days. The number of granulocytes was lower than C on the 21st day in groups E and L. The total number of newly formed vessels increased on the 21st day and was higher in both treatments when compared to C. The birefringent collagen fibers detected on the 21st day was lower in groups E and L than group C. The glycosaminoglycans and hydroxyproline amount was similar among groups throughout the experimental period. The active isoform of matrixmetallo proteinase (MMP)-2 and the latent isoform of MMP-9 did not show any differences among all groups and experimental periods. The amount of collagen Ⅰ presented higher values in the L group on the 14th day and in the E and L groups on the 21st day. For type III collagen, groups E and L presented values lower than C in all experimental periods. The amount of transforming growth factor-β1 in the E and L groups showed higher amounts than C on the 7th day and lower on the 14th and 21st days. For vascular endothelial growth factor, E and L groups presented similar and lower values than C on the 7th and 14th days, respectively, and similar on the 21st day. The therapies evaluated in this article stimulated fibroblast proliferation, neoangiogenesis and altered the reorganization of collagen fibers in the repair of excisional injuries in female adult rats.

Effects of adipose-derived mesenchymal stem cells on nociceptive sensitivity and repair processes at achilles tendon injury model in rats

On the model of Achilles tendon injury in rats, the antinociceptive and regenerative effect of allogeneic transplantation of various doses of adipose-derived mesenchymal stem cells (ADMSCs) into the lesion area was studied. It was found that netiher of tested regimens of allogeneic transplantation of ADMSCs contributed to a decrease in the edema of the injured limb. Among the studied regimens, only a twofold administration of 0.25 • 106 ADMSCs exhibited an antinociceptive effect. Administration of 0.50 • 106 ADMSCs promoted the accelerated onset of neovascularization of tendon tissue, while enhancing inflammation and the formation of granulation tissue. Double transplantation of 0.25 • 106 ADMSCs led to accelerated reorganization of collagen fibers, later neovascularization, however, there was an absence of inflammatory infiltrate, lipomatosis, and massive formation of granulation tissue in the lesion area.

The impact of periodontitis in the course of chronic obstructive pulmonary disease: Pulmonary and systemic effects

AimsThe aim of this study was to verify the impact of periodontitis in the course of chronic obstructive pulmonary disease (COPD) in C57Bl/6J mice. Main methodsThe animals were randomly divided into four groups (n = 8): Basal, Periodontitis (P), COPD and COPD+P. COPD was induced by orotracheal instillation of 30 μl of cigarette extract 3 times/week for 7 weeks. Periodontitis was induced by ligation technique for 22 days. Euthanasia was performed on 51st day. The analyzes were total/differential cells and cytokines recovered from bronchoalveolar lavage (BAL), total/differential blood cell count, platelets, total marrow cell count, airway collagen deposition, alveolar enlargement analyzed by mean linear intercept (Lm), mucus and bone crest reabsorption. One-way ANOVA followed by the Student-Newman-Keuls was used. Key findingsThe association COPD+P decreased macrophages (p = 0,0351), TNF-α (p = 0,0071) and INF-γ (p = 0,0004) in BAL, when compared to the COPD group maintaining emphysema levels by alveolar enlargement (p < .05) reorganization of collagen fibers (p = .001) and also mean linear intercept (lm) (p = .001) and mucus (p = .0001). The periodontitis group caused TNF-α increase (p = 0, 0001) in BAL. SignificancePeriodontitis, per se, does not alter any of the parameters analyzed, except for increased TNF-α in BAL. However, its association with COPD caused macrophages TNF-α and INF-γ alterations, when compared to the COPD group maintaining emphysema levels by alveolar enlargement and reorganization of collagen fibers. It seems that periodontitis is influencing the course of Th1 profile cell, and cytokines and pulmonary alterations. Further studies are needed to clarify the regulatory process underlying these two diseases.

Dynamic changes of the extracellular matrix during corneal wound healing

The extracellular matrix (ECM) confers transparency to the cornea because of the precise organization of collagen fibrils and a wide variety of proteoglycans. We monitored the corneal wound healing process after alkali burns in rabbits. We analyzed the location and expression of collagens and proteoglycans, the clinical impact, and the recovery of optical transparency.After the animals received both general and ocular topical anesthesia, the central cornea of the left eye was burned by placing an 8-mm diameter filter paper soaked in 0.5 N NaOH for 60 s. The eyes were evaluated under a surgical microscope at 1, 3, and 6 months after burning. At each time point, the clinical conditions of the burned and control corneas were observed. The arrangement of collagen fibers in the corneal stroma was visualized by Picrosirius-red staining, Gomori's silver impregnation and transmission electronic microscopy. Corneal light transmittance was also measured. Myofibroblasts presence was analyzed by immunohistochemistry. mRNA expression levels of collagen types I and III, lumican, decorin, keratocan and alpha-smooth muscle actin were determined by quantitative real-time polymerase chain reaction.One month after alkali burn, the ECM was disorganized and filled with lacunae containing different types of cells and collagen type III fibers in the wound area. Corneal opacities were present with attendant loss of light transmittance. Collagen and proteoglycan mRNA expression levels were up-regulated. After three months, wound healing progress was indicated by reduced corneal opacity, increased light transmittance, reorganization of collagen fibers and only collagen type I expression levels were at control levels. After six months, the wound area ECM morphology was similar to controls, but transmittance values remained low, denoting incomplete restoration of the stromal architecture.This multidisciplinary study of the stromal wound healing process revealed changes in corneal transmittance, collagen organization, myofibroblasts presence and ECM composition at 1, 3, and 6 months after alkali burning. Documenting wound resolution during the six-month period provided reliable information that can be used to test new therapies.

ПРИМЕНЕНИЕ ПОЛИМЕРНЫХ МАТЕРИАЛОВ В ЛЕЧЕНИИ ЗАБОЛЕВАНИЙ РОГОВИЦЫ

The article presents a critical assessment of the use of polymeric materials in keratoplasty, describing possible postoperative complications. The benefits and disadvantages of implanting various bio-compatible polymers: glyceryl-methacrylate, plastics ACR-7, plexiglass, polymethylmethacrylate, gelatin, hydrogel, and its modifications and others. The significant part of the article focuses on the use of biopolymers, many of which, according to studies, cause an inflammatory reaction, reorganization of collagen fibers and the change of the implant itself. A separate aspect of the article is the use of biodegradable polymers in keratoplasty. The great benefits of using these materials are the ability to control the rate of biodegradation by changing their structure by adding anions and cations superficially active substances. Of all the forms of polymers suitable for ophthalmology in keratoplasty, microspheres, films, and membranes have become very popular. Numerous studies using biodegradable materials have shown that despite the disadvantages (high cost, the complexity of manufacturing), of this type of polymers have great potential in the treatment of various corneal diseases. Keywords: Keratoplasty, polymeric materials, cornea, biodegradable materials, biocompatibility.

Targeted matrisome analysis identifies thrombospondin-2 and tenascin-C in aligned collagen stroma from invasive breast carcinoma

Increasing evidence demonstrates an important role for the extracellular matrix (ECM) in breast cancer progression. Collagen type I, a core constituent of the fibrous ECM, undergoes a significant set of changes that accompany tumor progression, termed Tumor Associated Collagen Signatures (TACS). Late stages of this progression are characterized by the presence of bundled, straight collagen (TACS-2) that become oriented perpendicular to the tumor-stromal boundary (TACS-3). Importantly, the presence of TACS-3 collagen is an independent predictor of poor patient outcome. At present, it remains unclear whether reorganization of the collagen matrix is the consequence of mechanical or compositional tissue remodeling. Here, we identify compositional changes in ECM correlating to collagen fiber reorganization from nineteen normal and invasive ductal carcinoma (IDC) patient biopsies using matrisome-targeted proteomics. Twenty-seven ECM proteins were significantly altered in IDC samples compared to normal tissue. Further, a set of nineteen matrisome proteins positively correlate and five proteins inversely correlate with IDC tissues containing straightened collagen fibers. Tenascin-C and thrombospondin-2 significantly co-localized with aligned collagen fibers in IDC tissues. This study highlights the compositional change in matrisome proteins accompanying collagen re-organization during breast cancer progression and provides candidate proteins for investigation into cellular and structural influences on collagen alignment.

Involvement of mast cells, CD68+ and VEGF+ expressions in response to Himatanthus drasticus commercial latex in mice wound healing model

ABSTRACT This study aimed to evaluate Himatanthus drasticus latex in a mice wound healing experimental model. Animals were divided into four groups (n=7) according to the treatments: GI - saline 0.9% (control), GII - mineral oil (vehicle), GIII - H. drasticus commercial latex (HdCL) and GIV - H. drasticus mixed isolated fraction (MIF, 1 mg/mL). The treatments were applied topically once daily, 50 µL for 14 consecutive days. Macroscopic lesions were evaluated, considering parameters such as swelling, redness, granulation tissue and reepithelialization. VEGF+, CD68+ expressions and mast cells (Toluidin blue stain) were evaluated. HdCL induced higher contraction and exuberant granulation tissue (P &gt; 0.05). HdCL showed a mild inflammatory process while MIF induced intense infiltrate inflammatory predominantly by lymphocytes, vascular congestion, bleeding and did not presented full reepithelialization. Reorganization of collagen fibers (red picrosirius stain) was observed. CD68+ expression and mast cells were presented as moderate, intense and mild in GI, GIII and GIV, respectively. Neovascularization occurred in all groups, while VEGF+ expression was intense in MIF in relation to HdCL. We concluded that HdCL presents wound healing potential, through modulation of mast cells, CD68+ and VEGF+ expressions that can be associated to triterpenes presence according MIF isolated from HdCL.

Multiscale mechanical integrity of human supraspinatus tendon in shear after elastin depletion

Human supraspinatus tendon (SST) exhibits region-specific nonlinear mechanical properties under tension, which have been attributed to its complex multiaxial physiological loading environment. However, the mechanical response and underlying multiscale mechanism regulating SST behavior under other loading scenarios are poorly understood. Furthermore, little is known about the contribution of elastin to tendon mechanics. We hypothesized that (1) SST exhibits region-specific shear mechanical properties, (2) fiber sliding is the predominant mode of local matrix deformation in SST in shear, and (3) elastin helps maintain SST mechanical integrity by facilitating force transfer among collagen fibers. Through the use of biomechanical testing and multiphoton microscopy, we measured the multiscale mechanical behavior of human SST in shear before and after elastase treatment. Three distinct SST regions showed similar stresses and microscale deformation. Collagen fiber reorganization and sliding were physical mechanisms observed as the SST response to shear loading. Measures of microscale deformation were highly variable, likely due to a high degree of extracellular matrix heterogeneity. After elastase treatment, tendon exhibited significantly decreased stresses under shear loading, particularly at low strains. These results show that elastin contributes to tendon mechanics in shear, further complementing our understanding of multiscale tendon structure-function relationships.

On the Spatiotemporal Material Anisotropy of the Vitreous Body in Tension and Compression.

Although linked to several vitreoretinal pathologies including traumatic retinal tears, breaks, and symptomatic vitreomacular traction, the dynamic material behavior of the vitreous body in response to mechanical loads is not well understood. The purpose of this study was to evaluate spatiotemporal patterns of collagen fiber reorganization and vitreous deformation (strain) in response to tensile and compressive forces. Using thick slabs of bovine eyes we examined collagen fiber reorganization following tensile and compressive step-loading with quantitative polarized light imaging. Strains were measured from sparse marker arrays and temporal collagen behavior was estimated from creep compliance rheological tests. Results showed that under applied loads (1) collagen fibers became significantly more aligned at the vitreous base (near the pars plana and the ciliary body), (2) vitreous located directly behind the lens deformed significantly more than surrounding regions, and (3) changes in collagen fiber alignment occurred on a short (<5s) timescale. Together these results show that, despite a homogeneous visual appearance, the vitreous body exhibits anisotropic material behavior in tension and compression. Spatiotemporal patterns of collagen rearrangement were consistent with epidemiological patterns of traumatic retinal damage and vitreoretinal topology. High strains in the vitreous corresponded with locations of lower collagen content that are prone to age-related degeneration. These data suggest that differential fiber alignment and mechanical deformation could contribute to the pathogenesis of these diseases. Computational models that incorporate these experimental data will help improve our understanding of the biomechanical mechanisms that contribute to the pathogenesis of traumatic retinal damage, vitreous degeneration, and vitreoretinal disease.

Matrix metalloproteinase 9 modulates collagen matrices and wound repair

Acute and chronic injuries are characterized by leukocyte infiltration into tissues. Although matrix metalloproteinase 9 (Mmp9) has been implicated in both conditions, its role in wound repair remains unclear. We previously reported a zebrafish chronic inflammation mutant caused by an insertion in the hepatocyte growth factor activator inhibitor gene 1 (hai1; also known as spint1) that is characterized by epithelial extrusions and neutrophil infiltration into the fin. Here, we performed a microarray analysis and found increased inflammatory gene expression in the mutant larvae, including a marked increase in mmp9 expression. Depletion of mmp9 partially rescued the chronic inflammation and epithelial phenotypes, in addition to restoring collagen fiber organization, as detected by second-harmonic generation imaging. Additionally, we found that acute wounding induces epithelial cell mmp9 expression and is associated with a thickening of collagen fibers. Interestingly, depletion of mmp9 impaired this collagen fiber reorganization. Moreover, mmp9 depletion impaired tissue regeneration after tail transection, implicating Mmp9 in acute wound repair. Thus, Mmp9 regulates both acute and chronic tissue damage and plays an essential role in collagen reorganization during wound repair.

Matrix metalloproteinase 9 modulates collagen matrices and wound repair.

Acute and chronic injuries are characterized by leukocyte infiltration into tissues. Although matrix metalloproteinase 9 (Mmp9) has been implicated in both conditions, its role in wound repair remains unclear. We previously reported a zebrafish chronic inflammation mutant caused by an insertion in the hepatocyte growth factor activator inhibitor gene 1 (hai1; also known as spint1) that is characterized by epithelial extrusions and neutrophil infiltration into the fin. Here, we performed a microarray analysis and found increased inflammatory gene expression in the mutant larvae, including a marked increase in mmp9 expression. Depletion of mmp9 partially rescued the chronic inflammation and epithelial phenotypes, in addition to restoring collagen fiber organization, as detected by second-harmonic generation imaging. Additionally, we found that acute wounding induces epithelial cell mmp9 expression and is associated with a thickening of collagen fibers. Interestingly, depletion of mmp9 impaired this collagen fiber reorganization. Moreover, mmp9 depletion impaired tissue regeneration after tail transection, implicating Mmp9 in acute wound repair. Thus, Mmp9 regulates both acute and chronic tissue damage and plays an essential role in collagen reorganization during wound repair.

SMAD3 deficiency promotes vessel wall remodeling, collagen fiber reorganization and leukocyte infiltration in an inflammatory abdominal aortic aneurysm mouse model

TGF-β signaling plays critical roles in the pathogenesis of aneurysms; however, it is still unclear whether its role is protective or destructive. In this study, we investigate the role of SMAD3 in the pathogenesis of calcium chloride (CaCl2)-induced abdominal aortic aneurysms (AAA) in Smad3−/−, Smad3+/− and Smad3+/+ mice. We find that loss of SMAD3 drastically increases wall thickening of the abdominal aorta. Histological analyses show significant vessel wall remodeling with elastic fiber fragmentation. Remarkably, under polarized light, collagen fibers in the hyperplastic adventitia of Smad3−/− mice show extensive reorganization accompanied by loosely packed thin and radial collagen fibers. The expressions of matrix metalloproteinases including MMP2, MMP9, and MMP12 and infiltration of macrophage/T cells are drastically enhanced in the vascular wall of Smad3−/− mice. We also observe marked increase of NF-κB and ERK1/2 signaling as well as the expression of nuclear Smad2, Smad4 and TGF-β1 in the vessel wall of Smad3−/− mice. In addition, we find that SMAD3 expression is reduced in the dedifferentiated medial smooth muscle-like cells of human AAA patients. These findings provide direct in vivo evidence to support the essential roles of SMAD3 in protecting vessel wall integrity and suppressing inflammation in the pathogenesis of AAAs.

Breast Cancer Subtype-Specific Interactions with the Microenvironment Dictate Mechanisms of Invasion

Most ductal breast carcinoma cells are weakly invasive in vitro and in vivo, suggesting that components of their microenvironment may facilitate a transition from in situ to invasive stages during progression. Here, we report that coculture of mammary fibroblasts specifically triggers invasive behavior in basal-type breast cancer cells through a ligand independent mechanism. When cultured alone in organotypic culture, both basal- and luminal-type breast cancer cells formed noninvasive spheroids with characteristics of ductal carcinoma in situ (DCIS). In contrast, when cocultured with mammary fibroblasts, basal-type spheroids exhibited invasive character whereas the luminal-type spheroids retained a benign and noninvasive duct-like architecture. Real-time imaging and functional studies revealed that the specificity of invasion was linked to a unique capacity of basal-type breast cancer cells to move within spheroids. Mammary fibroblasts induced invasion by triggering basal-type breast cancer cells to convert from a noninvasive program of mammary epithelial morphogenesis to an invasive program of sprouting endothelial angiogenesis. Contrary to the existing invasion models, soluble ligands produced by the fibroblasts were not sufficient to trigger invasion. Instead, basal-type invasion relied upon a Cdc42-dependent reorganization of collagen fibers in the extracellular matrix by fibroblasts. Inhibiting basal-type cell movement with clinically relevant drugs blocked invasion both in organotypic culture and in animals, suggesting a new treatment strategy for early-stage patients. Together our findings establish that fibroblast recruitment by basal-type breast cancer cells into early-stage tumors is sufficient to trigger their conversion from a benign, noninvasive DCIS-like stage to a malignant invasive stage. Furthermore, our findings suggest that different subtypes of breast cancer may require distinct types of contributions from the microenvironment to undergo malignant progression.

Evaluation of myelin sheath and collagen reorganization pattern in a model of peripheral nerve regeneration using an integrated histochemical approach

Peripheral nerves are complex histological structures that can be affected by a variety of conditions with different degree of axonal degeneration and demyelination. For the study of peripheral nerve regeneration in pathology and tissue engineering, it is necessary to evaluate the regeneration, remyelination and extracellular matrix reorganization of the neural tissue. Currently, different histochemical techniques must be used in parallel, and a correlation among their findings should be further performed. In this work, we describe a new histochemical method for myelin and collagen fibers based on luxol fast blue and picrosirius methods, for the evaluation of the morphology, the myelin sheath and the collagen fiber reorganization using a model of peripheral nerve regeneration. Whole brain, normal sciatic nerve and regenerating peripheral nerve samples were fixed in 10% neutral buffered formalin and paraffin-embedded, for the performance of the hematoxylin-eosin stain, the Luxol fast blue method and the new histochemical method for myelin and collagen. The results of this technique revealed that this new histochemical method allowed us to properly evaluate histological patterns, and simultaneously observe the histochemical reaction for myelin sheath and collagen fibers in normal tissue, and during the regeneration process. In conclusion, this new method combines morphological and histochemical properties that allowed us to determine with high accuracy the degree of remyelination and collagen fibers reorganization. For all these reasons, we hypothesize that this new histochemical method could be useful in pathology and tissue engineering.

Collagen‐agarose co‐gels as a model for collagen–matrix interaction in soft tissues subjected to indentation

The mechanical properties of soft tissues depend on the collagen fiber network and the surrounding non-fibrillar matrix. The mechanical role of non-fibrillar material remains poorly understood. Our recent study (Lake and Barocas, Ann Biomed Eng 2011) introduced collagen-agarose co-gels as a simple experimental model system to evaluate the mechanical contribution of non-fibrillar matrix, and evaluated co-gel properties in uniaxial tension. In this study, we utilized similar co-gels to examine collagen-matrix interaction in tissues subjected to incremental stress-relaxation indentation tests. Mechanical testing was performed using two orthogonal custom test devices, and polarized light imaging was used to quantify 3D collagen fiber kinematics under load. The addition of agarose led to concentration-dependent changes in the time-dependent mechanical response and magnitude/spread of collagen fiber reorganization of tissue analogs. Specifically, peak/relaxed loads increased, and relaxation rate decreased, with increasing agarose concentration. In addition, increasing agarose content led to larger magnitude changes in orientation direction and alignment strength that were more localized near the indenter. Results suggest that non-fibrillar material significantly contributes to the behavior of co-gels in indentation, likely by reducing permeability and resisting volume change, thereby providing insight into the properties of artificial and native tissues subjected to non-tensile loading.

Evaluation of the effect of Thai breadfruit's heartwood extract on the biological functions of fibroblasts from wrinkles

In previous studies, extract from Artocarpus incisus's heartwood (breadfruit tree) had antioxidant and antimelanogenic activities. Here, we investigated the extract's action on facial skin fibroblasts from wrinkled skin and nonwrinkled skin biopsies, particularly in the production of type I procollagen and metalloproteinase- 1 (MMP-1) and in the reorganization of collagen fibers. We found that the extract at a concentration of 50 microg/ml significantly enhanced percent viability and proliferation of wrinkled-skin fibroblasts. Flow cytometry showed that a 3.6-fold increased proportion of the wrinkled-skin fibroblasts were in their cell cycle S-phase, indicating increased proliferation. Type I procollagen synthesis by wrinkled-skin fibroblasts was augmented by the extract. Nonwrinkled-skin fibroblasts had higher synthesis and were unaffected by the extract. MMP-1 secretion was greater for wrinkled-skin fibroblasts, but the extract decreased its secretion for both fi broblasts samples. Fibroblasts were incorporated in collagen lattice disks. Lattices with nonwrinkled-skin fibroblasts contracted uniformly by 56% after a three-day culture and the extract had little effect. However, wrinkled-skin fi broblast lattices failed to show appreciable contractions (to 12% after three days). But remarkably, the extract conferred an ability of the wrinkled-skin fibroblast lattices to fully contract (to 53%). This shows that wrinkled-skin fi broblasts have the ability to reorganize collagen but that the extract can reactivate this latent potential. Our findings for the first time reveal that A. incisus's heartwood extract reversed the fibroblast deficiencies in the metabolism and reorganization of collagen and may underlie a wrinkle treatment.

Controlling the Spatial Organization of Cells and Extracellular Matrix Proteins in Engineered Tissues Using Ultrasound Standing Wave Fields

Tissue engineering holds great potential for saving the lives of thousands of organ transplant patients who die each year while waiting for donor organs. However, to successfully fabricate tissues and organs in vitro, methodologies that recreate appropriate extracellular microenvironments to promote tissue regeneration are needed. In this study, we have developed an application of ultrasound standing wave field (USWF) technology to the field of tissue engineering. Acoustic radiation forces associated with USWF were used to noninvasively control the spatial distribution of mammalian cells and cell-bound extracellular matrix proteins within three-dimensional (3-D) collagen-based engineered tissues. Cells were suspended in unpolymerized collagen solutions and were exposed to a continuous wave USWF, generated using a 1 MHz source, for 15 min at room temperature. Collagen polymerization occurred during USWF exposure resulting in the formation of 3-D collagen gels with distinct bands of aggregated cells. The density of cell bands was dependent on both the initial cell concentration and the pressure amplitude of the USWF. Importantly, USWF exposure did not decrease cell viability but rather enhanced cell function. Alignment of cells into loosely clustered, planar cell bands significantly increased levels of cell-mediated collagen gel contraction and collagen fiber reorganization compared with sham-exposed samples with a homogeneous cell distribution. Additionally, the extracellular matrix protein, fibronectin, was localized to cell banded areas by binding the protein to the cell surface prior to USWF exposure. By controlling cell and extracellular organization, this application of USWF technology is a promising approach for engineering tissues in vitro. (E-mail: dalecki@bme.rochester.edu)

Three‐dimensional collagen fiber remodeling by mesenchymal stem cells requires the integrin–matrix interaction

Tissue engineering aiming to repair or regenerate damaged tissues necessitates fabricating three-dimensional biomaterial scaffolds with controlled porosity for delivering cells. To facilitate cell distribution, a strategy using stem cell-based fabrication of biomaterials was tested in type II collagen fibers. Human mesenchymal stem cells when delivered in type II collagen assembled and reorganized these matrices and differentiated into spherical chondrocytes with the synthesis of cartilage proteins. The cell-mediated assembly and reorganization of collagen fibers was not limitless and only restricted to an appropriate ratio of cell number and collagen amount. The blocking of alpha2 or beta1-integrin function with specific antibodies significantly impeded the collagen-assembly effects. In vitro chondrogenesis or in vivo cartilage formation of human mesenchymal stem cells was also dependent on the interactions between cells and surrounding matrices. This method for three-dimensional fabricating collagen fibers may generally be applied to other biomaterials, when combined with surface modification or ligand addition for cell adhesion.