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Aim The aim of the present work was to explain the poor biointegration of acellular dermal xenogeneic matrix, leading to an unfavorable gingival healing following a grafting procedure for the treatment of soft tissue deficiencies. Background Numerous works have demonstrated the successful use of acellular dermal matrix (ADM) in soft tissue augmentation procedures. However, spare human investigations reported adverse healing outcomes at microscopic level. Case description Three patients showing various soft tissue deficiencies (recession, gingival thickening) requiring a gingival augmentation were grafted using an ADM porcine acellular dermal matrices (pADM) as a soft tissue substitute. For this purpose, appropriate soft tissue augmentation surgeries were performed and the grafted pADM was left for proper healing. Biopsies were harvested from two out of the three patients, respectively, at 11 and 27 weeks in order to conduct a histological evaluation of the pADM's doubtful biointegration. Moreover, the ultrastructural analysis of pADM was performed using scanning electron microscopy, and additional histological procedures were used to assess its ability to support human gingival fibroblast cultures. Signs of gingival inflammation persisted several months postoperatively. Histologically, numerous inflammatory cells characterized the grafted site. Indeed, the high number of foreign body giant cell granulomas and the very densified newly formed collagen fibers highlighted a fibrotic process within gingival connective tissue. The ultrastructural and histological analysis showed that pADM was characterized by very thick and dense collagen bundles demonstrating a nonphysiological collagen network organization. Cell culture experiments showed fibroblasts proliferating on the matrix surface, sparing its deeper part, even though the collagen matrix degradation seemed to occur following a gradient from the pADM surface inward. Conclusion The unfavorable clinical results may be caused by the poor colonization of matrix cells and poor angiogenesis leading to the inadequate biointegration of pADM. Hence, the pADM structure in terms of porosity and degradability should be further investigated. Clinical significance The present cases highlighted a poor integration of pADM following soft tissue grafting procedures, which was caused by the inadequate ultrastructure of the used pADM. Therefore, despite the utility of such tissue substitutes, their manufacturing improvement could be required to obtain a better biointegration.

Three-dimensional (3D) spheroid culture provides opportunities to model tumor growth closer to its natural context. The collagen network in the extracellular matrix supports autonomic tumor cell proliferation, but its presence and role in tumor spheroids remain unclear. In this research, we developed an in vitro 3D co-culture model in a microwell 3D (μ-well 3D) cell-culture array platform to mimic the tumor microenvironment (TME). The modular setup is used to characterize the paracrine signaling molecules and the role of the intraspheroidal collagen network in cancer drug resistance. The μ-well 3D platform is made up of poly(dimethylsiloxane) that contains 630 round wells for individual spheroid growth. Inside each well, the growth surface measured 500 μm in diameter and was functionalized with the amphiphilic copolymer. HCT-8 colon cancer cells and/or NIH3T3 fibroblasts were seeded in each well and incubated for up to 9 days for TME studies. It was observed that NIH3T3 cells promoted the kinetics of tumor organoid formation. The two types of cells self-organized into core-shell chimeric tumor spheroids (CTSs) with fibroblasts confined to the shell and cancer cells localized to the core. Confocal microscopy analysis indicated that a type-I collagen network developed inside the CTS along with increased TGF-β1 and α-SMA proteins. The results were correlated with a significantly increased stiffness in 3D co-cultured CTS up to 52 kPa as compared to two-dimensional (2D) co-culture. CTS was more resistant to 5-FU (IC50 = 14.0 ± 3.9 μM) and Regorafenib (IC50 = 49.8 ± 9.9 μM) compared to cells grown under the 2D condition 5-FU (IC50 = 12.2 ± 3.7 μM) and Regorafenib (IC50 = 5.9 ± 1.9 μM). Targeted collagen homeostasis with Sclerotiorin led to damaged collagen structure and disrupted the type-I collagen network within CTS. Such a treatment significantly sensitized collagen-supported CTS to 5-FU (IC50 = 4.4 ± 1.3 μM) and to Regorafenib (IC50 = 0.5 ± 0.2 μM). In summary, the efficient formation of colon cancer CTSs in a μ-well 3D culture platform allows exploration of the desmoplastic TME. The novel role of intratumor collagen quality as a drug sensitization target warrants further investigation.

Homograft availability and durability remain big challenges. Increasing the post-mortem ischaemic harvesting time beyond 24 h increases the potential donor pool. Cryopreservation, routinely used to preserve homografts, damages the extracellular matrix (ECM), contributing to valve degeneration. Decellularization might preserve the ECM, promoting host-cell infiltration and contributing towards better clinical outcomes. This study compared the performance of cryopreserved versus decellularized pulmonary homografts in the right ventricle outflow tract (RVOT) of a juvenile ovine model. Homografts (n = 10) were harvested from juvenile sheep, subjected to 48 h post-mortem cold ischaemia, cryopreserved or decellularized and implanted in the RVOT of juvenile sheep for 180 days. Valve performance was monitored echocardiographically. Explanted leaflet and wall tissue evaluated histologically, on electron microscopical appearance, mechanical properties and calcium content. In both groups the annulus diameter increased. Cryopreserved homografts developed significant (¾) pulmonary regurgitation, with trivial regurgitation (¼) in the decellularized group. Macroscopically, explanted cryopreserved valve leaflets retracted and thickened while decellularized leaflets remained thin and pliable with good coaptation. Cryopreserved leaflets and walls demonstrated loss of interstitial cells with collapsed collagen, and decellularized scaffolds extensive, uniform ingrowth of host-cells with an intact collagen network. Calcific deposits were shown only in leaflets and walls of cryopreserved explants. Young fibroblasts, with vacuoles and rough endoplasmic reticulum in the cytoplasm, repopulated the leaflets and walls of decellularized scaffolds. Young's modulus of wall tissue in both groups increased significantly. Cryopreserved valves deteriorate over time due to loss of cellularity and calcification, while decellularized scaffolds demonstrated host-cell repopulation, structural maintenance, tissue remodelling and growth potential.

Introduction and Aim:Isosorbide mononitrate, a nitric oxide donor, induces the enzyme cyclo-oxygenase-2, leading to production of local prostaglandins in human cervix and leads to structural rearrangement of the collagen network and thereby the cervix ripens.The aim of this study was to determine the role of Isosorbide mononitrate in induction of labour, determine induction delivery interval and obstetric and neonatal outcome. Materials and Methods:This study was an interventional study on 40 pregnant women conducted for a period of one year. Women requiring induction of labour, who met the inclusion criteria, were enrolled. Isosorbide mononitrate (40 milligrams) was inserted in the posterior fornix. If Bishop's score did not change after 24 hours of the initial dose, failure of induction of labour was considered.Those patients in whom labour pains started after isosorbide mononitrate alone were considered to be in successful group(A) while those who required additional drugs like prostaglandins were considered to be in the failed group(F). Results:The mean (SD) Bishop score after 1dose of isosorbide mononitrate in the successful group (A) was 6.04 (2.16) and in the failed group (F) was 3.81 (0.83). There was a significant difference in terms of Bishop Score after 1st Dose of IMN (W = 310.000, p = 0.001) and in terms of induction-delivery time in hours (t = -2.386, p = 0.023). Conclusion:This study concludes that Isosorbide mononitrate has significant role for induction of labor in terms of Bishop score changes and induction delivery interval.

Tissue injury results in profound alterations in the collagen network, associated with unfolding of the collagen triple helix, proteolytic degradation and generation of fragments. In the infarcted myocardium, changes in the collagen network are critically involved in the pathogenesis of left ventricular rupture, adverse remodeling and chronic dysfunction. We hypothesized that myocardial infarction is associated with temporally and spatially restricted patterns of collagen denaturation that may reflect distinct molecular mechanisms of collagen unfolding. We used a mouse model of non-reperfused myocardial infarction, and in vitro assays in fibroblast-populated collagen lattices. In healing infarcts, labeling with collagen hybridizing peptide (CHP) revealed two distinct patterns of collagen denaturation. During the inflammatory and proliferative phases of infarct healing, collagen denaturation was pericellular, localized in close proximity to macrophages and myofibroblasts. qPCR array analysis of genes associated with matrix remodeling showed that Membrane Type 1-Matrix Metalloproteinase (MT1-MMP) is markedly upregulated in infarct macrophages and fibroblasts, suggesting its involvement in pericellular collagen denaturation. In vitro, MT1-MMP-mediated pericellular collagen denaturation is involved in cardiac fibroblast migration. The effects of MT1-MMP on collagen denaturation and fibroblast migration involve the catalytic site, and require hemopexin domain-mediated actions. In contrast, during the maturation phase of infarct healing, extensive collagen denaturation was noted in the hypocellular infarct, in the infarct border zone and in the mitral valve annulus, in the absence of MT1-MMP. In vitro, mechanical tension in attached collagen lattices was sufficient to induce peripheral collagen denaturation. Our study suggests that in healing infarcts, early pericellular collagen denaturation may be important for migration of macrophages and reparative myofibroblasts in the infarct. Extensive denaturation of collagen fibers is noted in mature scars, likely reflecting mechanical tension. Chronic collagen denaturation may increase susceptibility of the matrix to proteolysis, thus contributing to progressive cardiac dilation and post-infarction heart failure.

Summary Objective: Knee osteoarthritis (OA) is associated with meniscal degeneration that may involve disorganization of the meniscal collagen fiber network. Our aims were to quantitatively analyze the microstructural organization of human meniscus samples in 3D using micro-computed tomography (μCT), and to compare the local microstructural organization between OA and donor samples. Method: We collected posterior horns of both medial and lateral human menisci from 10 end-stage medial compartment knee OA patients undergoing total knee replacement (medial & lateral OA) and 10 deceased donors without knee OA (medial & lateral donor). Posterior horns were dissected and fixed in formalin, dehydrated in ascending ethanol concentrations, treated with hexamethyldisilazane (HMDS), and imaged with μCT. We performed local orientation analysis of collagenous microstructure in 3D by calculating structure tensors from greyscale gradients within selected integration window to determine the polar angle for each voxel. Results: In donor samples, meniscus bundles were aligned circumferentially around the inner border of meniscus. In medial OA menisci, the organized structure of collagen network was lost, and main orientation was shifted away from the circumferential alignment. Quantitatively, medial OA menisci had the lowest mean orientation angle compared to all groups, −24° (95%CI -31 to −18) vs medial donor and −25° (95%CI -34 to −15) vs lateral OA. Conclusions: HMDS-based μCT imaging enabled quantitative analysis of meniscal collagen fiber bundles and their orientations in 3D. In human medial OA menisci, the collagen disorganization was profound with overall lower orientation angles, suggesting collagenous microstructure disorganization as an important part of meniscus degradation.

Osteoclasts, the multinucleated cells responsible for bone resorption, have an enormous destructive power which demands to be kept under tight control. Accordingly, the identification of molecular signals directing osteo- clastogenesis and switching on their resorptive activity have received much attention. Mandatory factors were identified, but a very essential aspect of the control mechanism of osteoclastic resorption, i.e. its spatial control, remains poorly understood. Under physiological conditions, multinucleated osteoclasts are only detected on the bone surface, while their mono-nucleated precursors are only in the bone marrow. How are pre-osteoclasts targeted to the bone surface? How is their progressive differentiation coordinated with their approach to the bone surface sites to be resorbed, which is where they finally fuse? Here we review the information on the bone marrow distribution of differentiating pre-osteoclasts relative to the position of the mandatory factors for their differentiation as well as relative to physical entities that may affect their access to the remodelling sites. This info allows recognizing an “osteoclastogenesis route” through the bone marrow and leading to the coincident fusion/resorption site – but also points to what still remains to be clarified regarding this route and regarding the restriction of fusion at the resorption site. Finally, we discuss the mechanism responsible for the start of re- sorption and its spatial extension. This review underscores that fully understanding the control of bone re- sorption requires to consider it in both space and time - which demands taking into account the context of bone tissue.

The present research aims to determine whether the application of non‐pigmented ciliary epithelium cells derived extracellular vesicles to human trabecular meshwork cells affects the formation and secretion of collagen type I to the extracellular matrix formation. Following the extraction of non‐pigmented ciliary epithelium derived extracellular vesicles by a precipitation method, their size and concentration were determined using tunable resistive pulse sensing technology. Extracellular vesicles were incubated with trabecular meshwork cells for 3 days. Morphological changes of collagen type I in the extracellular matrix of trabecular meshwork cells were visualized using confocal microscopy and scanning electron microscopy. A Sirius Red assay was used to determine the total amount of collagen. Finally, collagen type I expression levels in the extracellular matrix of trabecular meshwork cells were quantified by cell western analysis. We found that non‐pigmented ciliary epithelium extracellular vesicles were very effective at preventing collagen fibres formation by the trabecular meshwork cells, and their secretion to the extracellular matrix was significantly reduced (P

In recent years, engineering biomimetic cellular microenvironments have been a top priority for regenerative medicine. Collagen II, which is arranged in arches, forms the predominant fiber network in articular cartilage. Due to the shortage of suitable microfabrication techniques capable of producing 3D fibrous structures, in vitro replication of the arch-like cartilaginous tissue constitutes one of the major challenges. Hence, in the present study, we report a 3D bioprinting approach for fabricating arch-like constructs using two types of bioinks, gelatin methacryloyl (GelMa) and silk fibroin-gelatin (SF-G). The bioprinted SF-G constructs displayed increased proliferation of the encapsulated human bone marrow-derived mesenchymal stem cells compared to the GelMA constructs. Biochemical assays, gene, and protein expression exhibited the superior role of SF-G in forming the fibrous collagen network and chondrogenesis. Protein-protein interaction study using Metascape evaluated the function of the proteins involved. Further GeneMANIA and STRING analysis using Col 2A1, SOX 9, ACAN, and the genes upregulated on day 21 in RT-PCR, i.e. β-catenin, TGFβR1, Col 1A1 in SF-G and PRG4, Col 10A1, MMP 13 in GelMA validated our in vitro results. These findings emphasized the role of SF-G in regulating the Wnt/β-catenin and TGF-β signaling pathways. Hence, the 3D bioprinted arch-like constructs possess a substantial potential for cartilage regeneration.

Within the panniculus carnosus‐associated skeletal muscles in the human, the palmaris brevis and the platysma showed myotendinous/myofascial junctions with clear distance to the corium and the specific connection collagen type XXII. The orbicularis oris muscle, in contrast, contained bundles of striated muscle fibers reaching the corium at two distinct levels: the predominant inner ending was connected to the elastic network of the inner corium and the outer ending was within the more superficial collagen network. At both locations, the striated muscle fibers showed brush‐like cytoplasmic protrusions connecting a network which was not oriented toward the muscle fibers. Collagen type XXII was not present., While palmaris brevis and platysma muscles show true myotendinous junctions, the orbicularis oris muscle develops mimic muscle‐specific myocutaneous junctions. They insert brush like into the dermis and do not contain collagen XXII.

Intrahepatic cholangiocarcinomas (iCCs) are primary tumors of the liver characterized by the presence of a desmoplastic stroma. While tumor stroma may have a protective or a pejorative value depending on the type of malignant disease, the precise role of the desmoplastic stroma in iCC remains poorly understood. The aim of the present study was to evaluate the prognostic value of stromal compartment in iCC through a multiparametric morphological analysis. Forty-nine surgically resected iCCs were included. For all cases, tumor paraffin blocks of iCCs were selected for stromal morphological characterization through quantitative and qualitative approaches using immunohistochemistry and second-harmonic generation imaging. Intratumor heterogeneity was also evaluated in regards with the different stromal features. High proportionated stromal area (PSA) (defined by stromal to tumor area ratio) was inversely correlated with vascular invasion (62.5% vs 95.7%, p = 0.006) and positively correlated with well-differentiated grade (60% vs 12.5%, p = 0.001). Patients with high PSA had a better disease-free survival (DFS) than patients with low stromal area (60% vs 10%, p = 0.077). Low activated stroma index (defined by cancer-associated fibroblasts number to stromal area ratio) was associated with a better DFS (60% vs 10%, p = 0.05). High collagen reticulation index (CRI), defined as the number of collagen fiber branches within the entire length of the collagen network, was associated with a poorer overall survival (42% vs NR, p = 0.026). Furthermore, we showed that CRI was also an homogeneous marker throughout the tumor. Based on morphological features, desmoplastic stroma seems to exert a protective effect in patients with iCC. Stromal collagen reticulation may provide additional clinically relevant information. In addition, these data support the potential value to evaluate CRI in biopsy specimen.

Although experimental evidence has suggested that the polymer brush border (PBB) on the cartilage surface is important in regulating fluid permeability in the contact gap, the current theoretical understanding of joint lubrication is still limited. To address this research gap, a multiscale cartilage contact model that includes PBB, in particular its effect on the fluid permeability of the contact gap, is developed in this study. Microscale modeling is employed to estimate the permeability of the contact gap. This permeability is classified into two categories: For a gap size > 1 µm, the flow resistance is assumed to be dominated by the cartilage roughness; for gap size < 1 µm, flow resistance is assumed to be dominated by the surface polymers extending beyond the collagen network of the articular cartilage. For gap sizes of less than 1 µm, the gap permeability decreases exponentially with increasing aggrecan concentration, whereas the aggrecan concentration varies inversely with the gap size. Subsequently, the gap permeability is employed in a macroscale cartilage contact model, in which both the contact gap space and articular cartilage are modeled as two interacting poroelastic systems. The fluid exchange between these two media is achieved by imposing pressure and normal flux continuity boundary conditions. The model results suggest that PBB can substantially enhance cartilage lubrication by increasing the gap fluid load support (e.g., by 26 times after a 20-min indentation compared with the test model without a PBB). Additionally, the fluid flow resistance of PBB sustains the cartilage interstitial fluid pressure for a relatively long period, and hence reduces the vertical deformation of the tissue. Furthermore, it can be inferred that a reduction in the PBB thickness impairs cartilage lubrication ability.

The aim of this study was to characterize multiscale interactions between high intensity focused ultrasound (HIFU) and dentin collagen and associated matrix-metalloproteinases, in addition to the analysis of the effect of HIFU on bacterial biofilms and biological properties. Dentin specimens were subjected to 5, 10 or 20 s HIFU. XPS spectra were acquired and TEM was performed on dentin slabs. Collagen orientation was performed using Raman spectroscopy. Calcium measurements in human dental pulpal cells (hDPCs) were carried out after 7 and 14 days. For macrophages, CD36+ and CD163+ were analysed. Biofilms were analyzed using CLSM. Tandem mass spectroscopy was performed for the detection of hydroxyproline sequences along with human MMP-2 quantification. Phosphorus, calcium, and nitrogen were detected in HIFU specimens. TEM images demonstrated the collagen network appearing to be fused together in the HIFU 10 and 20 s specimens. The band associated with 960 cm-1 corresponds to the stretching ν1 PO43-. The control specimens showed intensive calcium staining followed by HIFU 20 s > HIFU 10 s > HIFU 5 s specimens. Macrophages in the HIFU specimens co-expressed CD80+ and CD163+ cells. CLSM images showed the HIFU treatment inhibiting bacterial growth. SiteScore propensity determined the effect of HIFU on the binding site with a higher DScore representing better site exposure on MMPs. Multiscale mapping of dentin collagen after HIFU treatment showed no deleterious alterations on the organic structure of dentin.

Lack of experimental human models hinders research on Lassa hemorrhagic fever and the development of treatment strategies. Here, we report the first chip‐based model for Lassa hemorrhagic syndrome. The chip features a microvessel interfacing collagen network as a simple mimic for extracellular matrix, allowing for quantitative and real‐time vascular integrity assessment. Luminal infusion of Lassa virus‐like particles led to a dramatic increase in vascular permeability in a viral load‐dependent manner. Using this platform, we showed that Fibrin‐derived peptide FX06 can be used to suppress the vascular integrity loss. This simple chip‐based model proved promising in the assessment of disease severity and provides an easy‐to‐use platform for future investigation of Lassa pathogenesis and drug development in a human‐like setting., An organ‐on‐a‐chip model that imitates Lassa hemorrhagic shock syndrome was developed and used to test drug candidate. The chip is composed of a microvessel interfacing collagen network as a simple mimic for extracellular matrix, which can recapitulate the physiological barrier function of a natural vessel and luminal infusion of Lassa virus‐like particles led to a dramatic increase in vascular permeability. Using this platform, a potential drug, Fibrin‐derived peptide FX06, shows its ability to suppress the Lassa‐induced vascular integrity loss.

Skin integrity and function depends to a large extent on the composition of the extracellular matrix, which regulates tissue organization. Collagen XII is a homotrimer with short collagenous domains that confer binding to the surface of collagen I-containing fibrils and extended flexible arms, which bind to non-collagenous matrix components. Thereby, collagen XII helps to maintain collagen suprastructure and to absorb stress. Mutant or absent collagen XII leads to reduced muscle and bone strength and lax skin, whereas increased collagen XII amounts are observed in tumor stroma, scarring and fibrosis. This study aimed at uncovering in vivo mechanisms by which collagen XII may achieve these contrasting outcomes. We analyzed skin as a model tissue that contains abundant fibrils, composed of collagen I, III and V with collagen XII decorating their surface, and which is subject to mechanical stress. The impact of different collagen XII levels was investigated in collagen XII-deficient (Col12-KO) mice and in mice with collagen XII overexpression in the dermis (Col12-OE). Unchallenged skin of these mice was histologically inconspicuous, but at the ultrastructural level revealed distinct aberrations in collagen network suprastructure. Repair of excisional wounds deviated from controls in both models by delayed healing kinetics, which was, however, caused by completely different mechanisms in the two mouse lines. The disorganized matrix in Col12-KO wounds failed to properly sequester TGFβ, resulting in elevated numbers of myofibroblasts. These are, however, unable to contract and remodel the collagen XII-deficient matrix. Excess of collagen XII, in contrast, promotes persistence of M1-like macrophages in the wound bed, thereby stalling the wounds in an early inflammatory stage of the repair process and delaying healing. Taken together, we demonstrate that collagen XII is a key component that assists in orchestrating proper skin matrix structure, controls growth factor availability and regulates cellular composition and function. Together, these functions are pivotal for re-establishing homeostasis after injury.

Background In vivo confocal Raman spectroscopy (CRS) revealed a clear correlation of age and dermal water content, indicating increasing water content of the dermis with increasing age. This enhancement of water has been interpreted as an age-dependent depletion, of proteins, mainly of collagen. Chronical sun exposure is known to destroy the collagen network of the skin, which leads to the signs of photoaging as the formation of wrinkles. Noninvasive in vivo measuring techniques for photoaging are limited. Therefore, sensitive techniques to quantify even mild degrees of photoaging in a clinical setting are of high interest. We used CRS to measure the water content in human dermis in vivo, assuming that additionally to the increase of water content in intrinsic aging, photoaging would lead to further collagen depletion and an additional increase in water content of the dermis. Materials and methods A panel of 24 female subjects of different age-groups and scores of photoaging was recruited. A ranking of high resolution dorsal forearm photographs was used to classify the degree of photoaging with high precision. After that, CRS water content and collagen measurements were performed in the photoexposed dorsal as well as the photoprotected volar dermis of the subjects. Results A positive correlation of water content in the dermis with age could be confirmed (r = .550). Further, a positive correlation between water content of dorsal dermis and photoaging ranks was observed (Pearson's r = .417). Conclusion Assessment of water content in the dermis with confocal Raman spectroscopy was found to be a promising method to measure the degree of photoaging in human subjects in vivo.

The pro-fibrotic milieu, as the result of the extracellular matrix remodeling, is a central feature in the pathophysiology of heart disease and contributes to its high morbidity and mortality. Fibrosis is a recognized risk factor for development of heart failure and arrythmias and is usually detected by cardiac magnetic resonance or endomyocardial biopsy. Collagen type I and type III are major components of the collagen network, and the assessment of their derived biomarkers could serve as estimate of the myocardial fibrotic content. This review summarizes data from numerous studies in which these biomarkers have proven their diagnostic and prognostic utility, setting the stage for further randomized clinical trials that might translate into early implementation of antifibrotic therapies.

Background Purple tulip extract is a rich source of flavonoids which are powerful antioxidants and can hence be considered as an ideal candidate for use in skin care products. Objectives We aimed to evaluate the effects of purple tulip extract on skin quality and to determine its molecular modes of interaction. Methods A pangenomic study on human skin fibroblasts was carried out to analyse multiple changes in gene expression. Ex vivo studies of human skin explants exposed to ultraviolet (UV) irradiation or H2 O2 were performed to assess modulations of protein expression. Finally, a clinical assay was carried out to evaluate the efficacy of purple tulip extract on skin appearance and condition of aged women. Results Genetic modulation analyses led us to infer the induction of many biological functions including cell differentiation, proliferation, migration, inflammatory responses, and matrix remodelling. The ex vivo studies revealed an enhancement of the collagen network and increased expression of glycosaminoglycans (GAG), fibronectin and collagen VI. Finally, the clinical study highlighted the potential anti-ageing properties of the purple tulip extract which decreased the relaxation of the oval face and improved skin elasticity after 28 days of treatment. Significant reductions of the length and depth of the nasolabial wrinkles were also observed. Conclusion Our genomics data on the effect of purple tulip extract on the ex vivo UV challenged skin, showed that genes responsible for, amongst others, the upkeep of the skin, such as collagen induction, immune cell proliferation and epidermal repair, were all upregulated. More importantly, the clinical study corroborated these data by the visible and measurable effects of the topical purple tulip extract on the aged skin of 22 women, further demonstrating the beneficial impact of the extract on aged skin.

Articular cartilage plays an important role in synovial joint function, but this function is diminished when cartilage tissue breaks down in osteoarthritis. Tissue engineering is a promising approach for replacing failed cartilage, as cartilage is a relatively simple tissue with no blood supply and very few biological cells. To imitate the structure of natural cartilage extracellular matrix material, three components must be included: the hydrated ground substance, the charges that contribute to compressive stiffness via electrostatic repulsion, and the nanofibrous collagen network that resists tensile deformation and failure. Here, the nanofiber network is considered, with examination of its fracture behavior in an as-electrospun state and following a mild chemical crosslinking process. Mode III fracture testing was used to quantify the tear toughness of the fibrous mats, and failure behavior was qualitatively examined with scanning electron microscopy. In ongoing work, this nanofibrous structure will be combined with a charged polyelectrolyte hydrogel gel to create a biomimetic cartilage-like material. By using biomimicry to replicate what is present in native cartilage tissue, a superior material can be designed and fabricated for use in tissue repair and replacement.

Over 1 million burn injuries are treated annually in the United States, and current tissue engineered skin fails to meet the need for full-thickness replacement. Bioprinting technology has allowed fabrication of full-thickness skin and has demonstrated the ability to close full-thickness wounds. However, analysis of collagen remodeling in wounds treated with bioprinted skin has not been reported. The purpose of this study is to demonstrate the utility of bioprinted skin for epidermal barrier formation and normal collagen remodeling in full-thickness wounds. Human keratinocytes, melanocytes, fibroblasts, dermal microvascular endothelial cells, follicle dermal papilla cells, and adipocytes were suspended in fibrinogen bioink and bioprinted to form a tri-layer skin structure. Bioprinted skin was implanted onto 2.5 × 2.5 cm full-thickness excisional wounds on athymic mice, compared with wounds treated with hydrogel only or untreated wounds. Total wound closure, epithelialization, and contraction were quantified, and skin samples were harvested at 21 days for histology. Picrosirius red staining was used to quantify collagen fiber orientation, length, and width. Immunohistochemical (IHC) staining was performed to confirm epidermal barrier formation, dermal maturation, vascularity, and human cell integration. All bioprinted skin treated wounds closed by day 21, compared with open control wounds. Wound closure in bioprinted skin treated wounds was primarily due to epithelialization. In contrast, control hydrogel and untreated groups had sparse wound coverage and incomplete closure driven primarily by contraction. Picrosirius red staining confirmed a normal basket weave collagen organization in bioprinted skin-treated wounds compared with parallel collagen fibers in hydrogel only and untreated wounds. IHC staining at day 21 demonstrated the presence of human cells in the regenerated dermis, the formation of a stratified epidermis, dermal maturation, and blood vessel formation in bioprinted skin, none of which was present in control hydrogel treated wounds. Bioprinted skin accelerated full-thickness wound closure by promoting epidermal barrier formation, without increasing contraction. This healing process is associated with human cells from the bioprinted skin laying down a healthy, basket-weave collagen network. The remodeled skin is phenotypically similar to human skin and composed of a composite of graft and infiltrating host cells. IMPACT STATEMENT: We have demonstrated the ability of bioprinted skin to enhance closure of full-thickness wounds through epithelialization and normal collagen remodeling. To our knowledge, this article is the first to quantify collagen remodeling by bioprinted skin in full-thickness wounds. Our methods and results can be used to guide further investigation of collagen remodeling by tissue engineered skin products to improve ongoing and future bioprinting skin studies. Ultimately, our skin bioprinting technology could translate into a new treatment for full-thickness wounds in human patients with the ability to recapitulate normal collagen remodeling in full-thickness wounds.

Extracellular matrix microstructure and mechanics are crucial to breast cancer progression and invasion into surrounding tissues. The peritumor collagen network is often dense and aligned, features which in vitro models lack. Aspiration of collagen hydrogels led to densification and alignment of microstructure surrounding embedded cancer cells. Two metastasis-derived breast cancer cell lines, MDA-MB-231 and MCF-7, were cultured in initially 4 mg/ml collagen gels for 3 days after aspiration, as well as in unaspirated control hydrogels. Videomicroscopy during aspiration, and at 0, 1, and 3 days after aspiration, epifluorescence microscopy of phalloidin-stained F-actin cytoskeleton, histological sections, and soluble metabolic byproducts from constructs were collected to characterize effects on the embedded cell morphology, the collagen network microstructure, and proliferation. Breast cancer cells remained viable after aspiration-ejection, proliferating slightly less than in unaspirated gels. Furthermore, MDA-MB-231 cells appear to partially relax the collagen network and lose alignment 3 days after aspiration. Aspiration-ejection generated aligned, compact collagen network microstructure with immediate cell co-orientation and higher cell number density apparently through purely physical means, though cell-collagen contact guidance and network remodeling influence cell organization and collagen network microstructure during subsequent culture. This study establishes a platform to determine the effects of collagen density and alignment on cancer cell behavior, with translational potential for anticancer drug screening in a biomimetic three-dimensional matrix microenvironment, or implantation in preclinical models.

Collagen is the most abundant secreted protein in vertebrates and persists throughout life without renewal. The permanency of collagen networks contrasts with both the continued synthesis of collagen throughout adulthood and the conventional transcriptional/translational homeostatic mechanisms that replace damaged proteins with new copies. Here, we show circadian clock regulation of endoplasmic reticulum-to-plasma membrane procollagen transport by the sequential rhythmic expression of SEC61, TANGO1, PDE4D and VPS33B. The result is nocturnal procollagen synthesis and daytime collagen fibril assembly in mice. Rhythmic collagen degradation by CTSK maintains collagen homeostasis. This circadian cycle of collagen synthesis and degradation affects a pool of newly synthesized collagen, while maintaining the persistent collagen network. Disabling the circadian clock causes abnormal collagen fibrils and collagen accumulation, which are reduced in vitro by the NR1D1 and CRY1/2 agonists SR9009 and KL001, respectively. In conclusion, our study has identified a circadian clock mechanism of protein homeostasis wherein a sacrificial pool of collagen maintains tissue function.

Purpose 3-Tesla magnetic resonance imaging (MRI) T1 and T2 mapping to detect and quantify cartilage matrix and meniscal degeneration between normal healthy volunteers and early osteoarthritis patients. Material and methods A prospective study including 25 patients and 10 healthy volunteers was done. Patients with symptoms of early osteoarthritis and Kellgren-Lawrence grade I-II on plain radiograph were included for MRI knee. Patients with inflammatory arthritis, infection, trauma, and history of knee surgery were excluded. Healthy, normal adult volunteers (preferably age and sex matched) without symptoms of osteoarthritis of the knee were drawn from patient's relatives/hospital employees/colleagues for MRI knee. Results T1 and T2 relaxation time values of articular cartilage and menisci were significantly higher in osteoarthritis patients as compared to healthy volunteers. No significant difference was found in morphological thickness of articular cartilage and menisci in early osteoarthritis patients and healthy volunteers. Conclusions T1 and T2 mapping are noninvasive MRI techniques reflecting changes in the biochemical composition of cartilage and menisci. T1 values reflect changes in proteoglycan content, and T2 values are sensitive to interaction between water molecules and collagen network. Mapping techniques assess early cartilage and meniscal matrix degeneration in osteoarthritis of the knee, and help in initiating treatment and monitoring disease progression. MRI is a sensitive modality for assessment of pathological changes in articular cartilage. With use of T1 and T2 mapping techniques, it is possible to evaluate the collagen network and proteoglycan content in articular cartilage and meniscal matrix.

Cardiac fibrosis is a heterogeneous disease, which is characterized by abundant proliferation of interstitial collagen, disordered arrangement, collagen network reconstruction, increased cardiac stiffness, and decreased systolic and diastolic functions, consequently developing into cardiac insufficiency. With several factors participating in and regulating the occurrence and development of cardiac fibrosis, a complex molecular mechanism underlies the disease. Moreover, cardiac fibrosis is closely related to hypertension, myocardial infarction, viral myocarditis, atherosclerosis, and diabetes, which can lead to serious complications such as heart failure, arrhythmia, and sudden cardiac death, thus seriously threatening human life and health. Resveratrol, with the chemical name 3,5,4′-trihydroxy-trans-stilbene, is a polyphenol abundantly present in grapes and red wine. It is known to prevent the occurrence and development of cardiovascular diseases. In addition, it may resist cardiac fibrosis through a variety of growth factors, cytokines, and several cell signaling pathways, thus exerting a protective effect on the heart.

Successful cartilage engineering requires the generation of biological grafts mimicking the structure, composition and mechanical behaviour of the native tissue. Here melt-electrowriting (MEW) was used to produce arrays of polymeric structures whose function was to orient the growth of cellular aggregates spontaneously generated within these structures, and to provide tensile reinforcement to the resulting tissues. Inkjeting was used to deposit defined numbers of cells into MEW structures, which self-assembled into an organized array of spheroids within hours, ultimately generating a hybrid tissue that was hyaline-like in composition. Structurally, the engineered cartilage mimicked the histotypical organization observed in skeletally immature synovial joints. This biofabrication framework was then used to generate scaled-up (50mm × 50mm) cartilage implants containing over 3,500 cellular aggregates in under 15 minutes. After 8 weeks in culture, a 50-fold increase in the compressive properties of these MEW reinforced tissues were observed, while the tensile properties were still dominated by the polymer network, resulting in a composite construct demonstrating tension-compression nonlinearity mimetic of the native tissue. Helium ion microscopy further demonstrated the development of an arcading collagen network within the engineered tissue. This hybrid bioprinting strategy provides a versatile and scalable approach to engineer cartilage biomimetic grafts for biological joint resurfacing.