Collagen Research Articles

Knockout of TMEM206 in mice associated with a loss of corneal transparency

AIM: To investigate the role of transmembrane protein 206 (TMEM206) in corneal edema in mice by knockout the TMEM206 gene using CRISPR/Cas9 editing technology. METHODS: TMEM206-knockout mice were generated using the CRISPR-Cas9 system. Variations in ophthalmic pathology were observed using slit lamp microscope and optical coherence tomography (OCT), intraocular pressure (IOP) was measured using a TonoLab Rebound Tonometer, and the ultrastructure of the corneal was observed using a transmission electron microscope. RESULTS: Corneal opacity was observed in 4/18 homozygous TMEM206-/- mice whereas a similar change was not observed in heterozygous TMEM206+/- mice and wild-type littermates. OCT examination showed that the mean central cornea thickness was 125±5.4 µm in 4 homozygous TMEM206-/- mice developed corneal edema and 115±1.2 µm in wild-type mice (t=3.468, P<0.05) at 43wk. The mean IOP was 12.08±0.07 mm Hg in four right eyes with corneal edema and 12.03±0.03 mm Hg in three normal left eyes (P>0.05). Transmission electron microscopy revealed a disruption in the organization of the collagen fibrils in the central part of the cornea in homozygous TMEM206-/- mice. CONCLUSION: TMEM206 is associated with corneal edema which caused organizational disruption of collagen fibrils in corneas of mice.

Identification of trait-associated microRNA modules in liver transcriptome of pig fed with PUFAs-enriched supplementary diet.

Dietary lipids provide energy, are cellular structural components, and are involved in physiological processes. Lipids are the dietary source in supplementary diet experiments in pigs. This study aims to investigate the dietary effects of PUFAs on the hepatic transcriptome and physiological pathways of two diets on two pig breeds. Polish Landrace (PL: n = 6) and six PLxDuroc (PLxD: n = 6) pigs were fed with a normal diet (n = 3) or PUFAs-enriched healthy diet (n = 3), and the hepatic miRNA profiles were studied for weighted gene co-expression network analysis biological interactions between gene networks and metabolic pathways of DE miRNA genes. The study identified trait-associated modules that were significantly associated with four phenotypic traits in the dietary groups of PL and PLxD: meat colour (a*), shoulder subcutaneous fat thickness, conductivity 24h post-mortem (PE24), and ashes. Trait-wise, a large set of co-expressed miRNAs of porcine liver were identified in these trait-associated significant modules (9, 7, 2, and 8) in PL and PLxD. Each module is represented by a module eigengene (ME). Forty-four miRNAs out of 94 miRNAs interacted with 6719 statistically significant target genes with a target score > 90. The GO/pathway analysis showed association with pathways including regulation of metallopeptidase activity, sebaceous gland development, collagen fibril organization, WNT signalling, epithelial tube morphogenesis, etc. The study showed the differences in miRNA expression between the dietary groups of PL and PLxD breeds. Hub genes of discovered miRNA clusters can be considered predicted miRNA genes associated with PE24, meat colour, shoulder subcutaneous fat thickness, and ashes. Discovered target genes for miRNA clusters play significant roles in biological functions such as (i) muscle and body growth development, (ii) different cellular processes and developments, (iii) system development, and (iv) metabolic processes.

Nanotechnology-Enhanced Orthopaedic Surgery

Nanomaterials hold significant promise for the future of orthopaedic implants due to their ability to mimic the nanoscale components of the bone, such as collagen fibrils and hydroxyapatite. Nanomaterials can regulate cell behaviour while offering mechanical strength and biocompatibility, making them ideal for bone repair and tissue regeneration. This comprehensive review explores the key existing and potential applications of nanotechnology in orthopaedics, including bone tissue engineering, drug delivery systems, systems combatting implant-related infections, and the surface preparation of implants to enhance osseointegration. These innovations are poised to revolutionise orthopaedic care by improving implant durability, reducing infection risks, and promoting bone regeneration to deliver personalised treatment and create better patient outcomes.

Nonlinear Stress-Induced Transformations in Collagen Fibrillar Organization, Disorder and Strain Mechanisms in the Bone-Cartilage Unit.

By developing a 3D X-ray modeling and spatially correlative imaging method for fibrous collagenous tissues, this study provides a comprehensive mapping of nanoscale deformation in the collagen fibril network across the intact bone-cartilage unit (BCU), whose healthy functioning is critical for joint function and preventing degeneration. Extracting the 3D fibril structure from 2D small-angle X-ray scattering before and during physiological compression reveals of dominant deformation modes, including crystallinity transitions, lateral fibril compression, and reorientation, which vary in a coupled, nonlinear, and correlated manner across the cartilage-bone interface. A distinct intermolecular arrangement of collagen molecules, and enhanced molecular-level disorder, is found in the cartilage (sliding) surface region. Just below, fibrils accommodate tissue strain by reorientation, which transitions molecular-level kinking or loss of crystallinity in the deep zone. Crystalline fibrils laterally shrink far more (20×) than they contract, possibly by water loss from between tropocollagen molecules. With the calcified plate acting as an anchor for surrounding tissue, a qualitative switch occurs in fibrillar deformation between the articular cartilage and calcified regions. These findings significantly advance this understanding of the complex, nonlinear ultrastructural dynamics at this critical interface, and opens avenues for developing targeted diagnostic and therapeutic strategies for musculoskeletal disorders.

Comparative ultrastructural and transcriptomic profile analysis of skin tissues from indigenous, improved meat, and dairy goat breeds.

High-quality goatskins are valuable byproducts usually produced by indigenous goat breeds with poorer production performance in Asia and Africa. However, the genetic and molecular mechanisms underpinning goatskin's biomechanical properties (e.g., tensile strength) remain elusive. Mechanistic exploration of these traits could greatly aid the genetic improvement and genetic resource conservation of native breeds in these regions. To fulfill this purpose, we collected skin tissues from three goat breeds: Huai goat (HG), a Chinese native variety producing high-quality goatskins; Yudong meat goat (YDMG), a crossbreed of HG and Boer goat; Henan dairy goat (HNDG), a dairy goat breed. Scanning electronic microscopy analysis of skin tissues found that the collagen fiber diameters, collagen fibril diameters, and crimps significantly differed among the three goat breeds; however, collagen fibril diameters are similar in HG and HNDG. A sum of 230, 775, and 86 differentially expressed genes (DEGs) were identified from YDMG versus HNDG, HG versus HNDG, and YDMG versus HG, respectively. Functional enrichment analysis suggested that signaling pathways involved in fatty acid, retinol, steroid metabolisms, and GO items related to the physical properties of the skin (e.g., collagen-containing extracellular matrix) are significantly overrepresented in DEGs identified from meat versus dairy goats. Furthermore, 106 DEGs (e.g., COL1A1, COL1A2, and SPARC) showed specific expression patterns in HG and YDMG versus HNDG. Items about biophysical features of skin (e.g., extracellular matrix organization and ECM proteoglycans) are markedly enriched. Protein-protein interaction analysis suggested that two growth factors (IGF1 and PDGFD) are latent collagen and other ECM protein expression modulators. Ultrastructural analysis of goat skin tissues suggested that collagen fibril diameter is not a major factor affecting goatskin quality. Transcriptomic profiles unveiled core genes and associated biological processes potentially involved in regulating goatskin quality. These discoveries shined new light on deeper understanding the mechanisms of hide-related traits in goat and other livestock.

Sex Differences in Response to Diet Enriched with Glutathione Precursors in the Aging Heart.

Common features of the aging heart are dysregulated metabolism, inflammation, and fibrosis. Elevated oxidative stress is another hallmark of cardiac aging that can exacerbate each of these conditions. We hypothesize that by increasing natural antioxidant levels (glutathione), we will improve cardiac function. Twenty-one-month-old mice were fed Glycine and N-Acetyl Cysteine (GlyNAC) (glutathione precursors)-supplemented or control diets for 12 weeks. Heart function was monitored longitudinally, and the exercise performance was determined at the end of the study. We found that the GlyNAC diet was beneficial for old male but not old female mice, leading to an increase of Ndufb8 expression (a subunit of the mitochondrial respiratory chain complex), and higher enzymatic activity for CPT1b and CrAT, two carnitine acyltransferases that are critical to cardiomyocyte metabolism. Although no quantifiable change of collagen turnover was detected, hearts from GlyNAC-fed old males exhibited a slight but significant enrichment in Fmod, a protein that can inhibit collagen fibril formation, possibly reducing extracellular matrix (ECM) stiffness and thus improving diastolic function. Cardiac diastolic function was modestly improved in males but not females, and surprisingly GlyNAC-fed female mice showed a decline in exercise performance. In summary, our work supports the concept that aged male and female hearts are phenotypically different. These basic differences may affect the response to pharmacological and diet interventions, including antioxidants.

Restoration of Corneal Stiffness in Rabbits Following Withdrawal of Travoprost.

To evaluate if the effect of travoprost on corneal material stiffness could be restored after drug withdrawal. Seventy-two rabbits were randomly allocated into three groups: medicine (M), medicine withdrawal (MW), and blank (B). Within the M and MW groups, treatment with travoprost was administered to the right eyes (MT and MWT) over a period of 12 weeks. Subsequently, the M group was killed, but the MW group underwent an additional 12-week period for treatment withdrawal. No treatment was given to the contralateral eyes (MC and MWC) in the M and MW groups. A separate blank control (BC) group remained untreated for the entire 24-week duration. In each group, corneas from 18 rabbits were tested mechanically under inflation conditions to estimate their tangent modulus (Et). The corneas of the remaining six rabbits underwent electron microscopy analysis, which focused on fibril diameter and interfibrillar spacing. Central corneal thickness (CCT) of the treated eyes (MT and MWT groups) decreased with 12 weeks of travoprost treatment (P < 0.05). The CCT in the MWT group increased after 12 weeks of withdrawal but was still lower than that in the BC group (P < 0.05). The Et of the MT group was significantly lower than that of the MC group at mean tissue stresses of 2, 4, and 6 kPa (P < 0.05). Conversely, no significant difference in Et values was observed between the MWT, MWC, and BC groups, indicating recovery after treatment cessation. Furthermore, the stromal interfibrillar spacing of the treated MT group was significantly larger (P < 0.05) than that of the control MC group, but no disparity was noted among the MWT, MWC, and BC groups following treatment withdrawal. Additionally, there were no significant differences in the mean diameter of collagen fibrils among all groups (all P > 0.05). Travoprost treatment appears to soften corneal tissue, decrease tissue thickness, and reduce the density of stromal collagen fibers by increasing the interfibrillar spacing. These changes were partially reversed after treatment cessation. Travoprost could further inhibit corneal growth, so its use in childhood and adolescence should be carefully considered. Additionally, the effect of travoprost in reducing corneal stiffness may lead to underestimations of intraocular pressure (IOP) measurement and hence overestimations in the effect of treatment in lowering IOP.

Electrospun hydrophilic PAN/CO/TA composite nanofibrous membrane for adsorbing Cu(II) in water

In this paper, a novel polyacrylonitrile (PAN)/collagen (CO)/tannic acid (TA) composite nanofiber membrane for the adsorption of Cu(II) in water was prepared. The nanofibrous membranes were characterized by Fourier transform infrared (FTIR), scanning electron microscopy (SEM), thermogravimetric (TG), and water contact angle (WCA) techniques, and their adsorption performance on Cu(II) in water was tested. The results showed that incorporating CO improved the hydrophilic performance of the membranes, and the contact angle of PAN/CO decreased from 60.57° to 39.16° with the increase of CO content, which exhibited good hydrophilicity. Moreover, CO effectively crosslinked TA, so that TA was well fixed on the surface of the fiber membrane, and the resulting PAN/CO/TA composite nanofiber membrane had good fiber morphology, uniform fiber diameter distribution, with an average diameter of 292.22 nm, and good adsorption performance for Cu(II), up to 133.26 mg/g. The adsorption kinetics fitting showed that the adsorption mechanism was mainly electrostatic adsorption and chelation of Cu(II) by phenoxy anion. PAN/CO/TA and PAN/CO/TA/Cu nanofiber membranes showed bacteriostatic effects on E. coli and S. aureus, with PAN/CO/TA/Cu nanofiber membranes being particularly effective, with the average inhibition bands for E. coli and S. aureus being 7.50 mm and 10.05 mm, respectively. The distribution of the electric field during the spinning process was also simulated by finite element analysis in this study. Since TA is a natural polymer of plant origin and CO is of animal origin, it provides an environmentally friendly and cost-effective method to remove Cu(II) from water.

Challenges of the minimum postmortem interval (PMImin) estimation in forensic scenarios: Is dental histology an alternative?

Postmortem interval (PMI) estimates the time since death. Teeth are perennial elements capable of remaining intact in taphonomic environmental circumstances. The objective was to evaluate the feasibility of estimating the minimum postmortem interval (PMImin) through histological analyses of dental tissues exposed to burial and drowning conditions, simulating common scenarios in forensic practice. A total of n = 99 teeth were analyzed and divided into four groups: control (T0), one month (T1), three months (T2), and six months (T3). The control sample comprised 10 teeth, while T1, T2 e T3 were divided into three different subgroups: controlled environment, buried, and drowned. For each subgroup, ten samples were used. Following exposure to taphonomic conditions, the specimens were processed, and histological sections were obtained. The two-way ANOVA test and the Tukey’s post-hoc test were employed for the quantitative analysis of dentin collagen fibrils, revealing statistically significant differences (α = 5 %). This allowed for the estimation of the PMImin at three months by observing pixel counts exceeding 13e+05 in drowned teeth and greater than 8e+05 in buried teeth. Qualitative analysis revealed that the PMImin of drowned teeth was estimated at one month due to the absence of the periodontal ligament (PDL) and at six months due to the absence of predentin and partial degradation of the cementum. For buried teeth, the three-month PMImin was indicated by the absence of PDL and partial cementum degradation. The absence of pulp and remnants of predentin characterized the six-month PMImin. Qualitative and quantitative histological characteristics and parameters are potential to estimate PMImin in forensic scenarios spanning up to six months.

Endochondral ossification: insights into the cartilage mineralization processes achieved by an anhydrous freeze substitution protocol

Growth plate cartilage (GP) serves as a dynamic site of active mineralization and offers a unique opportunity to investigate the cell-regulated matrix mineralization process. Transmission electron microscopy (TEM) provides a means for the direct observation of these mechanisms, offering the necessary resolution and chemical analysis capabilities. However, as mineral crystallinity is prone to artifacts using aqueous fixation protocols, sample preparation techniques are critical to preserve the mineralized tissue in its native form. We optimized cryofixation by high-pressure freezing followed by freeze substitution in anhydrous acetone containing 0.5% uranyl acetate to prepare murine GP for TEM analysis. This sample preparation workflow maintains cellular and extracellular protein structural integrity with sufficient contrast for observation and without compromising mineral crystallinity. By employing appropriate sample preparation techniques, we were able to observe two parallel mineralization processes driven by chondrocytes: 1) intracellular- and 2) extracellular-originating mineralized vesicles. Both mechanisms are based on sequestering calcium phosphate (CaP) within a membrane-limited structure, albeit originating from different compartments of the chondrocytes. In the intracellular originating pathway, CaP accumulates within mitochondria as globular CaP granules, which are incorporated into intracellular vesicles (500-1000 nm) and transported as granules to the extracellular matrix (ECM). In contrast, membrane budding vesicles with a size of approximately 100-200nm, filled with needle-shaped minerals were observed only in the ECM. Both processes transport CaP to the collagenous matrix via vesicles, they can be differentiated based on the vesicle size and mineral morphologies. Their individual importance to the cartilage mineralization process is yet to be determined. Statement of SignificanceWe do not fully understand the process by which epiphyseal cartilage mineralizes - a vital step in endochondral bone formation. Previous work has proposed that mitochondria and intracellular vesicles are storage sites for the delivery of mineral to collagen fibrils. However, these concepts are founded on results from in vitro models of mineralization; no prior work has observed mineral-containing intracellular vesicles or mitochondria in developing epiphyseal cartilage. Here we developed a new cryofixation preparation route for transmission electron microscopy (TEM) imaging that has disclosed a cell-regulated process of mineralization in epiphyseal cartilage. High resolution TEM images revealed an involvement of mitochondria and intracellular and extracellular vesicles in delivering transient mineral phases to the collagen fibrils to promote cartilage mineralization.

Induced collagen type-I secretion by hepatocytes of the melanoma liver metastasis is associated with a reduction in tumour-infiltrating lymphocytes.

Overall patients with melanoma liver metastasis (MLiM) have a dismal prognosis and poor responses to the standard of care treatment. Understanding the role of the tumour microenvironment (TME) is critical for discovering better strategies to overcome intrinsic therapy resistance in MLiM. The aim was to understand the crosstalk signalling pathways between hepatocytes and metastatic melanoma cells in the TME of MLiM. Hepatocytes and melanoma tumour cells of MLiM were assessed using transcriptomic NanoString GeoMx digital spatial profiling (NGDSP) assay. Functional assays were performed using normal hepatocytes and MLiM-derived cell lines. Validation was performed using multiplex immunofluorescence. In NGDSP analysis adjacent normal hepatocytes (ANH) had higher CXCR4 and COL1A1/2 levels than distant normal hepatocytes (DNH), while melanoma cells had higher TNF-α levels. In vitro, MLiM cell lines released TNF-α which upregulated CXCR4 and CXCL12 levels in ANH. CXCL12 activated CXCR4, which triggered AKT and NFκB signalling pathways. Consequently, AKT signalling induced the upregulation of collagen type I. MLiM were significantly encircled by a shield of collagen, whereas other liver metastases showed reduced levels of collagen. Of all the liver metastasis analyzed, the presence of collagen in melanoma liver metastasis was associated with a reduction in tumour-infiltrating lymphocytes. MLiM modified ANH to increase collagen production and created a physical barrier. The collagen barrier was associated with a reduction of immune cell infiltration which could potentially deter MLiM immune surveillance and treatment responses. Spatial analyses of melanoma liver metastasis show that adjacent normal hepatocytes have increased collagen-type I levels. Melanoma liver metastases tumour cells secrete enhanced levels of TNF-α to stimulate CXCR4/CXCL12 upregulation in adjacent normal hepatocytes. Activation of CXCR4 promotes AKT and NF-κB signalling pathways to promote collagen-type I secretion in adjacent normal hepatocytes. Elevated collagen levels were associated with reduced tumour-infiltrating lymphocytes.

Ultrastructure and immunohistochemistry of apteric skin in ratites and its epidermal soft cornification

An electron microscopy and immunohistochemistry study has been conducted to acquire comparative information on the structure of apteric skin in ratites, ostrich and emu. The epidermis is thin in the neck of both species and thicker in the dorsal region where acidic and neutral keratins are present in the viable epidermis and stratum corneum. The dermis in both species is mostly occupied by collagen fibrils that form large bundles, often organized in alternated layers in the deeper part of the dermis. Numerous collagen fibrils contact the basement membrane of the epidermis. Sparse tactile Meissner or Krause sensilli are present among the thick collagen bundles. The ostrich epidermis in the dorsal skin is thicker than in the neck, with a columnar basal layer, 3–5 intermediate suprabasal layers and a thick corneous layer. The epidermis of the neck in emu is very thin, featuring two-three narrow cell layers above a flat basal layer and a relatively thick corneous layer. Basal and suprabasal keratinocytes contain lipid droplets and small keratin bundles but no keratohyalin accumulates in pre-corneous cells. The thin corneocytes form a multilayered corneous layer. Loricrine is present in pre-corneous and corneous layers while CBPs, formerly indicated as beta-keratins, are absent in apteric epidermis.

Tailored 3D Agarose-Well Integrated with Human Skin Equivalents for Enhanced Skin Penetration Assessment

We developed a tailored 3D Agarose-well system integrated with reconstructed human skin equivalents to enhance skin penetration assessments. This system addresses common limitations in traditional trans-well reconstructions, such as dermal layer contraction and limited lateral diffusion, by entangling collagen fibrils within the Agarose-well. We evaluated the penetration behavior of three peptides, with and without skin-penetrating peptide (SPP) sequences, alongside adenosine, a known anti-wrinkle agent. Despite a SPP having a molecular weight approximately four times greater than that of adenosine, its kinetic constant was similar, with values of about 39 and 34, respectively. Moreover, this living skin equivalent system not only allowed for the evaluation of adenosine penetration, but also demonstrated its biological effects, with adenosine significantly enhancing procollagen synthesis by approximately 23% compared to the control. Overall, this novel strategy holds the potential for tailoring 3D Agarose-wells and advancing high-performance gel development, making it a promising approach for applications in tissue engineering, medical science, regenerative medicine, and cosmetics.

A unique and biocompatible corneal collagen crosslinking in vivo

Corneal crosslinking (CXL) is a widely applied technique to halt the progression of ectatic diseases through increasing the thickness and mechanical stiffness of the cornea. This study investigated the biocompatibility and efficiency of a novel CXL procedure using ruthenium and blue light in rat corneas and evaluated parameters important for clinical application. To perform the CXL procedure, the corneal epithelium of rats was removed under anaesthesia, followed by the application of a solution containing ruthenium and sodium persulfate (SPS). The corneas were then exposed to blue light at 430 nm at 3 mW/cm2 for 5 min. Rat corneas were examined and evaluated for corneal opacity, corneal and limbal neovascularization, and corneal epithelial regeneration on days 0, 1, 3, 6, 8, and 14. On day 28, the corneas were isolated for subsequent tissue follow-up and analysis. CXL with ruthenium and blue light showed rapid epithelial healing, with 100% regeneration of the corneal epithelium and no corneal opacity on day 6. The ruthenium group also exhibited significantly reduced corneal (p < 0.01) and limbal neovascularization (p < 0.001). Histological analysis revealed no signs of cellular damage or apoptosis, which further confirms the biocompatibility and nontoxicity of our method. Confocal and scanning electron microscopy (SEM) images confirmed high density of collagen fibrils, indicating efficient crosslinking and enhanced structural integrity. This study is unique that demonstrates in vivo safety, biocompatibility, and functionality of ruthenium and blue light CXL. This approach can prevent toxicity caused by UV-A light and can be an immediate alternative compared to the existing crosslinking procedures that have side effects and clinical risks for the patients.

Mechanistic insights on stabilization and destabilization effect of ionic liquids on type I collagen fibrils

Tuned assembly of collagen has tremendous applications in the field of biomedical and tissue engineering owing to its targeted biological functionalities. In this study, ionic liquids choline dihydrogen citrate (CDHC) and diethyl methyl ammonium methane sulfonate (AMS) have been used to regulate the self-assembly of collagen at its physiological pH by probing the assembled systems at certain concentration ratios of ionic liquids and the systems were studied using various characterization methods. Due to interaction with collagen, choline dihydrogen citrate causes delay in the collagen fibrillisation process showing no binding interactions with collagen. In contrast, diethyl methyl ammonium methane sulfonate shows crosslinking effect on collagen fibrillisation due to the electrostatic interaction with the tetrahedral hydration shell of collagen moieties. From rheological studies it was observed that the AMS treated collagen fibril at 1:1 % (w/v) has highest linear viscoelastic range, this can bear the stress under high strain compare to native collagen fibril as well as all CDHC composites. For a sustainable biomaterial or bio-scaffold, mechanical property plays pivotal role on it and from our experimental analysis we found certain composites of ionic liquid treated collagen fibrillar assembly which may act as a sustainable biomaterial or bio-scaffold. It was also evolved that, how the structure-function relationship of ionic force modulated fibrillar assembly controlling the mechanical properties of the tuned system. This self-assembled, ionic-liquid treated collagen-fibrillar system would accelerate various force modulated fibrillar network study, for mimicking the ECM and tissue engineering application.

Self-Assembly of Cyclic-Bending Collagen Fibrils by Polyimide Films.

The cyclic-bending morphologies of the fibrils formed by the self-assembly of type I collagen (Col) are closely related to the mechanisms of various diseases. Therefore, studies that allow the self-assembly of Col molecules to form cyclic-bending fibrils in vitro are vitally important. In this study, we successfully achieved the cyclic-bending shapes (specifically, a regular hexagonal shape) of Col molecules by controlling the steric structures of polyimide (PI) molecular chains through the film formation process. Specifically, when a single layer of PI film was baked, the PI molecular chains within the film bent in the direction parallel to the substrate surface plane. Repeating the layering and baking processes resulted in 3D structures of the PI molecular chains, which were oriented in the direction perpendicular to the substrate surface plane. This three-dimensional bending would result from the PI molecular chain interactions between the upper and lower layers. When the Col molecules were reacted on these film surfaces, they recognized the structures of the PI molecular chains and self-assembled to form cyclic-bending Col fibrils. Especially, in PI films subjected to three cycles of layering and baking, hemicircular-shaped Col fibrils were observed to be regularly arrayed. Additionally, these regularly cyclic-bending fibrils were aligned in the uniaxial direction through a uniaxial rubbing treatment of the PI films. This successful research is significant both as a method for controlling the morphologies of Col fibrils and as a study that explores the biomedical implications of Col fibril cyclic-bending in the living body.

Cellular solids and prestressed affine networks as models of the elastic behavior of soft biological structures.

We reviewed two microstructural models, cellular solid models and prestressed affine network models, that have been used previously in studies of elastic behavior of soft biological materials. These models provide simple and mathematically transparent equations that can be used to interpret experimental data and to obtain quantitative predictions of the elastic properties of biological structures. In both models, volumetric density and elastic properties of the microstructure are key determinants of the macroscopic elastic properties. In the prestressed network model, geometrical rearrangement of the microstructure (kinematic stiffness) is also important. As examples of application of these models, we considered the shear behavior of the cytoskeleton of adherent cells, of the collagen network of articular cartilage, and of the lung parenchymal network since their ability to resist shear is important for their normal biological and physiological functions. All three networks carry a pre-existing stress (prestress). We predicted their shear moduli using the microstructural models and compared those predictions with existing experimental data. Prestressed network models of the cytoskeleton and of the lung parenchyma provided a better correspondence to experimental data than cellular solid models. Both cellular solid and prestressed network models of the cartilage collagen network provided reasonable agreements with experimental values. These findings suggested that the kinematic stiffness and material stiffness of microstructural elements were both important determinants of the shear modulus of the cytoskeleton and of the lung parenchyma, whereas elasticity of collagen fibrils had a predominant role in the cartilage shear behavior.

Accessory Ligament of the Deep Digital Flexor Tendon of the Horse Forelimb and Its Relationship with the Superficial Digital Flexor Tendon: A Plastination, Histological, and Morphometry Study.

The accessory ligament of the deep digital flexor tendon (AL-DDFT) plays a crucial role in the stay apparatus of the horse. This study aimed to investigate the anatomical relationship between the AL-DDFT, the superficial digital flexor tendon (SDFT), and other structures in the metacarpal region. Sixteen distal forelimbs from eight horses, aged 1 to 6 years, were evaluated through macroscopic, microscopic, and morphometric analyses, utilizing detailed dissection, E12 plastinated sections, and histological analysis. During lateral dissection, a connection was observed between the AL-DDFT and the SDFT. Histological evaluation revealed that this connection was a fibrous band (FB), extending the common synovial sheath (CSS) to the SDFT, along with associated collagen fibrils of the epiligament and peritenon. Additionally, two distinct forms of the AL-DDFT were identified, Type I and Type II, with Type II showing a greater cross-sectional area (CSA) than Type I. While numerous morphological and morphometric studies have explored the AL-DDFT and related structures, research incorporating plastination-based morphological and histological evaluations remains scarce. The findings provide valuable insights for both the morphological and clinical assessment of structures within the metacarpal region.

Comparative Histopathological Analysis of Nasal Osteotomy Techniques in a Rabbit Model.

Objective: Nasal osteotomy is a crucial step in rhinoplasty, explicitly addressing the open roof deformity in nasal bones. This study aimed to assess and compare the recovery patterns associated with 4 distinct osteotomy techniques in a rabbit model. Materials and Methods: Twenty white New Zealand rabbits weighing 2.5 to 3.0 kg were included in the study. Median osteotomies were performed on the left side using piezosurgery and on the right side using an oscillating micro-saw. Additionally, internal osteotomy was executed on the right side and external osteotomy on the left side, both performed with a chisel across all rabbits. Ten rabbits were sacrificed on the 7th day, while the remaining 10 were sacrificed on the 15th day for subsequent histological examination. The granulation tissue formation (GTF), vascularization, inflammation, collagen fibril content, osteotomy gap width, and new bone formation were evaluated in hematoxylin and eosin and Masson's trichrome-stained sections. Results: On the 7th day, piezosurgery demonstrated statistically-significant advantages in terms of GTF, neovascularization, and collagen fibril quantity when compared to the oscillating micro-saw (P < .05). Furthermore, a statistically-significant difference was observed among the groups concerning the distribution of connective tissue organization between the 7th and 15th days (P < .05). Conclusions: This study's findings unequivocally establish that bone healing after osteotomy with piezosurgery surpasses that achieved with an oscillating micro-saw in this rabbit model.

Growth Differentiation Factor 5-Induced Mesenchymal Stromal Cells Enhance Tendon Healing.

Mesenchymal stromal cells (MSCs) have immense potential for use in musculoskeletal tissue regeneration; however, there is still a paucity of evidence on the effect of tenogenic MSCs (TMSCs) in tendon healing invivo. This study aimed to determine the effects of growth differentiation factor 5 (GDF5)-induced rabbit MSCs (rbMSCs) on infraspinatus tendon healing in a New Zealand white rabbit model. In this study, bone marrow-derived rbMSCs were isolated, and 100 ng/mL GDF5 was used to induce tenogenic differentiation in rbMSC. The effects of GDF5 on rbMSC in vitro were assessed by total collagen assay, gene expression analysis, and immunofluorescence staining of tenogenic markers; native tenocytes isolated from rabbit tendon were used as a positive control. In invivo, a window defect was created on the infraspinatus tendons bilaterally. After 3 weeks, the rabbits (n = 18) were randomly divided into six groups and repaired with various interventions: (1) surgical suture; (2) fibrin glue (FG); (3) suture and FG; (4) suture, FG, and rabbit tenocytes (rbTenocyte); (5) suture, FG, and rbMSCs, and (6) suture, FG, and TMSC. All animals were euthanized at 6 weeks postoperatively. The in vitro GDF5-induced rbMSCs (or TMSC) showed increased total collagen expression, augmented scleraxis (SCX), and type-I collagen (COL1A1) mRNA gene expression levels. Immunofluorescence showed similar expression in GDF5-induced rbMSC to that of rbTenocyte. In vivo histological analysis showed progressive tendon healing in the TMSC-treated group; cells with elongated nuclei aligned parallel to the collagen fibers, and the collagen fibers were in a more organized orientation, along with macroscopic evidence of tendon callus formation. Significant differences were observed in the cell-treated groups compared with the non-cell-treated groups. Histological scoring showed a significantly enhanced tendon healing in the TMSC- and rbMSC-treated groups compared with the rbTenocyte group. The SCX mRNA expression levels, at 6 weeks following repair, were significantly upregulated in the TMSC group. Immunofluorescence showed COL-1 bundles aligned in parallel orientation; this was further confirmed in atomic force microscopy imaging. SCX, TNC, and TNMD were detected in the TMSC group. In conclusion, GDF5 induces tenogenic differentiation in rbMSCs, and TMSC enhances tendon healing invivo compared with conventional suture repair. Impact Statement Tendon tears and degeneration are debilitating clinical conditions. To date, the suture method is the only gold standard for repairing tendons. Mesenchymal stromal cells (MSCs) have been suggested for many years for their potential in tissue regeneration, especially in tendon-degenerative conditions. Growth differentiation factor 5 (GDF5) has been reported to induce human MSC into a tenogenic lineage (or TMSC), hence a potential cell source for tendon regeneration. This study reported on the potential of rabbit MSC to differentiate into TMSC via GDF5 induction and the potential of TMSC in tendon healing in a New Zealand white rabbit infraspinatus tendon model fulfilled with the 3R principle (reduce, reuse, and replace).