Microplastics (MPs) are environmental contaminants with sizes of the order of less than 5 mm that can enter the human body through inhalation and ingestion. Studies have shown that MPs can pose a threat to human health, and thus evaluation of the presence and potential adverse effects of MPs in tissues is critical. Typical MP studies include enzymatic or chemical tissue digestion that can lead to the loss of some MPs. Moreover, digestion does not allow mapping the location of the contaminant within the tissue architecture. This study aimed to develop a method to evaluate the presence of MPs in histological (thin) sections of tissues without digestion using optical photothermal infrared (O-PTIR) microspectroscopy at sub-micron (500 nm) spatial resolution. Tissue phantoms containing specific amounts and types of MPs and biological tissues were evaluated using polarized light microscopy (PLM) and O-PTIR, and several data analysis approaches were employed to detect MPs in non-digested samples. MPs of sizes from 3 to 85 µm were detected and characterized in tissue phantoms. Furthermore, we detected MPs related to the breakdown products of nylon and cellulose particles in thin sections of biological tissues and discussed obstacles related to the use of database spectra for comparison with O-PTIR spectra, demonstrating the potential of this novel approach and the associated challenges.

Collagen II is a vital structural component in developing bones and mature cartilage. Mutations in this protein cause spondyloepiphyseal dysplasia, a disease characterized primarily by altered skeletal growth and manifesting with a range of phenotypes, from lethal to mild. This study examined transgenic mice harboring the R992C (p.R1124C) substitution in collagen II. Previous research demonstrated significant growth abnormalities and disorganized growth plate structure in these mice, and histological signs of osteoarthritic changes in the knee joints of 9-month-old mice with the R992C mutation. Our study focuses on detecting early structural changes in the articular cartilage that occur before histological signs become apparent. Through microscopic and spectroscopic analyses, we observed significant alterations in the distribution gradients of collagenous proteins and proteoglycans in the cartilage of R992C mutant mice. We propose that these early changes, eventually leading to articular cartilage degeneration in older mice, underscore the progressive nature of osteoarthritic changes linked to collagen II mutations. By identifying these early structural aberrations, our findings emphasize the importance of early detection of osteoarthritic changes, potentially facilitating timely, non-surgical interventions.

You have accessJournal of UrologyInfertility: Basic Research & Pathophysiology (PD53)1 May 2024PD53-01 ARE MICROPLASTICS PRESENT IN HUMAN TESTICLE TISSUE? ANALYSIS USING INFRARED SPECTROSCOPY John M. Masterson, Azita HassanMazandarani, William Querido, Andrzej Steplewski, Yi Zhang, Carissa Huynh, Andrzej Fertala, Nancy Pleshko, and Maurice M. Garcia John M. MastersonJohn M. Masterson , Azita HassanMazandaraniAzita HassanMazandarani , William QueridoWilliam Querido , Andrzej SteplewskiAndrzej Steplewski , Yi ZhangYi Zhang , Carissa HuynhCarissa Huynh , Andrzej FertalaAndrzej Fertala , Nancy PleshkoNancy Pleshko , and Maurice M. GarciaMaurice M. Garcia View All Author Informationhttps://doi.org/10.1097/01.JU.0001009456.01770.91.01AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Environmental microplastics are a consequence of plastic pollution and represent a growing public health concern. Microplastic particles, have been found in human tissues of varying organ systems and have almost uniformly been identified via infrared or Raman spectroscopy. However, little is known about the ability of microplastic particles to infiltrate the male reproductive system. We analyzed male testicle tissue to identify microplastics using standard Fourier-transform infrared (FTIR) spectroscopy followed by high-resolution optical photothermal infrared (O-PTIR) spectroscopy of sectioned tissue. METHODS: Fresh tissue from orchiectomy specimen was paraffin-embedded, sectioned into 5-micron slices, and mounted on low-e microscope slides. Polarizing microscopy was performed using the Nikon Eclipse LV 100N POL light microscope and Perkin Elmer Spotlight 400 imaging spectrometer to identify foreign particles within the tissue. The FTIR spectra of the identified particles were obtained with a resolution of 6.25 µm. These spectra were then compared to entries in a standard Perkin Elmer database. Additionally, spectral data at a spatial resolution of 500 nm were acquired from the same particle using O-PTIR spectroscopy. RESULTS: The comparison of the FTIR spectra with the Perkin Elmer database revealed that the analyzed particle exhibited spectral characteristics matching Azlon (search score 0.67), polyamide resin (search score 0.67), nylon (search score 0.65), and ecteola-modified cellulose (search score 0.65). Refined analyses using O-PTIR demonstrated variations in the spectra from different particle regions, and absorbance peaks were present at 1062, 1166, 1540, and 1650 cm-1 (Figure 1). We propose that broad absorbance centered at∼1062 cm-1 likely indicates modification of the nylon structure, possibly by oxidation. CONCLUSIONS: This is the first report to demonstrate that microplastic materials are indeed able to infiltrate testicular tissue. The clinical consequences of tissue infiltration by microplastics are currently unknown, including whether this phenomenon results in as-yet undescribed clinical harms or benefits. The clinical consequence of this phenomenon remains an area of active research. Download PPT Source of Funding: N/A © 2024 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 211Issue 5SMay 2024Page: e1138 Advertisement Copyright & Permissions© 2024 by American Urological Association Education and Research, Inc.Metrics Author Information John M. Masterson More articles by this author Azita HassanMazandarani More articles by this author William Querido More articles by this author Andrzej Steplewski More articles by this author Yi Zhang More articles by this author Carissa Huynh More articles by this author Andrzej Fertala More articles by this author Nancy Pleshko More articles by this author Maurice M. Garcia More articles by this author Expand All Advertisement PDF downloadLoading ...

Excessive posttraumatic scarring in orthopedic tissues, such as joint capsules, ligaments, tendons, muscles, and peripheral nerves, presents a significant medical problem, resulting in pain, restricted joint mobility, and impaired musculoskeletal function. Current treatments for excessive scarring are often ineffective and require the surgical removal of fibrotic tissue, which can aggravate the problem. The primary component of orthopedic scars is collagen I-rich fibrils. Our research team has developed a monoclonal anti-collagen antibody (ACA) that alleviates posttraumatic scarring by inhibiting collagen fibril formation. We previously established the safety and efficacy of ACA in a rabbit-based arthrofibrosis model. In this study, we evaluate the utility of a well-characterized thermoresponsive hydrogel (THG) as a delivery vehicle for ACA to injury sites. Crucial components of the hydrogel included N-isopropylacrylamide, poly(ethylene glycol) diacrylate, and hyaluronic acid. Our investigation focused on in vitro ACA release kinetics, stability, and activity. Additionally, we examined the antigen-binding characteristics of ACA post-release from the THG in an in vivo context. Our preliminary findings suggest that the THG construct exhibits promise as a delivery platform for antibody-based therapeutics to reduce excessive scarring in orthopedic tissues.

Highly organized collagen fibrils interlacing with proteoglycans form the crucial architecture of the cornea and facilitate its transparency. Corneal scarring from accidental injury, surgery, or infection alters this highly organized tissue, causing severe consequences, including blindness. There are no pharmacological or surgical methods to effectively and safely treat excessive corneal scarring. Thus, we tested the anticorneal scarring utility of a rationally designed anticollagen antibody (ACA) whose antifibrotic effects have already been demonstrated in nonocular models. Utilizing a rabbit model with an incisional corneal wound, we analyzed ACA's effects on forming collagen and proteoglycan-rich extracellular matrices in scar neotissue. We used microscopic and spectroscopic techniques to quantify these components and measure crucial parameters characterizing the structure and organization of collagen fibrils. Moreover, we analyzed the spatial distribution of collagen and proteoglycans in normal and healing corneas. Our study demonstrated significant changes in the quality and quantity of the analyzed molecules synthesized in scar neotissue. It showed that these changes extend beyond incision margins. It also showed ACA's positive impact on some crucial parameters defining proper cornea structure. This pilot study provides a stepping stone for future tests of therapeutic approaches that target corneal extracellular scar matrix assembly.

Purpose: This study aimed to evaluate the utility of a rationally engineered antibody that directly blocks collagen fibrillogenesis to reduce scar tissue formation associated with subconjunctival glaucoma surgery. Material and methods: Fourteen eyes of 7 adult rabbits underwent glaucoma filtering surgery using XEN 45 Gel Stent. The rabbits’ eyes were divided randomly into three treatment groups: (i) treated with the antibody, (ii) treated with mitomycin C, and (iii) treated with the antibody and mitomycin C. Following surgeries, the intraocular pressure and bleb appearance were evaluated in vivo. The rabbits were sacrificed 8 weeks after the surgery, and their eyes were harvested and processed for tissue analysis. Subsequently, tissue samples were analyzed microscopically for fibrotic tissue and cellular markers of inflammation. Moreover, the collagen-rich fibrotic tissue formed around the stents was analyzed using quantitative histology and infrared spectroscopy. The outcomes of this study were analyzed using the ANOVA test. Results: This study demonstrated no significant differences in intraocular pressure, bleb appearance, or presence of complications such as bleb leak among the treatment groups. In contrast, we observed significant differences among the subpopulations of collagen fibrils formed within scar neo-tissue. Based on the spectroscopic analyses, we determined that the relative content of mature collagen cross-links in the antibody-treated group was significantly reduced compared to other groups. Conclusions: Direct blocking of collagen fibrillogenesis with the anti-collagen antibody offers potentially beneficial effects that may reduce the negative impact of the subconjunctival scarring associated with glaucoma filtering surgery.

Dupuytren's disease is a benign fibroproliferative disorder of the hand that results in disabling digital contractures that impair function and diminish the quality of life. The incidence of this disease has been correlated with chronic inflammatory states, but any direct association between inflammatory cytokines and Dupuytren's disease is not known. We hypothesized that advanced fibroproliferation is associated with increased levels of circulating inflammatory cytokines. Blood and fibrotic cord tissue were collected preoperatively from patients with severe contracture and control patients. Blood plasma concentrations of known inflammatory cytokines were evaluated using a multiplex immunoassay. Proteins from the cord tissue were analyzed by RNA sequencing and immunohistochemistry. Moreover, collagen-rich cords were analyzed using Fourier-transform infrared spectroscopy. The results indicate that patients exhibited significantly elevated circulating inflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin (IL)-2, and IL-12p70, as compared with controls. Similarly, IL-4 and IL-13 were detected significantly more frequently in Dupuytren's disease as compared with control. RNA sequencing revealed 5311 differentially expressed genes and distinct clustering between diseased and control samples. In addition to increased expression of genes associated with fibroproliferation, we also observed upregulation of transcripts activated by inflammatory cytokines, including prolactin inducible protein and keratin intermediate filaments. IL-2, but not TNF-α, was detected in fibrotic cord tissue by immunohistochemistry. Finally, spectroscopic assays revealed a significant reduction of the collagen content and alterations of collagen cross-linking within the Dupuytren's disease tissues. In total, our results illustrate that patients with severe Dupuytren's disease exhibit substantially elevated circulating inflammatory cytokines that may drive fibroproliferation. Clinical Significance: The results from this study establish the basis for a specific cytokine profile that may be useful for diagnostic testing and therapeutic intervention in Dupuytren's disease.

Posttraumatic fibrotic scarring is a significant medical problem that alters the proper functioning of injured tissues. Current methods to reduce posttraumatic fibrosis rely on anti-inflammatory and anti-proliferative agents with broad intracellular targets. As a result, their use is not fully effective and may cause unwanted side effects. Our group previously demonstrated that extracellular collagen fibrillogenesis is a valid and specific target to reduce collagen-rich scar buildup. Our previous studies showed that a rationally designed antibody that binds the C-terminal telopeptide of the α2(I) chain involved in the aggregation of collagen molecules limits fibril assembly in vitro and reduces scar formation in vivo . Here, we have utilized a clinically relevant arthrofibrosis model to study the broad mechanisms of the anti-scarring activity of this antibody. Moreover, we analyzed the effects of targeting collagen fibril formation on the quality of healed joint tissues, including the posterior capsule, patellar tendon, and subchondral bone. Our results show that blocking collagen fibrillogenesis not only reduces collagen content in the scar, but also accelerates the remodeling of healing tissues and changes the collagen fibrils’ cross-linking. In total, this study demonstrated that targeting collagen fibrillogenesis to limit arthrofibrosis affects neither the quality of healing of the joint tissues nor disturbs vital tissues and organs.

INTRODUCTION:Posttraumatic scarring of peripheral nerves produces unwanted adhesions that block axonal growth. In the context of surgical nerve repair, the organization of the scar tissue adjacent to conduits used to span the gap between the stumps of transected nerves is poorly understood. The goal of this study was to elucidate the patterns of distribution of collagen-rich scar tissue and analyze the spatial organization of cells that produce fibrotic deposits around and within the conduit's lumen. METHODS:Employing a rabbit model of sciatic nerve transection injury, we studied the formation of collagen-rich scar tissue both inside and outside conduits used to bridge the injury sites. Utilizing quantitative immunohistology and Fourier-transform infrared spectroscopy methods, we measured cellular and structural elements present in the extraneural and the intraneural scar of the proximal and distal nerve fragments. RESULTS:Analysis of cells producing collagen-rich deposits revealed that alpha-smooth muscle actin-positive myofibroblasts were only present in the margins of the stumps. In contrast, heat shock protein 47-positive fibroblasts actively producing collagenous proteins were abundant within the entire scar tissue. The most prominent site of transected sciatic nerves with the highest number of cells actively producing collagen-rich scar was the proximal stump. CONCLUSION:Our findings suggest the proximal region of the injury site plays a prominent role in pro-fibrotic processes associated with the formation of collagen-rich deposits. Moreover, they show that the role of canonical myofibroblasts in peripheral nerve regeneration is limited to wound contracture and that a distinct population of fibroblastic cells produce the collagenous proteins that form scar tissue. As scarring after nerve injury remains a clinical problem with poor outcomes due to incomplete nerve recovery, further elucidation of the cellular and spatial aspects of neural fibrosis will lead to more targeted treatments in the clinical setting.

BACKGROUND:Increased tendon pain and tendon damage is a significant complication related to hyperlipidemia. Unlike the well-established pathogenesis associated with increased serum concentrations of total cholesterol, triglycerides, and low-density lipoprotein in atherosclerotic cardiovascular disease, the role of hyperlipidemia in promoting tendon damage remains controversial and requires mechanistic clarity.METHODS:In this study, we analyzed the consequences of hypercholesterolemia on the integrity of the collagen-based architecture of the Achilles tendon. The Achilles tendons from rabbits fed with normal-cholesterol (nCH) and high-cholesterol (hCH) diets were analyzed. We studied the morphology of tendons, distribution of lipids within their collagen-rich milieu, the relative amounts of fibrillar collagen I and collagen III, and selected biomechanical parameters of the tendons at the macroscale and the nanoscale.RESULTS:Histological assays of hCH tendons and tenosynovium demonstrated hypercellular areas with increased numbers of macrophages infiltrating the tendon structure as compared to the nCH tendons. While Oil Red staining revealed lipid-rich deposits in the hCH tendons, hybridization of tendon tissue with the collagen hybridizing peptide (CHP) demonstrated damage to the collagen fibers. Fourier-transform infrared (FTIR) spectra showed the presence of distinct peaks consistent with the presence of cholesterol ester. Additionally, the hCH tendons displayed regions of poor collagen content that overlapped with lipid-rich regions. The hCH tendons had a substantial fourfold increase in the collage III to collagen I ratio as compared to the nCH tendons. Tendons from the hCH rabbits showed poor biomechanical characteristics in comparison with control. The biomechanical changes were evident at the macrolevel and the nanolevel of tendon structure.CONCLUSIONS:Our findings support the hypothesis that hypercholesterolemia coincides with the weakening of the tendons. It is likely that the intimate contact between collagen fibrils and cholesterol deposits contributes to the weakening of the fibrillar structure of the tendons.

Age-related tendon degeneration (tendinosis) is characterized by a phenotypic change in which tenocytes display characteristics of fibrochondrocytes and mineralized fibrochondrocytes. As tendon degeneration has been noted in vivo in areas of decreased tendon vascularity, we hypothesized that hypoxia is responsible for the development of the tendinosis phenotype, and that these effects are more pronounced in aged tenocytes. Hypoxic (1% O2 ) culture of aged, tendinotic, and young human tenocytes resulted in a mineralized fibrochondrocyte phenotype in aged tenocytes, and a fibrochondrocyte phenotype in young and tendinotic tenocytes. Investigation of the molecular mechanism responsible for this phenotype change revealed that the fibrochondrocyte phenotype in aged tenocytes occurs with decreased Rac1 activity in response to hypoxia. In young hypoxic tenocytes, however, the fibrochondrocyte phenotype occurs with concomitant decreased Rac1 activity coupled with increased RhoA activity. Using pharmacologic and adenoviral manipulation, we confirmed that these hypoxic effects on the tenocyte phenotype are linked directly to the activity of RhoA/Rac1 GTPase in in vitro human cell culture and tendon explants. These results demonstrate that hypoxia drives tenocyte phenotypic changes, and provide a molecular insight into the development of human tendinosis that occurs with aging.

Skeletal dysplasias form a group of skeletal disorders caused by mutations in macromolecules of cartilage and bone. The severity of skeletal dysplasias ranges from precocious arthropathy to perinatal lethality. Although the pathomechanisms of these disorders are generally well defined, the feasibility of repairing established aberrant skeletal tissues that developed in the presence of mutant molecules is currently unknown. Here, we employed a validated mouse model of spondyloepiphyseal dysplasia (SED) that enables temporal control of the production of the R992C (p.R1192C) collagen II mutant that causes this disease. Although in our earlier studies we determined that blocking the expression of this mutant at the early prenatal stages prevents a SED phenotype, the utility of blocking the R992C collagen II at the postnatal stages is not known. Here, by switching off the expression of R992C collagen II at various postnatal stages of skeletal development, we determined that significant improvements of cartilage and bone morphology were achieved only when blocking the production of the mutant molecules was initiated in newborn mice. Our study indicates that future therapies of skeletal dysplasias may require defining a specific time window when interventions should be applied to be successful.

Experimental approaches to improving tissue repair utilize cells and growth factors needed to restore the architecture and function of damaged tissues and organs. Key limitations of these approaches include poor delivery of therapeutic cells and growth factors into injury sites, as well as their short-term retention in target areas. In our earlier studies, we demonstrated that artificial collagen-specific anchor (ACSA) expressed on the surface of therapeutic cells directs them into collagen-rich sites of injury. Moreover, we demonstrated that the ACSA improves the retention of these cells in target sites, thereby promoting tissue repair. To advance the ACSA-based technology, we engineered the second generation of the ACSA-expressing cells able to deliver growth factors to target sites. In this study, we specifically focused on insulin growth factor 1 (IGF1), which enhances the repair of a number of collagen-rich connective tissues, including ligament and tendon. Utilizing gene engineering, we produced IGF1 in the ACSA-expressing cells. Using relevant experimental models, we demonstrated that recombinant IGF1 secreted by these cells maintains its specificity and biological activity. Moreover, our studies show that IGF1 produced by the ACSA-expressing cells cultured in three-dimensional environment promotes the formation of the collagen-rich fibrillar matrix. Furthermore, the engineered cells integrated well with the native collagen-rich tendon tissue. Our study provides strong evidence for the great potential of cells with rationally engineered target-specific receptors to restore damaged connective tissues. Future studies in relevant animal models will determine the utility of these cells in vivo.

Spondyloepiphyseal dysplasia (SED) exemplifies a group of heritable diseases caused by mutations in collagenous proteins of the skeletal system. Its main feature is altered skeletal growth. Pathomechanisms of SED include: changes in the stability of collagen II molecules, inability to form proper collagen fibrils, excessive intracellular retention of mutant molecules, and endoplasmic reticulum stress. The complexity of this patho-mechanism presents a challenge for designing therapies for SED. Our earlier research tested whether such therapies only succeed when applied during a limited window of development. Here, employing an inducible mouse model of SED caused by the R992C mutation in collagen II, we corroborate our earlier observations that a therapy must be applied at the prenatal or early postnatal stages of skeletal growth in order to be successful. Moreover, we demonstrate that blocking the expression of the R992C collagen II mutant at the early prenatal stages leads to long-term positive effects. Although, we could not precisely mark the start of the expression of the mutant, these effects are not significantly changed by switching on the mutant production at the early postnatal stages. By demonstrating the need for early therapeutic interventions, our study provides, for the first time, empirically-based directions for designing effective therapies for SED and, quite likely, for other skeletal dysplasias caused by mutations in key macromolecules of the skeletal system.

Skeletal dysplasias form a group of skeletal disorders caused by mutations in macromolecules of cartilage and bone. The severity of skeletal dysplasias ranges from precocious arthropathy to perinatal lethality. Although the pathomechanisms of these disorders are generally well defined, the feasibility of repairing established aberrant skeletal tissues that developed in the presence of mutant molecules is currently unknown. Here, we employed a validated mouse model of spondyloepiphyseal dysplasia (SED) that enables temporal control of the production of the R992C (p.R1192C) collagen II mutant that causes this disease. Although in our earlier studies we determined that blocking the expression of this mutant at the early prenatal stages prevents a SED phenotype, the utility of blocking the R992C collagen II at the postnatal stages is not known. Here, by switching off the expression of R992C collagen II at various postnatal stages of skeletal development, we determined that significant improvements of cartilage and bone morphology were achieved only when blocking the production of the mutant molecules was initiated in newborn mice. Our study indicates that future therapies of skeletal dysplasias may require defining a specific time window when interventions should be applied to be successful.

Post-traumatic joint contracture is a frequent orthopaedic complication that limits the movement of injured joints, thereby severely impairing affected patients. Non-surgical and surgical treatments for joint contracture often fail to improve the range of motion. In this study, we tested a hypothesis that limiting the formation of collagen-rich tissue in the capsules of injured joints would reduce the consequences of the fibrotic response and improve joint mobility. We targeted the formation of collagen fibrils, the main component of fibrotic deposits formed within the tissues of injured joints, by employing a relevant rabbit model to test the utility of a custom-engineered antibody. The antibody was delivered directly to the cavities of injured knees in order to block the formation of collagen fibrils produced in response to injury. In comparison to the non-treated control, mechanical tests of the antibody-treated knees demonstrated a significant reduction of flexion contracture. Detailed microscopic and biochemical studies verified that this reduction resulted from the antibody-mediated blocking of the assembly of collagen fibrils. These findings indicate that extracellular processes associated with excessive formation of fibrotic tissue represent a valid target for limiting post-traumatic joint stiffness. (c) 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1038-1046, 2017.

Introduction Although collagen-rich deposits are the main component of neural scars, the patterns of their formation are ill defined. Essential to the biosynthesis of collagen fibrils are enzymes catalyzing posttranslational modifications and chaperones that control the formation of the collagen triple helix. Prolyl-4-hydroxylase (P4H) and heat shock protein-47 (HSP47) play a key role, and their production is upregulated during scar formation in human tissues. Alpha smooth muscle actin (SMA) is also produced during fibrotic processes in myofibroblasts that participate in fibrotic response. In injured peripheral nerves, however, the distribution of cells that produce these markers is poorly understood.Methods The goal of this study was to determine the distribution of the SMA-positive, HSP47-positive, and the P4H-positive cells to better understand the formation of collagen-rich fibrotic tissue (FT) in response to peripheral nerve injury. To reach this goal, we employed a rabbit model of crush-injury and partial-transection injury of the sciatic nerves.Results Our study demonstrated that SMA is expressed in a relatively small number of cells seen in neural FT. In contrast, cells producing P4H and HSP47 are ubiquitously present in sites of injury of the sciatic nerves.Conclusion We contemplate that these proteins may serve as valuable markers that define fibrotic activities in the injured peripheral nerves.

ABSTRACTPost‐traumatic joint contracture is a debilitating consequence of trauma or surgical procedures. It is associated with fibrosis that develops regardless of the nature of initial trauma and results from complex biological processes associated with inflammation and cell activation. These processes accelerate production of structural elements of the extracellular matrix, particularly collagen fibrils. Although the increased production of collagenous proteins has been demonstrated in tissues of contracted joints, researchers have not yet determined the complex protein machinery needed for the biosynthesis of collagen molecules and for their assembly into fibrils. Consequently, the purpose of our study was to investigate key enzymes and protein chaperones needed to produce collagen‐rich deposits. Using a rabbit model of joint contracture, our biochemical and histological assays indicated changes in the expression patterns of heat shock protein 47 and the α‐subunit of prolyl 4‐hydroxylase, key proteins in processing nascent collagen chains. Moreover, our study shows that the abnormal organization of collagen fibrils in the posterior capsules of injured knees, rather than excessive formation of fibril‐stabilizing cross‐links, may be a key reason for observed changes in the mechanical characteristics of injured joints. This result sheds new light on pathomechanisms of joint contraction, and identifies potentially attractive anti‐fibrotic targets. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:489–501, 2016.

Mutations in collagen II, a main structural protein of cartilage, are associated with various forms of spondyloepiphyseal dysplasia (SED), whose main features include aberrations of linear growth. Here, we analyzed the pathomechanisms responsible for growth alterations in transgenic mice with conditional expression of the R992C collagen II mutation. Specifically, we studied the alterations of the growth plates of mutant mice in which chondrocytes lacked their typical columnar arrangement. Our studies demonstrated that chondrocytes expressing the thermolabile R992C mutant collagen II molecules endured endoplasmic reticulum stress, had atypical polarization, and had reduced proliferation. Moreover, we demonstrated aberrant organization and morphology of primary cilia. Analyses of the extracellular collagenous deposits in mice expressing the R992C mutant collagen II molecules indicated their poor formation and distribution. By contrast, transgenic mice expressing wild-type collagen II and mice in which the expression of the transgene encoding the R992C collagen II was switched off were characterized by normal growth, and the morphology of their growth plates was correct. Our study with the use of a conditional mouse SED model not only indicates a direct relation between the observed aberration of skeletal tissues and the presence of mutant collagen II, but also identifies cellular and matrix elements of the pathomechanism of SED.