OBJECTIVE

This review article aims to determine the properties, uses, toxicity, and other side effects of crosslinking agents in tissue scaffolds when applied in vitro and in vivo.

A literature search was performed using the PubMed-NCBI (MEDLINE) database (https://pubmed.ncbi.nlm. nih.gov/) with keywords: crosslinking reagent, collagen, hydroxyapatite, and bone regeneration. GRADE criteria were used to assess the quality of evidence.

A total of six articles were included in the study. Improved mechanical properties of collagen-hydroxyapatite scaffolds with high porosity can be achieved by employing crosslinking methods, including physical dehydrothermal (DHT) treatment, chemical treatment with glutaraldehyde (GA), Microbial Transglutaminase (mTGase), 1‐ethyl‐3‐(3‐ dimethylaminopropyl) carbodiimide (EDAC), or a combination of both DHT and EDAC. Furthermore, the crosslinking of EDAC and DHT can lead to forming ester bonds between activated carboxyl groups and hydroxyl groups.

The combination of DHT and EDAC crosslinking can increase mechanical strength, make the pore size appropriate, make the scaffold more stable, and support cell adhesion so that new cells can grow, and the process of osteogenesis can run more optimally.

Riboflavin-ultraviolet A (UVA) collagen cross-linking has not only achieved good clinical efficacy in the treatment of corneal diseases such as dilatation keratopathy, bullae keratopathy, infectious keratopathy, and in the combined treatment of corneal refractive surgeries, but also its efficacy and safety in scleral collagen cross-linking have been initially confirmed. To better promote the application of cross-linking in the clinical treatment of corneal and scleral diseases, exploring controllability and predictability of the surgical efficacy are both important for evaluating the surgical efficacy and personalized precision treatment. In this paper, the progress on the cross-linking depth of riboflavin-UVA collagen cross-linking, and its relationship with the cross-linking effect will be reviewed. It will provide the reference for further application of this procedure in ophthalmology clinics.
Riboflavin/pharmacology* ; Humans ; Collagen/radiation effects* ; Ultraviolet Rays ; Cross-Linking Reagents ; Corneal Diseases/drug therapy* ; Photosensitizing Agents/therapeutic use*

The three-dimensional (3D) printed bone tissue repair guide scaffold is considered a promising method for treating bone defect repair. In this experiment, chitosan (CS), sodium alginate (SA), and mineralized collagen (MC) were combined and 3D printed to form scaffolds. The experimental results showed that the printability of the scaffold was improved with the increase of chitosan concentration. Infrared spectroscopy analysis confirmed that the scaffold formed a cross-linked network through electrostatic interaction between chitosan and sodium alginate under acidic conditions, and X-ray diffraction results showed the presence of characteristic peaks of hydroxyapatite, indicating the incorporation of mineralized collagen into the scaffold system. In the <i>in vitroi> collagen release experiments, a weakly alkaline environment was found to accelerate the release rate of collagen, and the release amount increased significantly with a lower concentration of chitosan. Cell experiments showed that scaffolds loaded with mineralized collagen could significantly promote cell proliferation activity and alkaline phosphatase expression. The subcutaneous implantation experiment further verified the biocompatibility of the material, and the implantation of printed scaffolds did not cause significant inflammatory reactions. Histological analysis showed no abnormal pathological changes in the surrounding tissues. Therefore, incorporating mineralized collagen into sodium alginate/chitosan scaffolds is believed to be a new tissue engineering and regeneration strategy for achieving enhanced osteogenic differentiation through the slow release of collagen.
Chitosan/chemistry* ; Alginates/chemistry* ; Tissue Scaffolds/chemistry* ; Printing, Three-Dimensional ; Osteogenesis ; Collagen/chemistry* ; Cell Differentiation ; Animals ; Tissue Engineering/methods* ; Cell Proliferation ; Biocompatible Materials ; Glucuronic Acid/chemistry* ; Hexuronic Acids/chemistry*

Abnormal accumulation of collagen fibrils is a hallmark feature of oral submucous fibrosis (OSF). However, the precise characteristics and underlying mechanisms remain unclear, impeding the advancement of potential therapeutic approaches. Here, we observed that collagen I, the main component of the extracellular matrix, first accumulated in the lamina propria and subsequently in the submucosa of OSF specimens as the disease progressed. Using RNA-seq and Immunofluorescence in OSF specimens, we screened the cartilage oligomeric matrix protein (COMP) responsible for the abnormal collagen accumulation. Genetic COMP deficiency reduced arecoline-stimulated collagen I deposition significantly in vivo. In comparison, both COMP and collagen I were upregulated under arecoline stimulation in wild-type mice. Human oral buccal mucosal fibroblasts (hBMFs) also exhibited increased secretion of COMP and collagen I after stimulation in vitro. COMP knockdown in hBMFs downregulates arecoline-stimulated collagen I secretion. We further demonstrated that hBMFs present heterogeneous responses to arecoline stimulation, of which COMP-positive fibroblasts secrete more collagen I. Since COMP is a molecular bridge with Fibril-associated collagens with Interrupted Triple helices (FACIT) in the collagen network, we further screened and identified collagen XIV, a FACIT member, co-localizing with both COMP and collagen I. Collagen XIV expression increased under arecoline stimulation in wild-type mice, whereas it was hardly expressed in the Comp^-/- mice, even with under stimulation. In summary, we found that COMP may mediates abnormal collagen I deposition by functions with collagen XIV during the progression of OSF, suggesting its potential to be targeted in treating OSF.
Oral Submucous Fibrosis/pathology* ; Cartilage Oligomeric Matrix Protein/genetics* ; Animals ; Mice ; Humans ; Fibroblasts/metabolism* ; Collagen Type I/metabolism* ; Arecoline/pharmacology* ; Mouth Mucosa/metabolism* ; Cells, Cultured ; Fluorescent Antibody Technique

Temporomandibular joint (TMJ) disc displacement is one of the most significant subtypes of temporomandibular joint disorders, but its etiology and mechanism are poorly understood. In this study, we elucidated the mechanisms by which destruction of inflamed collagen fibrils induces alterations in the mechanical properties and positioning of the TMJ disc. By constructing a rat model of TMJ arthritis, we observed anteriorly dislocated TMJ discs with aggravated deformity in vivo from five weeks to six months after a local injection of Freund's complete adjuvant. By mimicking inflammatory conditions with interleukin-1 beta in vitro, we observed enhanced expression of collagen-synthesis markers in primary TMJ disc cells cultured in a conventional two-dimensional environment. In contrast, three-dimensional (3D)-cultivated disc cell sheets demonstrated the disordered assembly of inflamed collagen fibrils, inappropriate arrangement, and decreased Young's modulus. Mechanistically, inflammation-related activation of the nuclear factor kappa-B (NF-κB) pathway occurs during the progression of TMJ arthritis. NF-κB inhibition reduced the collagen fibril destruction in the inflamed disc cell sheets in vitro, and early NF-κB blockade alleviated collagen degeneration and dislocation of the TMJ discs in vivo. Therefore, the NF-κB pathway participates in the collagen remodeling in inflamed TMJ discs, offering a potential therapeutic target for disc displacement.
Animals ; NF-kappa B/metabolism* ; Temporomandibular Joint Disorders/pathology* ; Temporomandibular Joint Disc/metabolism* ; Rats ; Rats, Sprague-Dawley ; Disease Models, Animal ; Male ; Collagen/metabolism* ; Cells, Cultured ; Joint Dislocations/pathology* ; Interleukin-1beta ; Arthritis, Experimental

Xerostomia (dry mouth) is frequently experienced by patients treated with radiotherapy for head and neck cancers or with Sjögren's syndrome, with no permanent cure existing for this debilitating condition. To this end, in vitro platforms are needed to test therapies directed at salivary (fluid-secreting) cells. However, since these are highly differentiated secretory cells, the maintenance of their differentiated state while expanding in numbers is challenging. In this study, the efficiency of three reversible thermo-ionically crosslinked gels: (1) alginate-gelatin (AG), (2) collagen-containing AG (AGC), and (3) hyaluronic acid-containing AG (AGHA), to recapitulate a native-like environment for human salivary gland (SG) cell expansion and 3D spheroid formation was compared. Although all gels were of mechanical properties comparable to human SG tissue (~11 kPa) and promoted the formation of 3D spheroids, AGHA gels produced larger (>100 cells/spheroid), viable (>93%), proliferative, and well-organized 3D SG spheroids while spatially and temporally maintaining the high expression of key SG proteins (aquaporin-5, NKCC1, ZO-1, α-amylase) for 14 days in culture. Moreover, the spheroids responded to agonist-induced stimulation by increasing α-amylase secretory granules. Here, we propose alternative low-cost, reproducible, and reversible AG-based 3D hydrogels that allow the facile and rapid retrieval of intact, highly viable 3D-SG spheroids.
Humans ; Hydrogels/chemistry* ; Acinar Cells/cytology* ; Spheroids, Cellular/cytology* ; Salivary Glands/cytology* ; Gelatin/chemistry* ; Collagen/chemistry* ; Alginates/chemistry* ; Cell Culture Techniques/methods* ; Hyaluronic Acid/chemistry* ; Cell Proliferation ; Cell Survival ; Cells, Cultured

Five previously undescribed diterpenoids, named succipenoids D‒H (1‒5), along with four undescribed lignans, named succignans A‒D (6‒9), were isolated from the dichloromethane extract of Chinese medicinal succinum. Compounds 1‒5 were characterized as nor-abietane diterpenoids, while compounds 6‒9 were identified as lignans polymerized from two groups of phenylpropanoid units. The structures of these novel compounds, including their absolute configurations, were determined through spectroscopic and computational methods. Biological assessments of renal fibrosis demonstrated that compounds 6 and 7 effectively reduce the expression of proteins associated with renal fibrosis, including α-smooth muscle actin (α-SMA), collagen I, and fibronectin in transforming growth factor-β1 (TGF-β1) induced normal rat kidney proximal tubular epithelial cells (NRK-52e).
Animals ; Rats ; Lignans/isolation & purification* ; Diterpenes/isolation & purification* ; Fibrosis/drug therapy* ; Drugs, Chinese Herbal/pharmacology* ; Molecular Structure ; Cell Line ; Kidney Diseases/pathology* ; Transforming Growth Factor beta1/genetics* ; Kidney/metabolism* ; Actins/genetics* ; Fibronectins/genetics* ; Collagen Type I/genetics* ; Epithelial Cells/metabolism*

OBJECTIVES: To compare the clinical effects of concentrated growth factor (CGF) membrane and Bio-Gide ^® collagen membrane, combined with Bio-Oss ^® sticky bone respectively in alveolar ridge preservation (ARP) of maxillary anterior teeth. METHODS: Thirty patients who needed alveolar ridge preservation after maxillary anterior tooth extraction were selected and randomly assigned to the Bio-Gide group and the CGF group. In both groups, the extraction sockets were tightly filled with the Bio-Oss^® sticky bone. In the Bio-Gide group used Bio-Gide^® collagen membrane to cover the upper edge of the Bio-Oss^® sticky bone and closed the wound. The CGF group, the CGF membrane was covered on the upper edge of the Bio-Oss^® sticky bone and the wound was closed. The soft tissue wound healing status at 10 days after ARP, the changes in alveolar ridge height and width immediately after ARP and at 6 months after ARP, and the doctor-patient satisfaction at 6 months after ARP were compared and evaluated between the two groups. RESULTS: At 6 months after ARP, there was no statistically significant difference in the changes of alveolar bone width and height between the two groups (<i>Pi>>0.05). However, the CGF group showed better performance in soft tissue healing after ARP and doctor-patient satisfaction, and the differences were statistically significant (<i>Pi><0.05). CONCLUSIONS Compared with the Bio-Gide^® collagen membrane, the combined application of CGF membrane and Bio-Oss^® sticky bone can lead to better soft tissue healing after ARP of maxillary anterior teeth and higher doctor-patient satisfaction, showing obvious advantages in ARP of maxillary anterior teeth.
Humans ; Maxilla/surgery* ; Tooth Extraction ; Alveolar Process/surgery* ; Membranes, Artificial ; Alveolar Ridge Augmentation/methods* ; Intercellular Signaling Peptides and Proteins/therapeutic use* ; Minerals/therapeutic use* ; Collagen ; Wound Healing ; Tooth Socket/surgery* ; Bone Substitutes/therapeutic use* ; Male ; Female ; Middle Aged ; Alveolar Bone Loss/prevention & control* ; Adult

Collagen, a vital matrix protein for various tissue and functions in animals, is widely applied in biomaterials. In type Ⅰ collagen, missense mutations of glycine (Gly) in the Gly-Xaa-Yaa triplet of the triple helix are a major cause of osteogenesis imperfecta (OI). Clinical manifestations exhibit marked heterogeneity, spanning a broad disease spectrum from mild skeletal fragility (Type Ⅰ) to severe limb deformities (Type Ⅲ) and perinatal lethal forms (Type Ⅱ). This study utilized recombinant collagen as a model to further elucidate whether Gly→Ala/Val mutations at the N-terminus of the integrin-binding sequence GFPGER affect collagen structure and function, and to explore the underlying mechanisms by which missense mutations impact the biological function of collagen. By introducing Ala and Val substitutions at seven Gly positions N-terminal to the GFPGER sequence, we systematically assessed the effects of these amino acid replacements on the triple-helical structure, thermal stability, integrin-binding ability, and cell adhesion of recombinant collagen. All constructs formed a stable triple-helix structure, with slightly compromised thermal stability. Gly→Val substitutions increased the susceptibility of recombinant collagen to trypsin, which suggested local conformational perturbations in the triple helix. In addition, Gly→Val substitutions significantly reduced the integrin-binding affinity and decreased HT1080 cell adhesion, with the effects stronger than Gly→Ala substitutions. Compared with Gly→Ala substitutions, substitution of Gly with the larger residue Val had enhanced negative effects on the structure and function of recombinant collagen. These findings provide new insights into the molecular mechanisms of osteogenesis imperfecta and offer theoretical references and experimental foundations for the design of collagen sequences and the development of collagen-based biomaterials.
Recombinant Proteins/biosynthesis* ; Glycine/genetics* ; Humans ; Osteogenesis Imperfecta/genetics* ; Integrins/metabolism* ; Collagen/metabolism* ; Collagen Type I/metabolism* ; Amino Acid Substitution ; Mutation ; Mutation, Missense

OBJECTIVE: To review the research progress of pathological changes of glenohumeral capsule in patients with recurrent shoulder anterior dislocation (RSAD). METHODS: The literature on shoulder capsules, both domestic and international, was reviewed. The anatomy, histology, and molecular biology characteristics of the glenohumeral capsule in RSAD patients were summarized. RESULTS: Anatomically, the glenohumeral capsule is composed of four distinct parts: the upper, lower, anterior, and posterior sections. The thickness of these sections is uneven, and the stability of the capsule is further enhanced by the presence of the glenohumeral and coracohumeral ligaments. Histologically, the capsule tissue undergoes adaptive changes following RSAD, which improve its ability to withstand stretching and deformation. In the realm of molecular biology, genes associated with the regulation of structure formation, function, and extracellular matrix homeostasis of the shoulder capsule's collagen fibers exhibit varying degrees of expression changes. Specifically, the up-regulation of transforming growth factor β ₁ (TGF-β ₁), TGF-β receptor 1, lysyl oxidase, and procollagen-lysine, 2-oxoglutarate 5-dioxygenase 1 facilitates the repair of the joint capsule, thereby contributing to the maintenance of shoulder joint stability. Conversely, the up-regulation of collagen type Ⅰ alpha 1 (COL1A1), COL3A1, and COL5A1 is linked to the recurrence of shoulder anterior dislocation, as these changes reflect the joint capsule's response to dislocation. Additionally, the expressions of tenascin C and fibronectin 1 may play a role in the pathological processes occurring during the early stages of RSAD. CONCLUSION Glenohumeral capsular laxity is both a consequence of RSAD and a significant factor contributing to its recurrence. While numerous studies have documented alterations in the shoulder capsule following RSAD, further research is necessary to confirm the specific pathological anatomy, histological, and molecular biological changes involved.
Humans ; Joint Capsule/metabolism* ; Shoulder Dislocation/metabolism* ; Recurrence ; Shoulder Joint/metabolism* ; Tenascin/metabolism* ; Transforming Growth Factor beta1/genetics* ; Collagen Type I/genetics* ; Extracellular Matrix/metabolism*