Effect Of Collagen Cross-linking Research Articles

Investigating the influence of collagen cross-linking on mechanical properties of thoracic aortic tissue.

Vascular diseases, such as abdominal aortic aneurysms, are associated with tissue degeneration of the aortic wall, resulting in variations in mechanical properties, such as tissue ultimate stress and a high slope. Variations in the mechanical properties of tissues may be associated with an increase in the number of collagen cross-links. Understanding the effect of collagen cross-linking on tissue mechanical properties can significantly aid in predicting diseased aortic tissue rupture and improve the clarity of decisions regarding surgical procedures. Therefore, this study focused on increasing the density of the aortic tissue through cross-linking and investigating the mechanical properties of the thoracic aortic tissue in relation to density. Uniaxial tensile tests were conducted on the porcine thoracic aorta in four test regions (anterior, posterior, distal, and proximal), two loading directions (circumferential and longitudinal), and density increase rates (0%-12%). As a result, the PPC (Posterior/Proximal/Circumferential) group experienced a higher ultimate stress than the PDC (Posterior/Distal/Circumferential) group. However, this relationship reversed when the specimen density exceeded 3%. In addition, the ultimate stress of the ADC (Anterior/Distal/Circumferential) and PPC group was greater than that of the APC (Anterior/Proximal/Circumferential) group, while these findings were reversed when the specimen density exceeded 6% and 9%, respectively. Finally, the high slope of the PDL (Posterior/Distal/Longitudinal) group was lower than that of the ADL (Anterior/Distal/Longitudinal) group, but the high slope of the PDL group appeared larger due to the stabilization treatment. This highlights the potential impact of density variations on the mechanical properties of specific specimen groups.

Transglutaminase-induced Cornea Collagen Crosslinking Effect on Central Corneal Thickness and Keratocyte Cell Density

Abstract
 Introduction & Objectives : This research is aimed to evaluate the effect of transglutaminase-induced corneal collagen crosslinking (CXL) on central corneal thickness (CCT) and keratocyte cell density in vivo.
 Methods : Twenty-eight white New Zealand rabbits were divided into four groups: the transglutaminase- induced CXL group, the epithelial-off CXL group, the transepithelial CXL group, and the control group. The ocular surface was treated with a 1 U/mL microbial transglutaminase solution, and both the epithelial-off and transepithelial groups were exposed to clinical ultraviolet A-riboflavin (UVA/RF). The efficacy of each group was evaluated on the 14th day after the procedures. Central corneal thickness was evaluated with Corneal Visualization Scheimpflug Technology (Corvis ST) and keratocyte cell density wad evaluated with histopathology examination.
 Results : Transglutaminase-induced CXL group exhibited the highest mean biomechanical CCT (370.14 ± 38.85) in comparison to the UVA/RF epithelial-off group (368.00 ± 25.48), the UVA/RF transepithelial group (369.86 ± 23.43), and the control group (365.14 ± 28.74). Still, there was no significant differences in both biomechanical CCT (p=0.990). Transglutaminase-induced CXL group had the highest mean of keratocyte cell density (43.26 ± 10.65) compared to UVA/RF epithelial-off (29.99 ± 4.79), UVA/RF transepithelial group (42.03 ± 6.55), and control group (34.36 ± 6.76). There was a significant difference between the group, with a p-value of 0.008.
 Conclusion : Transglutaminase shown that it produces favorable results for CCT and keratocyte density after CXL, which are two factors contributing to corneal rigidity. The outcomes are equivalent to riboflavin used as a conventional element in CXL.

Effect of Corneal Collagen Cross Linking For Keratoconus on The Ocular Higher Order Aberrations Corneal Collagen Cross Linking effect

Effect of Corneal Collagen Cross Linking For Keratoconus on The Ocular Higher Order Aberrations Corneal Collagen Cross Linking effect

Questionable effect of collagen cross-linking for keratoconus in a pregnant female: A case report

Questionable effect of collagen cross-linking for keratoconus in a pregnant female: A case report

Effectiveness of collagen cross-linking induced by two-photon absorption properties of a femtosecond laser in ex vivo human corneal stroma.

This study aimed to investigate the effectiveness of two-photon induced collagen cross-linking (CXL) using femtosecond lasers in human corneal stroma. An 800-nm femtosecond laser optical path for CXL was established. Corneal samples that received two-photon induced CXL and ultraviolet-A (UVA) CXL underwent uniaxial stretching experiments, proteolytic resistance assays and observation of collagen fiber structure changes. Two-photon induced CXL can achieve corneal stiffening effects comparable to UVA CXL and showed better advantages at low strains. The cornea after two-photon induced CXL exhibited high enzymatic resistance and tight collagen fiber arrangement. Two-photon induced CXL promises to be a new option for keratoconus.

The histopathological findings in excised upper eyelids of patients with dermatochalasis following collagen cross-linking treatment.

To evaluate the histopathological effects of collagen cross-linking (CCL) on excised skin samples of patients undergoing upper eyelid blepharoplasty due to dermatochalasis. This study examined 74 excised eyelid skin samples from 37 dermatochalasis patients. Following an upper eyelid blepharoplasty, CCL with hypotonic riboflavin (0.1%) was applied. Both treated (right eyelid, CCL group) and untreated eyelid specimen (left eyelid, non-CCL group) sections were stained with hematoxylin-eosin and Masson's trichrome. The sections were evaluated for the following parameters: the collagen status (parallel, oblique, and perpendicular), the distance between collagen fibers, the diameter of collagen fibers, and the length of collagen fibers. There were no statistically significant differences in the collagen status, the distance between collagen fibers, the diameter of collagen fibers, and the length of collagen fibers between the CCL and non-CCL groups (p > 0.05 for all). Although the lack of statistically significant differences, the structure of the treated eyelid collagen fibers was more parallel in 48% of the participants than in the untreated ones. For male patients, a statistically significant shorter distance between collagen fibers was observed in the CCL group (8.05 ± 2.04µm) compared to the non-CCL group (9.97 ± 2.33µm) (p = 0.042). In this study, more parallel collagen structures and tightly packed collagen fibers were detected in eyelid samples following CCL treatment. The authors note that the results of this study may be promising for further research, so the effect of CCL therapy on the eyelid may be an interesting subject for the treatment of non-severe or surgically inadequately corrected dermatochalasis.

Visual and refractive effects of collagen cross-linking in keratoconus

Visual and refractive effects of collagen cross-linking in keratoconus

Modulatory Effect of Glycated Collagen on Oral Streptococcal Nanoadhesion

Biofilm-mediated oral diseases such as dental caries and periodontal disease remain highly prevalent in populations worldwide. Biofilm formation initiates with the attachment of primary colonizers onto surfaces, and in the context of caries, the adhesion of oral streptococci to dentinal collagen is crucial for biofilm progression. It is known that dentinal collagen suffers from glucose-associated crosslinking as a function of aging or disease; however, the effect of collagen crosslinking on the early adhesion and subsequent biofilm formation of relevant oral streptococci remains unknown. Therefore, the aim of this work was to determine the impact of collagen glycation on the initial adhesion of primary colonizers such as Streptococcus mutans UA159 and Streptococcus sanguinis SK 36, as well as its effect on the early stages of streptococcal biofilm formation in vitro. Type I collagen matrices were crosslinked with either glucose or methylglyoxal. Atomic force microscopy nanocharacterization revealed morphologic and mechanical changes within the collagen matrix as a function of crosslinking, such as a significantly increased elastic modulus in crosslinked fibrils. Increased nanoadhesion forces were observed for S. mutans on crosslinked collagen surfaces as compared with the control, and retraction curves obtained for both streptococcal strains demonstrated nanoscale unbinding behavior consistent with bacterial adhesin-substrate coupling. Overall, glucose-crosslinked substrates specifically promoted the initial adhesion, biofilm formation, and insoluble extracellular polysaccharide production of S. mutans, while methylglyoxal treatment reduced biofilm formation for both strains. Changes in the adhesion behavior and biofilm formation of oral streptococci as a function of collagen glycation could help explain the biofilm dysbiosis seen in older people and patients with diabetes. Further studies are necessary to determine the influence of collagen crosslinking on the balance between acidogenic and nonacidogenic streptococci to aid in the development of novel preventive and therapeutic treatment against dental caries in these patients.

A study on corneal collagen cross-linking effect on quality of life in bullous keratopathy patients

A study on corneal collagen cross-linking effect on quality of life in bullous keratopathy patients

Biomechanical and Morphologic Effects of Collagen Cross-Linking in Human Tarsus.

PurposeTo investigate the feasibility of increasing the stiffness of human tarsal tissue following treatment with riboflavin and ultraviolet A (UVA) to induce cross-linking of collagen fibers.MethodsIn this case control study, 18 right and left upper eyelids were excised en bloc from 18 fresh-frozen cadavers. One side served as the control while the samples from the opposite side were cross-linked. Four 2 × 6-mm vertical strips of central tarsus were cut from the superior to inferior border of each tarsal plate. Sample tissue was irradiated with UVA at 6 mW/cm2 for 18 minutes. A microtensile load cell and an optical coherence tomography scanner allowed calculation of stiffness (Young's modulus). Six cross-linked samples and corresponding controls were stained with hematoxylin and eosin (H&E) and Masson trichrome stains. Four controls and four cross-linked samples were also reviewed with a transmission electron microscope.ResultsMean Young's modulus in the linear region for controls was 28 ± 9 MPa and was much higher at 138 ± 8 MPa for cross-linked samples (P < 0.001), yielding a 493% mean stiffness increase. Staining with H&E and Masson did not reveal any histologic changes. Transmission electron microscopy showed a decrease in average diameter of 50 randomly selected collagen fibers from 47.2 ± 1.9 nm prior to cross-linking to 34.2 ± 1.1 nm post cross-linking (P < 0.001). Qualitatively, the collagen fibers appeared more closely packed following cross-linking.ConclusionsThe findings of this study suggest that collagen cross-linking is a viable and effective modality for increasing the stiffness of human tarsal plates.Translational RelevanceThis work provides proof that collagen cross-linking produces stiffening of the human tarsal plate and may be used in disorders that cause eyelid laxity.

Effects of collagen crosslinking on porcine and human tarsal plate

BackgroundFloppy eyelid syndrome is a disorder in which the tarsal plate is easily distensible and is currently treated with conservative or surgical measures. Human tarsal plate contains type I collagen, which is crosslinked in corneal tissue as a treatment for keratoconus. We hypothesized that collagen crosslinking would similarly stiffen tarsal plate tissue and investigated this in porcine and human tarsal plate specimens.MethodsRiboflavin-sensitized porcine and human tarsus samples were irradiated with ultraviolet-A light. Porcine experiments were analyzed with gross photographs, anterior segment optical computed tomography (AS-OCT) imaging, and tensile testing. A prospective study of human tarsus was performed on samples from patients undergoing wedge resection for floppy eyelid syndrome and was analyzed with AS-OCT and tensile testing.Results73 porcine adnexa and 9 patients (16 eyelids) who underwent wedge excision were included in the study. Grossly, greater stiffness was observed in crosslinked porcine tissue. AS-OCT imaging in porcine tissue showed a distinct hyperreflective band in crosslinked specimens whose area and intensity increased with longer treatment time (P = 0.003); this band was also visible in crosslinked human specimens. Tensile testing was performed, but results were not statistically significant.ConclusionsAS-OCT imaging, which has not been previously described for tarsal plate, showed a characteristic change in crosslinked porcine and human specimens. Tissue stiffness was increased grossly, but changes in tensile properties were not statistically significant. Further study is warranted to determine relevance as a potential treatment for floppy eyelid syndrome.

Effects of scleral collagen crosslinking with different carbohydrate on chemical bond and ultrastructure of rabbit sclera: Future treatment for myopia progression.

BackgroundMyopia is the most common ocular disorder and is mainly caused by axial elongation of the sclera. If the stiffness of sclera increased, it can inhibit myopia progression. The aim of this study is to compare the effect of the collagen crosslinking with different types and concentrations of carbohydrates on chemical bond and ultrastructural change of rabbit sclera.MethodsNine New Zealand white rabbits were treated with five, sequential sub-Tenon injections of 0.15 mL solutions of ribose, sucrose, and glycogen of 0.1, 0.2 and 0.4 M concentration at the right eye over 14 days. Ten weeks after the last injection, the rabbits were sacrificed and chemical bond and ultrastructural changes were compared with those of the untreated left sclera using Raman spectroscopy, atomic force microscopy (AFM), and histology.ResultsRaman spectroscopy of the control and cross-linked rabbit sclera tissue revealed different types of collagen interactions. Raman shift of 919 cm-1 (C-C stretching and vibration of the proline ring in collagen) was the highest in ribose, followed by sucrose and glycogen. Total energy intensity was also highest in ribose, followed by sucrose and glycogen, and showed a tendency to increase at higher concentrations. AFM revealed interlocking arrangements of collagen fibrils. The collagen fibril diameter was 105.6 ± 21.2 nm, 109.4 ± 28.8 nm, 113.1 ± 30.8 nm and 137.6 ± 25.3 nm for control group, 0.4 M glycogen, sucrose, and ribose, respectively. Histology indicated increased density of the collagen bundle and no increase in inflammatory cell recruitment compared to control at high concentrations of ribose.ConclusionsScleral crosslinking using glycation increased the scleral biomechanical rigidity and these results were particularly pronounced in ribose. Scleral crosslinking using glycation may be a promising method for inhibiting high myopia progression.

Dentine collagen cross-linking using tiopronin-protected Au/EDC nanoparticles formulations

ObjectiveThe aim of this study was to investigate EDC-assisted collagen crosslinking effect with different concentrations of tiopronin-protected gold (TPAu) nanoparticles on demineralized dentine. MethodsTPAu nanoparticles were fabricated from 0.31-g tetrachloroauric acid and 0.38-g of N-(2-mercaptopropionyl) glycine (2.4-mmol). Then co-dissolved using 35-mL of 6:1 methanol/acetic acid and mixed using NaBH4. EDC (0.3-M) was conjugated to TPAu nanoparticles at TPAU/EDC-0.25:1, and TPAU/EDC-0.5:1 treatment formulations ratios. Dentin specimens treated with 0.3-M EDC solution alone or left untreated were used as control. Nanoparticles formulations were characterized in term of particles morphology and size, Zeta potential, thermogravimetric analysis and small-angle X-ray scattering. Dentin substrates were characterized in term of TEM investigation, dentin proteases characterization, hydroxyproline liberation, elastic modulus measurement, Raman analysis and confocal microscopy viewing. ResultsTEM evaluation of tiopronin protected gold nanoparticles dispersion revealed nano-clusters formations in both groups. However, based on our TEM measurements, the particle-size was ranging from ˜20 to 50 nm with spherical core-shape which were almost similar for both TPAu/EDC ratios (0.5:1 and 0.25:1). Zeta potential measurements indicate negative nanoparticles surface charge. SAXS profiles for both formulations, suggest a typical profile for uni-lamellar nanoparticles. Superior dentin collagen cross-linking effect was found with the TPAu/EDC nanoparticles formulations compared to the control and EDC treated groups. SignificanceCross-linking of dentin collagen using TPAu coupled with EDC through TPAu/EDC nanoparticles formulations is of potential significance in improving the biodegradation resistance, proteases inhibition, mechanical and structural stability of demineralized dentin substrates. In addition, the cross-linking effect is dependent on TPAu/EDC ratio, whereas higher cross-linking effect was found at TPAu/EDC ratio of 0.5:1.

Effect of Collagen Cross-Linking on Alkali Burn-Induced Corneal Neovascularization in Rabbits.

Objective This study aims at investigating the effects and molecular mechanism of riboflavin-ultraviolet-A-induced cross-linking (corneal collagen cross-linking, CXL) on corneal neovascularization (CNV) in a rabbit alkali burn model. Methods A total of 60 rabbits were injured with alkali burns to induce CNV in the right eye and were randomly divided into six groups: Group A—injury and no treatment; Groups B, C, and D—CXL treatment for 30 min, 15 min, and 45 min administered immediately after injury, respectively; and Groups E and F—CXL treatment for 30 min administered 1 day and 3 days after injury, respectively. CNV area, corneal edema, and corneal epithelial defects were observed on days 4, 7, 10, and 14 after injury. Western blot was used to detect expression of the vascular endothelial growth factor (VEGF), matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-2 (MMP-9), and tissue inhibitor of metalloproteinases 1 (TIMP-1) at 7 and 14 days after injury. Results CXL treatment decreased CNV and corneal edema in all groups compared to Group A. On day 7, MMP-9 expression was significantly increased in all CXL treatment groups, and TIMP-1 was upregulated in Groups D and F compared to Group A. In addition, VEGF, MMP-2, MMP-9, and TIMP-1 expression were increased in Group A on day 14 after injury. Conclusions Our results indicate that riboflavin-ultraviolet-A-induced cross-linking (corneal collagen cross-linking, CXL) significantly inhibits alkali burn-induced CNV in rabbits, possibly through downregulating VEGF, MMP-2, MMP-9, and TIMP-1 expression.

Extracellular matrix with defective collagen cross-linking affects the differentiation of bone cells.

Fibrillar type I collagen, the predominant organic component in bone, is stabilized by lysyl oxidase (LOX)-initiated covalent intermolecular cross-linking, an important determinant of bone quality. However, the impact of collagen cross-linking on the activity of bone cells and subsequent tissue remodeling is not well understood. In this study, we investigated the effect of collagen cross-linking on bone cellular activities employing a loss-of-function approach, using a potent LOX inhibitor, β-aminopropionitrile (BAPN). Osteoblastic cells (MC3T3-E1) were cultured for 2 weeks in the presence of 0–2 mM BAPN to obtain low cross-linked collagen matrices. The addition of BAPN to the cultures diminished collagen cross-links in a dose-dependent manner and, at 1 mM level, none of the major cross-links were detected without affecting collagen production. After the removal of cellular components from these cultures, MC3T3-E1, osteoclasts (RAW264.7), or mouse primary bone marrow-derived stromal cells (BMSCs) were seeded. MC3T3-E1 cells grown on low cross-link matrices showed increased alkaline phosphatase (ALP) activity. The number of multinucleate tartrate-resistant acid phosphatase (TRAP)-positive cells increased in RAW264.7 cells. Initial adhesion, proliferation, and ALP activity of BMSCs also increased. In the animal experiments, 4-week-old C57BL/6 mice were fed with BAPN-containing diet for 8 weeks. At this point, biochemical analysis of bone demonstrated that collagen cross-links decreased without affecting collagen content. Then, the diet was changed to a control diet to minimize the direct effect of BAPN. At 2 and 4 weeks after the change, histological samples were prepared. Histological examination of femur samples at 4 weeks showed a significant increase in the number of bone surface osteoblasts, while the bone volume and surface osteoclast numbers were not significantly affected. These results clearly demonstrated that the extent of collagen cross-linking of bone matrix affected the differentiation of bone cells, underscoring the importance of collagen cross-linking in the regulation of cell behaviors and tissue remodeling in bone. Characterization of collagen cross-linking in bone may be beneficial to obtain insight into not only bone mechanical property, but also bone cellular activities.

PACK-CXL: corneal collagen cross-linking for infectious keratitis

Infectious keratitis is one of the leading causes of irreversible vision loss.With the emergence of multidrug-resistant pathogen, despite appropriate diagnosis and antimicrobial treatment, infectious keratitis can still result in corneal opacity and even melting and perforation.Therefore, it is imperative to develop novel treatment for infectious keratitis.More recently, several articles have demonstrated the effectiveness of collagen cross-linking (CXL) in treating infectious keratitis via direct antimicrobial activity and inhibition of corneal enzymatic degradation.In order to highlight the use of CXL for infectious keratitis treatment, a new term was proposed at the 9th CXL congress to rename its use in infections as photoactivated chromophore for infectious keratitis-CXL (PACK-CXL). Based on the current literature, PACK-CXL appears to be effective for bacterial keratitis, especially on inhibiting corneal perforation.The effect was limited for fungal and amoebic keratitis, and maybe aggravative on viral keratitis.In this article, we will review the basic profile, technical operation, prevention of complication of CXL, the principle mechanism of PACK-CXL and its clinical application for bacterial, fungal keratitis, amoebic and viral keratitis. Key words: Keratitis/therapy; Cross-linking reagents/therapeutic use; Review

The effects of collagen cross-linking on corneal density: a comparison between accelerated and conventional methods.

To compare the corneal density before and after accelerated versus conventional corneal cross-linking (CXL). Pentacam densitometry and corneal mapping were performed before and 6months following the randomly assigned intervention. Corneal density was measured over different zones and layers of the cornea by two independent ophthalmologists. Total corneal density was calculated. The patients were followed up for the occurrence of procedure-related and unrelated complication 1week, 1month and a final visit around 6months (180 ± 10days) after the procedure, during which the corneal density was remeasured and recorded. Twenty-three patients were enrolled. There was a significant increase in corneal density over the anterior layer (P < 0.001) and middle corneal layer (P = 0.035). Additionally, the corneal density significantly increased in 0-2 and 2-6mm zones (P < 0.001), as well as total density (P = 0.002) following corneal CXL. Although there was no significant difference in the corneal density between the two CXL groups, the increase in corneal density was only significant in group A for the 6-10mm zone and the posterior corneal layer. Both the conventional and accelerated CXL methods increase the overall corneal density with exception of posterior central corneal layer and the 6-10mm zone, which is only increased in the accelerated method.

Effects of collagen cross-linking on the keratoconus metabolic network.

Keratoconus (KC) is a multifactorial, ectatic corneal disease. Metabolic changes in the corneal stroma with alterations in collagen fibril stability, oxidative stress, and urea cycle, have previously been reported as key players in KC pathobiology. Recently, corneal collagen cross-linking (CXL) has been introduced as a treatment that can address the progressive nature of KC. While the treatment has been successful in the early days, it is not without clinical ramifications. In this study, we investigated the alterations in KC metabolic profiles due to CXL. Primary human corneal fibroblasts (HCFs) from healthy donors and human KC fibroblasts (HKCs) from KC donor patients were plated on transwell polycarbonate membranes and stimulated by a stable vitamin C. At 4 weeks, riboflavin was added to the cultures followed by UVA irradiation (365 nm). Using mass spectrometry, we measured the major differences in metabolites in HKCs compared to HCFs pre- and post CXL. The analysis of 276 metabolites in HCFs and HKCs revealed that the most affected metabolites due to CXL were glutathione disulfide, ascorbic acid, proline, and lysine. A significant decrease in the pro-inflammatory biomarkers (myo-inositol and histidine) was also observed. Furthermore, a significant downregulation of many amino acids, lactate levels, and other water-soluble metabolites was noted in HKCs following CXL. CXL is a KC treatment available to patients within certain criteria. Surprisingly, the cellular and molecular mechanisms are considerably understudied limiting our ability for more precise and targeted CXL treatments. In this study, for the first time, we report the effects of CXL on KC metabolism.

Human in vitro Model Reveals the Effects of Collagen Cross-linking on Keratoconus Pathogenesis

Keratoconus (KC) is a corneal thinning disorder that leads to severe vision impairment As opposed to corneal transplantation; corneal collagen crosslinking (CXL) is a relatively non-invasive procedure that leads to an increase in corneal stiffness. In order to evaluate the effect of CXL on human corneal stromal cells in vitro, we developed a 3-D in vitro CXL model, using primary Human corneal fibroblasts (HCFs) from healthy patients and Human Keratoconus fibroblasts (HKCs) from KC patients. Cells were plated on transwell polycarbonate membranes and stimulated by a stable vitamin C. CXL was performed using a mixed riboflavin 0.1% PBS solution followed by UVA irradiation. Our data revealed no significant apoptosis in either HCFs or HKCs following CXL. However, corneal fibrosis markers, Collagen III and α-smooth muscle actin, were significantly downregulated in CXL HKCs. Furthermore, a significant downregulation was seen in SMAD3, SMAD7, and phosphorylated SMADs -2 and -3 expression in CXL HKCs, contrary to a significant upregulation in both SMAD2 and Lysyl oxidase expression, compared to HCFs. Our novel 3-D in vitro model can be utilized to determine the cellular and molecular effects on the human corneal stroma post CXL, and promises to establish optimized treatment modalities in patients with KC.

Assessing the effect of collagen cross-linking by quantitative measurement of corneal viscoelasticity

Assessing the effect of collagen cross-linking by quantitative measurement of corneal viscoelasticity