Collagen Cross-linking Research Articles

Efficacy of enzyme‑induced collagen crosslinking on porcine cornea.

The purpose of the present study was to investigate the effect of a new crosslinking (CXL) method, induced by enzymes, on porcine corneas. Corneal strip (10x3 mm) pairs obtained from 60 fresh porcine eyes were harvested and divided into four groups, Groups A-D. Each pair of corneal strips was incised from the central part of the same cornea; one was incubated in transglutaminase (Tgase) solution (microbial Tgase 2 produced by tissue engineering) and the other remained untreated as a control. CXL strips of Groups A-D were incubated with 2, 1, 0.5 and 0.25 U/ml Tgase solution, respectively at 37˚C for 30 min. After that, tensile strain measurements were performed for all strips. One cornea from each group was chosen randomly for hematoxylin and eosin, and Masson staining to identify histological morphology changes. The elastic modulus of treated corneas of Groups A-D were 6.56±2.93, 4.72±1.29, 5.24±2.13 and 3.48±1.60 MPa (mean ± SD), respectively at a strain of 20%, and had a 66, 43, 36 and -6% increase compared with those of their control strips. Compared with the control strips, the elastic modulus of the treated strips significantly increased in Groups A-C. The central corneal thickness of the treated corneas in Groups A-D were 1.54±0.14, 1.41±0.15, 1.47±0.11 and 1.43±0.13 µm, respectively; however, there was not a statistically significant difference compared with the control group. No reduction in corneal transparency was observed, and no obvious abnormalities were found in corneal morphology. CXL mediated by enzymes can lead to a notable enhancement in the biomechanical characteristics of the cornea while maintaining its structural integrity. Enzyme-induced CXL could be a new generation CXL method for strengthening the cornea.

Implantation of Adipose-Derived Mesenchymal Stromal Cells (ADSCs)-Lining Prosthetic Graft Promotes Vascular Regeneration in Monkeys and Pigs

Background:Current replacement procedures for stenosis or occluded arteries using prosthetic grafts have serious limitations in clinical applications, particularly, endothelialization of the luminal surface is a long-standing unresolved problem.Method:We produced a cell-based hybrid vascular graft using a bioink engulfing adipose-derived mesenchymal stromal cells (ADSCs) and a 3D bioprinting process lining the ADSCs on the luminal surface of GORE-Tex grafts. The hybrid graft was implanted as an interposition conduit to replace a 3-cm-long segment of the infrarenal abdominal aorta in Rhesus monkeys.Results:Complete endothelium layer and smooth muscle layer were fully developed within 21 days post-implantation, along with normalized collagen deposition and crosslinking in the regenerated vasculature in all monkeys. The regenerated blood vessels showed normal functionality for the longest observation of more than 1650 days. The same procedure was also conducted in miniature pigs for the interposition replacement of a 10-cm-long right iliac artery and showed the same long-term effective and safe outcome.Conclusion:This cell-based vascular graft is ready to undergo clinical trials for human patients.

Inhibiting Lysyl Oxidases prevents pathologic cartilage calcification

Lysyl oxidases (LOX(L)) are enzymes that catalyze the formation of cross-links in collagen and elastin fibers during physiologic calcification of bone. However, it remains unknown whether they may promote pathologic calcification of articular cartilage, an important hallmark of debilitating arthropathies. Here, we have studied the possible roles of LOX(L) in cartilage calcification, related and not related to their cross-linking activity. We first demonstrated that inhibition of LOX(L) by β-aminoproprionitrile (BAPN) significantly reduced calcification in murine and human chondrocytes, and in joint of meniscectomized mice. These BAPN’s effects on calcification were accounted for by different LOX(L) roles. Firstly, reduced LOX(L)-mediated extracellular matrix cross-links downregulated Anx5, Pit1 and Pit2 calcification genes. Secondly, BAPN reduced collagen fibrotic markers Col1 and Col3. Additionally, LOX(L) inhibition blocked chondrocytes hypertrophic differentiation (Runx2 and COL10), pro-inflammatory IL-6 release and reactive oxygen species (ROS) production, all triggers of chondrocyte calcification. Through unbiased transcriptomic analysis we confirmed a positive correlation between LOX(L) genes and genes for calcification, hypertrophy and extracellular matrix catabolism. This association was conserved throughout species (mouse, human) and tissues that can undergo pathologic calcification (kidney, arteries, skin). Overall, LOX(L) play a critical role in the process of chondrocyte calcification and may be therapeutic targets to treat cartilage calcification in arthropathies.

Dysregulation of extracellular matrix and Lysyl Oxidase in Ehlers-Danlos syndrome type IV skin fibroblasts

BackgroundEhlers-Danlos syndrome Type IV (aka Vascular Ehlers Danlos, or vEDS) is a dominantly inherited mutation in the Collagen 3A1 gene (COL3A1). The disease is characterized by tissue friability and age-related susceptibility to arterial aneurysm, dissection and rupture as well as uterine and bowl tears. These clinical manifestations result in major surgical intervention and decreased life expectancy. Understanding how mutations in COL3A1 impact the structure and function of the extracellular matrix (ECM) is important to managing the disease and finding treatments.ResultsSkin fibroblasts from vEDS subjects heterozygous for the p.G588S pathogenic variant in the COL3A1 gene and a normal individual were cultured and studied. Proteomics analysis identified dozens of upregulated proteins related to extracellular matrix dysregulation that is characteristic of fibrosis. Gene expression libraries from cultured primary fibroblasts were screened for messenger RNA (mRNA) markers of ECM degradation. The proteomics and targeted gene expression array results were largely consistent with dysregulation of the extracellular matrix in vEDS. The data show upregulation of multiple Collagen proteins and genes, other ECM components, and enzymes related to ECM processing and turn-over. vEDS fibroblasts expressed significantly more cross linked C-Telopeptide of Collagen III (CTXIII) than normal fibroblasts, indicative of Collagen III degradation and turn-over. Further, the expression and activity of Lysyl Oxidase (LOX), an enzyme that initiates covalent cross-linking of soluble collagen and elastin into protease resistant fibers, is elevated in vEDS fibroblasts compared to normal fibroblasts.ConclusionTogether, these findings suggest dysregulated ECM deposition and processing, reminiscent of a state of fibrosis. Therapeutics that target the dysregulated ECM proteins or help replace damaged tissue may improve clinical outcomes.

LOX, but not LOXL2, promotes bone metastasis formation and bone destruction in triple-negative breast cancer

The primary function of the lysyl oxidase (LOX) family, including LOX and its paralogue LOX-like (LOXL)-2, is to catalyze the covalent crosslinking of collagen and elastin in the extracellular matrix. LOX and LOXL2 are also facilitating breast cancer invasion and metastatic spread to visceral organs (lungs, liver) in vivo. Conversely, the contribution of LOX and LOXL2 to breast cancer bone metastasis remains scant. Here, using gene overexpression or silencing strategies, we investigated the role of LOX and LOXL2 on the formation of metastatic osteolytic lesions in animal models of triple negative breast cancer. In vivo, the extent of radiographic metastatic osteolytic lesions in animals injected with LOX-overexpressing [LOX(+)] tumor cells was 3-fold higher than that observed in animals bearing tumors silenced for LOX [LOX(−)]. By contrast, the extent of osteolytic lesions between LOXL2(+) and LOXL2(−) tumor-bearing animals did not differ, and was comparable to that observed with LOX(−) tumor-bearing animals. In situ, TRAP staining of bone tissue sections from the hind limbs of LOX(+) tumor-bearing animals was substantially increased compared to LOX(−), LOXL2(+) and LOXL2(−)-tumor-bearing animals, which was indicative of enhanced active-osteoclast resorption. In vitro, tumor-secreted LOX increased osteoclast differentiation induced by RANKL, whereas LOXL2 seemed to counteract LOX’s pro-osteoclastic activity. Furthermore, LOX (but not LOXL2) overexpression in tumor cells induced a robust production of IL-6, the latter being a pro-osteoclastic cytokine. Based on these findings, we propose a model in which LOX and IL-6 secreted from tumor cells act in concert to enhance osteoclast-mediated bone resorption that, in turn, promotes metastatic bone destruction in vivo.

Glycosaminoglycans and collagen fibril distribution at various depths of the corneal stroma of normal and CXL treated rats

Corneal collagen cross-linking (CXL) is widely used to treat keratoconus and ecstatic corneal disorders. The present studies were carried out to investigate the distribution of glycosaminoglycans (GAGs) and collagen fibril (CF) at different depths of the normal and CXL treated corneal stroma of four week old rats 7 days after standard CXL application. Ten Wistar rats’ corneas were used for the study. The epithelium of the cornea from the left eye of each rat was removed and treated with standard CXL application using riboflavin and Ultraviolet-A (UVA) (3 mW/cm2 for 30 min). The cornea from the right eye was used as the control cornea. The cornea was removed from the eye and processed for transmission electron microscopy. A bottom mounted Quemesa camera was used to capture digital images and these images were analysed using iTEM software. In the control cornea, the GAGs area size was not significantly different in the anterior, middle, and posterior stroma. In the CXL treated rats the GAGs area size gradually increased from the anterior to the posterior stroma whereas the spacing between the GAGs gradually decreased. There were very large GAGs present in the posterior stroma of the CXL treated rats. When comparing the control and CXL cornea, the GAGs area in the CXL cornea was significantly higher and inter-GAGs-spacing was smaller than in the control cornea. In the control cornea, the collagen fibrils diameter was higher in the anterior stroma and lowest in the posterior stroma. In the CXL treated cornea, the CF diameter and the interfibrillar spacing gradually decreased from the anterior to the posterior stroma. On comparison between the control and the CXL treated cornea, the interfibrillar spacing was significantly smaller in the CXL treated cornea than the control cornea in the anterior, middle, and posterior stroma but there was no difference in the diameter. The CXL treatment significantly increased the GAGs area and decreased the inter-GAGs-spacing, and inter–CF–spacing. This could be due to the gradual decline in the availability of riboflavin, UVA, and oxygen in the middle and posterior stroma. Further studies are required to investigate the role of keratan sulphate and chondroitin sulphate by using monoclonal antibodies with immunogold technique.

Safety and efficacy of a combined treatment with corneal collagen cross-linking and lamellar keratoplasty in patients with keratitis

Introduction and Aim: There is a link between an increase in eye infections, environmental deterioration, increased intensity of eye surgery, and the diversification of diagnostic techniques and the development of corneal disease and eye injuries. This study aims to assess the safety and efficacy of a complex treatment for corneal ulcers (keratitis) that combines corneal collagen cross-linking (CXL) with lamellar keratoplasty (LK). Materials and Methods: This study consisted of 28 eyes from 28 patients with keratitis. All patients were divided into the main and control groups; each group had 14 patients (14 eyes). All patients underwent a standard ophthalmological examination, which included visometry without correction, with correction, tonometry, biomicroscopy, and ophthalmoscopy. Results: High postoperative astigmatism was found in 50% of cases (7 eyes) of the control group and in 21.4% of cases (3 eyes) of the main group over the 1-6 months follow-up period. Both groups visual acuity development dynamics were very similar from the three-month to three-year follow-up period, and the average visual acuity with maximal correction was between 0.4 - 0.5. Conclusion: Ultraviolet radiation exposure during CXL increases the graft's strength, the recipient's corneal tissue in the suture area, and the pathological flora's direct bactericidal and virucidal effects, reducing infectious process recurrences.

Fungal Keratitis: Diagnosis, Management, and Recent Advances.

Fungal keratitis is one of the major causes of microbial keratitis that may lead to corneal blindness. Many problems related to diagnosis and therapy are encountered in fungal keratitis, including difficulty in obtaining laboratory diagnoses and the availability and efficacy of antifungal medications. Intensive and prolonged use of antifungal topical preparations may not be enough. The use of antifungal medications is considered the main treatment for fungal keratitis. It is recommended to start antifungal therapy after confirmation of the clinical diagnosis with a smear or positive cultures. Topical application of antifungal medications is a mainstay for the treatment of fungal keratitis; however, systemic, intra-stromal, or intra-cameral routes may be used. Therapeutic keratoplasty is the main surgical procedure approved for the management of fungal keratitis with good success rate. Intrastromal corneal injection of antifungal medications may result in steady-state drug levels within the corneal tissue and prevent intervals of decreased antifungal drug concentration below its therapeutic level. In cases of severe fungal keratitis with deep stromal infiltration not responding to treatment, intracameral injection of antifungal agents may be effective. Collagen cross-linking has been proposed to be beneficial for cases of fungal keratitis as a stand-alone therapy or as an adjunct to antifungal medications. Although collagen cross-linking has been extensively studied in the past few years, its protocol still needs many modifications to optimize UV fluence levels, irradiation time, and concentration of riboflavin to achieve 100% microbial killing.

Copper chelation reduces early collagen deposition and preserves saliva secretion in irradiated salivary glands

Radiation therapy is a first-line treatment for head and neck cancer; however, it typically leads to hyposalivation stemming from fibrosis of the salivary gland. Current strategies to restore glandular function are dependent on the presence of residual functional salivary gland tissue, a condition commonly not met in patients with extensive fibrotic coverage of the salivary gland resulting from radiation therapy. Fibrosis is defined by the pathological accumulation of connective tissue (i.e., extracellular matrix) and excessive deposition of crosslinked (fibrillar) collagen that can impact a range of tissues and given that collagen crosslinking is necessary for fibrosis formation, inhibiting this process is a reasonable focus for developing anti-fibrotic therapies. Collagen crosslinking is catalyzed by the lysyl oxidase family of secreted copper-dependent metalloenzymes, and since that copper is an essential cofactor in all lysyl oxidase family members, we tested whether localized delivery of a copper chelator into the submandibular gland of irradiated mice could suppress collagen deposition and preserve the structure and function of this organ. Our results demonstrate that transdermal injection of tetrathiomolybdate into salivary glands significantly reduced the early deposition of fibrillar collagen in irradiated mice and preserved the integrity and function of submandibular gland epithelial tissue. Together, these studies identify copper metabolism as a novel therapeutic target to control radiation induced damage to the salivary gland and the current findings further indicate the therapeutic potential of repurposing clinically approved copper chelators as neoadjuvant treatments for radiation therapy.

Pan-Cancer Analysis of the LOX Family Reveals that LOX Affects Tumor Prognosis by Affecting Immune Infiltration.

The lysyl oxidase (LOX) gene family encodes for a group of copper-dependent enzymes that play a crucial role in the cross-linking of collagen and elastin fibers in the extracellular matrix (ECM). Dysregulation of LOX gene expression has been implicated in various pathological conditions, including cancer. Several studies have shown that the LOX gene family is involved in cancer progression and metastasis. The goal of this article is to conduct a comprehensive analysis of the LOX family's role in pan-cancer multiplexes. We utilized pan-cancer multi-omics sequencing data from TCGA to investigate the relationship between LOX family genes and tumors at four different levels: mutation, copy number variation, methylation, and gene expression. In addition, we also examined the relationship between LOX family genes and tumors at the cell line level using tumor cell line sequencing data from CCLE. Taking into account the impact of LOX family genes on lung cancer, we developed a LOX family lung cancer prognostic model to forecast the disease's prognosis. Our findings revealed that LOXL2 had the highest mutation frequency in tumors, while all four LOX family genes experienced some degree of copy number variation in diverse tumors. We observed that LOX, LOXL1 to LOXL3 were predominantly highly expressed in tumors including LUAD. The expression trends of LOX and LOXL1 to LOXL3 were consistent across tumor cell lines, but differed somewhat from LOXL4. Utilizing 25 LOX family-related genes, we constructed a LOX family prognostic model that performed well in predicting the prognosis of lung cancer. Through pan-cancer analysis, we gain further knowledge of the role of LOX family genes in different tumors, offering a novel pathway for future research into the relationship between LOX family genes and tumors.

Ab. No. 161 An Immediate Effect of Active Release Technique [ART] Versus Graston Technique [GT] on Neck Pain in Individuals Having Myofascial Trigger Point of Upper Trapezius

Introduction: Myofascial trigger points (MTrPs) are hyperirritable, palpable nodules found along taut bands of muscle fibers. It has been estimated that 85% of people who come to pain in clinics have trigger points in the neck, and they occur more often in women than in men. MTrPs are classified clinically into active and latent. latent trigger points produce pain when palpated and cause restriction of movement. ART can identify and reduce fibrous adhesions, microscopic tissue injury and inflammation. Graston technique is an Instrument assisted soft Tissue Mobilization method. It helps the healing process by intentional micro-trauma, breaking down collagen cross-linkage, increasing blood flow and cellular regeneration. The purpose is to compare the immediate Effect of Active Release Technique Methods: 30 participants age (18-34) years were included, 15 per group. Ethical approval was taken. Groups A received ART & Group B - Graston technique. Pre and post evaluation was done using pain pressure threshold (PPT) and Visual Analogue Scale (VAS) for pain. Result: Within group analysis showed Group A PPT (p>0.05), VAS (p<0.05) and Group B PPT p>0.05), VAS (< 0.05). Between group showed statistical significance in PPT (p<0.05) but did not in VAS (p>0.05). Conclusion: Graston technique is more effective in increasing PPT. Implications: Graston technique can be used to get faster results.

Keratoconus: imaging modalities and management.

Keratoconus (KCN) is characterized by gradual thinning and steepening of the cornea, which can lead to significant vision problems owing to high astigmatism, corneal scarring, or even corneal perforation. The detection of KCN in its early stages is crucial for effective treatment. In this review, we describe current advances in the diagnosis and treatment of KCN. This narrative review focuses on recent advancements in the diagnosis and treatment of KCN, especially evolving approaches and strategies. To ensure the inclusion of the most recent literature, relevant publications discussing advanced imaging techniques and treatment options for KCN were extensively gathered from the PubMed/MEDLINE and Google Scholar databases. The following index terms and keywords were used for the online search: keratoconus, diagnosis of keratoconus, advances in the diagnosis of keratoconus, topography or tomography, anterior segment optical coherence tomography, treatment of keratoconus, advances in the treatment of keratoconus, collagen crosslinking, intrastromal ring, keratoplasty, and new techniques in keratoconus. Various screening methods such as corneal topography, tomography, anterior segment optical coherence tomography, and assessment of corneal biomechanics have been developed to identify KCN in its early stages. After diagnosis, KCN management focuses on preventing disease progression. Corneal collagen crosslinking is a minimally invasive treatment that can slow or stop the progression of the condition. Recent research has also explored the use of copper sulfate eye drops (IVMED-80) as a noninvasive treatment to prevent the progression of KCN. Current treatment options for visual improvement include scleral lenses, intracorneal ring segments, corneal allogeneic intrastromal ring segments, and deep anterior lamellar keratoplasty. Recently, novel alternative procedures, such as isolated Bowman layer transplantation, either as a corneal stromal inlay or onlay, have demonstrated encouraging outcomes. Artificial intelligence has gained acceptance for providing best practices for the diagnosis and management of KCN, and the science of its application is contentiously debated; however, it may not have been sufficiently developed. Early detection and advancements in screening methods using current imaging modalities have improved diagnosis of KCN. Improvement in the accuracy of current screening or diagnostic tests and comparison of their validities are achievable by well-designed, large-scale, prospective studies. The safety and effectiveness of emerging treatments for KCN are currently being investigated. There is an ongoing need for studies to track progress and evaluate clinicians' knowledge and practices in treating patients with KCN. Artificial intelligence capabilities in management approach considering the currently available imaging modalities and treatment options would best benefit the patient.

Exploring the effects of homocysteine metabolism in osteoporosis management in Indian adult females.

Objectives: Women are at a significantly higher risk of osteoporotic fractures, largely due to progressive bone demineralization and impaired bone microarchitecture. Low bone mineral density (BMD) is a common condition in women worldwide. Disrupted homocysteine (Hcy) metabolism has been linked to reduced BMD and increased risk of osteoporotic fractures. Hyperhomocysteinemia (Hhcy) affects osteoblast and osteoclast activity, interferes with collagen cross-linking in the extracellular matrix, and has a detrimental effect on bone health. This study aimed to establish the association between hematological and biochemical parameters and osteoporosis in adult females. Methods: We measured Hcy, creatinine, uric acid (UA), vitamin B12, and vitamin D levels. Significantly elevated Hcy (27.322 ± 0.816 vs 10.152 ± 0.381 µmol/L), creatinine (0.670 ± 0.012 vs 0.587 ± 0.011 mg/dL), and UA (5.118 ± 0.083 vs 2.786 ± 0.060 mg/dL) were found in osteoporotic females, while reduced concentrations of vitamin B12 (148.883 ± 2.192 vs 294.14 ± 6.505 pg/mL) and vitamin D (24.98 ± 0.621 vs 33.7 ± 0.652 ng/mL) were observed. Results: Hematological parameters were found differentially expressed in osteoporotic females. Elevated Hcy levels, combined with reduced vitamin B12 and vitamin D, were strongly associated with decreased BMD and a higher susceptibility to osteoporotic fractures. Women with increased Hcy levels also had lower T-scores compared to those without Hhcy. Conclusions: These findings suggest that Hcy plays a critical role in bone resorption and osteoporotic fractures. Regulating Hcy metabolism may serve as an effective therapeutic strategy for managing bone resorption and osteoporosis. We hypothesize that elevated Hcy levels are closely related to low BMD and an increased risk of osteoporosis.

Effect of Corneal Collagen Crosslinking with Riboflavin in Patients of Progressive Keratoconus: A Prospective Cohort Study

Introduction: Keratoconus is a non inflammatory disease characterised by progressive corneal thinning that results in a cone-like ectasia, irregular astigmatism, and decreased vision. Advanced disease can result in corneal scarring and blindness, necessitating penetrating keratoplasty. The only treatment believed to have the ability to stop or decrease the progression of keratoconus is Collagen Cross-Linking (CXL). Aim: To evaluate the efficacy of corneal collagen cross-linking with riboflavin (C3R) in patients with keratoconus by studying the outcomes in terms of Uncorrected Visual Acuity (UCVA), Best Corrected Visual Acuity (BCVA), manifest refraction, and corneal topography. Materials and Methods: A hospital-based prospective cohort study was conducted at the Regional Institute of Ophthalmology, Government Medical College, Thiruvananthapuram, Kerala, India from June 2017 to June 2018. Fifteen eyes of 12 patients who underwent corneal collagen cross-linking for progressive keratoconus were included. Routine ophthalmological examinations was done for all patients. Patients under the age of 35 years with progressive keratoconus, an average K value not exceeding 60 D, and who had not undergone any other surgical modalities of treatment were included in the study. Patients with scarred cornea or hydrops, active ocular infection, or pachymetry <400 µm were excluded. Detailed informed consent was obtained from all patients before surgery. Preoperative UCVA, BCVA cylinder, Spherical Equivalent (SE), and topography values were obtained. All patients were followed-up at three and six months postoperative period to look for changes in UCVA, BCVA, manifest refraction, and corneal topography. Data were expressed as the mean±Standard Deviation (SD) pre- and postoperatively and analysed statistically using the paired t-test, with p<0.05 considered statistically significant. Analysis was conducted using Statistical Package for Social Sciences (SPSS) version 16.0. Results: On follow-up at six months post-treatment, UCVA (log MAR) improved from 0.87±0.40 to 0.65±0.35 (p-value=0.001), and BCVA (log MAR) changed from 0.37±0.21 to 0.17±0.22 (p-value=0.001). K max decreased from 54.90±6.05 to 52.67±5.02 (p-value=0.001), and K min decreased from 48.14±4.24 to 45.93±4.07 (p-value=0.001). Astigmatism and SE also showed statistically significant improvement post-procedure (p-value 0.001 and 0.002, respectively). Astigmatism reduced from -3.50±1.58 to -2.90±1.37. SE reduced from -4.93±3.92 to -4.28±3.48. No significant complications were noted in any of the patients. Conclusion: The CXL is an effective procedure for stabilising the progression of keratoconus, thus reducing the burden of preventable blindness and the need for keratoplasty.

Deep learning in early keratoconus/corneal ectasia

Machine learning algorithms have been gaining tremendous relevance in the last decade in more or less all disciplines in medicine. In most cases supervised learning techniques are used to perform classification or graduation of diseases. Early detection of keratoconus in a screening environment is crucial as in this stage several therapeutic options such as collagen crosslinking or intracorneal ring implantation are available which could prevent further progression of the disease. In this talk we focus on new techniques of automatic identification of ectatic corneal diseases such as keratoconus in an early stage based on anterior segment optical coherence measurements with the Casia 2 tomographer. A neural network approach is implemented and fed with data of corneal front surface height and curvature, back surface height and curvature, and thickness data (in total 5 data layers) and the similarity of the measurement to normal or abnormal situation is provided. Performance data are shown for our training, validation, and test dataset.

Advances in Noninvasive Molecular Imaging Probes for Liver Fibrosis Diagnosis.

Liver fibrosis is a wound-healing response to chronic liver injury, which may lead to cirrhosis and cancer. Early-stage fibrosis is reversible, and it is difficult to precisely diagnose with conventional imaging modalities such as magnetic resonance imaging, positron emission tomography, single-photon emission computed tomography, and ultrasound imaging. In contrast, probe-assisted molecular imaging offers a promising noninvasive approach to visualize early fibrosis changes invivo, thus facilitating early diagnosis and staging liver fibrosis, and even monitoring of the treatment response. Here, the most recent progress in molecular imaging technologies for liver fibrosis is updated. We start by illustrating pathogenesis for liver fibrosis, which includes capillarization of liver sinusoidal endothelial cells, cellular and molecular processes involved in inflammation and fibrogenesis, as well as processes of collagen synthesis, oxidation, and cross-linking. Furthermore, the biological targets used in molecular imaging of liver fibrosis are summarized, which are composed of receptors on hepatic stellate cells, macrophages, and even liver collagen. Notably, the focus is on insights into the advances in imaging modalities developed for liver fibrosis diagnosis and the update in the corresponding contrast agents. In addition, challenges and opportunities for future research and clinical translation of the molecular imaging modalities and the contrast agents are pointed out. We hope that this review would serve as a guide for scientists and students who are interested in liver fibrosis imaging and treatment, and as well expedite the translation of molecular imaging technologies from bench to bedside.

Expanding limits of artificial enzymes: unprecedented catalysis by an oxidase nanozyme in activating a structural protein for covalent crosslinking and conferring remarkable proteolytic resistance.

Nature has endowed us with some complex enzymes capable of utilizing proteins as their substrates to generate functional proteins through post-translational modification. However, nanozymes' interplay with proteins as substrates is scarce, with their chemistry predominantly established using only small molecule substrates, featuring a significant gap in this area. Due to the huge prospects of nanozymes in biotechnological and therapeutic interventions, studies establishing the unexplored roles of nanozymes in the biological environments and their interplay beyond small molecule substrates warrant immediate attention. In this study, we unveil the unprecedented role of a Mn-based oxidase nanozyme (MnN) in activating a structural protein, collagen, and covalently crosslinking its tyrosine residues with only a trace amount of tannic acid (TA) without compromising its triple-helical structural integrity. While therapeutic applications demand materials prepared from collagen, the current chemical and physical crosslinking of collagen often presents significant challenges such as toxicity, denaturation, or high costs. MnN lucidly accomplishes crosslinking interplay at its 101 facets using oxygen as a co-substrate under mild conditions. This process takes advantage of MnN being active at mild acidic pH where collagen preferentially exists as a soluble triple helix (monomeric form), exposing functionalities and enhancing the crosslinking degree. Importantly, this reaction also confers 100% resistance to collagenase attack on the collagen tendon-derived biological material. The catalyzed TA-tyrosine linkage in the telopeptide region of collagen probably impedes the initial recognition step of collagenase, providing robust protection against its degradative action. Our study not only expands the repertoire of nanozymes' substrates beyond the existing library of small molecules but also establishes a significant step toward designing a gold standard for collagen crosslinking. With biomedical applications demanding biomaterials derived from protein scaffolds with preserved structural integrity, our investigation bridges the gap between nanozymes' chemistry and crosslinking proteins, opening exciting prospects for biomaterial development.

Application of Rapid Corneal Collagen Crosslinking (CXL) Combined with FS-LASIK in the Diagnosis and Treatment of Myopia

Application of Rapid Corneal Collagen Crosslinking (CXL) Combined with FS-LASIK in the Diagnosis and Treatment of Myopia

Alternative methods of surgical treatment of keratoconus

The introduction of early diagnostic methods for keratoconus into clinical practice has become the basis for the development of surgical treatment techniques for this pathology, such as corneal collagen crosslinking and interlamellar keratoplasty with implantation of intrastromal segments. The article analyzes the results of research by Russian and foreign specialists in these areas and presents the data on the combination of SMILE surgery and corneal crosslinking, the Rome protocol of corneal crosslinking, modifications of interlamellar keratoplasty, the use of femtosecond laser technologies, and some pilot studies. Modern requirements for ophthalmological care require a personalized approach to each patient, and therefore the surgeon should have a wide range of surgical methods of treatment applicable to different patient cohorts. The described methods of treatment, according to the authors, are the most promising.

Comparative analysis of clinical and functional parameters in patients with stable stage II-III keratoconus after implantation of corneal segments and fitting of scleral rigid contact lenses

This study compares the effect of implantation of intrastromal corneal ring segment (ICRS) and fitting of individual scleral rigid contact lenses (RCLs) on visual functions in patients with stage II-III KC after previously performed corneal collagen cross-linking. The Helmholtz National Medical Research Center of Eye Diseases examined and treated 34 patients (69 eyes) aged 18 to 33 years with stage II-III KC. The study included patients who had previously undergone standard corneal collagen cross-linking. Depending on the type of optical correction, the patients were divided into two groups: patients in group 1 underwent ICRS implantation using a femtosecond laser; patients in group 2 were fitted with individual scleral RCLs. Improvement in clinical and functional parameters was observed in both groups. A higher clinical and functional result was achieved in group 2. For patients with stable stage II-III KC, it is advisable to recommend fitting of individual scleral RCLs for visual rehabilitation.