Fibroblasts' secretome from calcified and non-calcified dermis in Pseudoxanthoma elasticum differently contributes to elastin calcification.

Pseudoxanthoma elasticum (PXE) is a rare disease characterized by ectopic calcification, however, despite the widely spread effect of pro/anti-calcifying systemic factors associated with this genetic metabolic condition, it is not known why elastic fibers in the same patient are mainly fragmented or highly mineralized in clinically unaffected (CUS) and affected (CAS) skin, respectively. Cellular morphology and secretome are investigated in vitro in CUS and CAS fibroblasts. Here we show that, compared to CUS, CAS fibroblasts exhibit: a) differently distributed and organized focal adhesions and stress fibers; b) modified cell-matrix interactions (i.e., collagen gel retraction); c) imbalance between matrix metalloproteinases and tissue inhibitor of metalloproteinases; d) differentially expressed pro- and anti-calcifying proteoglycans and elastic-fibers associated glycoproteins. These data emphasize that in the development of pathologic mineral deposition fibroblasts play an active role altering the stability of elastic fibers and of the extracellular matrix milieu creating a local microenvironment guiding the level of matrix remodeling at an extent that may lead to degradation (in CUS) or to degradation and calcification (in CAS) of the elastic component. In conclusion, this study contributes to a better understanding of the mechanisms of the mineral deposition that can be also associated with several inherited or age-related diseases (e.g., diabetes, atherosclerosis, chronic kidney diseases).

Assessment of the association of myofibroblasts and structural components of the extracellular matrix with histopathological parameters of actinic cheilitis and lower lip squamous cell carcinoma.

BACKGROUND: To evaluate the presence of myofibroblasts (MFs) in the development of lip carcinogenesis, through the correlation of clinical, histomorphometric and immunohistochemical parameters, in actinic cheilitis (ACs) and lower lip squamous cell carcinomas (LLSCCs). METHODS: Samples of ACs, LLSCCs, and control group (CG) were prepared by tissue microarray (TMA) for immunohistochemical TGF-ß, α-SMA, and Ki-67 and histochemical hematoxylin and eosin, picrosirius red, and verhoeff van gieson reactions. Clinical and microscopic data were associated using the Mann-Whitney, Kruskal-Wallis/Dunn, and Spearman correlation tests (SPSS, p < 0.05). RESULTS: ACs showed higher number of α-SMA+ MFs when compared to CG (p = 0.034), and these cells were associated with the vertical expansion of solar elastosis (SE) itself (p = 0.027). Areas of SE had lower deposits of collagen (p < 0.001), immunostaining for TGF-ß (p < 0.001), and higher density of elastic fibers (p < 0.05) when compared to areas without SE. A positive correlation was observed between high-risk epithelial dysplasia (ED) and the proximity of SE to the dysplastic epithelium (p = 0.027). LLSCCs showed a higher number of α-SMA+ MFs about CG (p = 0.034), as well as a reduction in the deposition of total collagen (p = 0.009) in relation to ACs and CG. There was also a negative correlation between the amount of α-SMA+ cells and the accumulation of total collagen (p = 0.041). Collagen and elastic density loss was higher in larger tumors (p = 0.045) with nodal invasion (p = 0.047). CONCLUSIONS: Our findings show the possible role of MFs, collagen fibers, and elastosis areas in the lip carcinogenesis process.

Morphological study of incompetent saphenous veins: apoptosis and ultrastructural changes of smooth muscle cells.

BACKGROUND: Varicose veins affect approximately 25% of people in industrialized countries. METHODS: The study aimed at detecting apoptotic cells and histopathological changes in varicose vein walls. Patients (N.=41) with varicose veins and 30 control group patients were divided into two groups according to their age (younger and older than 50 years). Apoptosis was determined by the TUNEL assay, elastin and collagen IV expression by immunohistochemistry and ultrastructural changes by transmission electron microscopy. RESULTS: The results show that the number of apoptotic cells in the layers of varicose veins increased, in particular in a group of patients aged over 50 years. In the varicose veins as compared to control veins the elastic fibers were found to be thinner, more fragmented and disorderly arranged. Elastin and collagen IV expression was found to decline in the intima and the media of varicose veins in both age groups. Electron microscopy demonstrated hypertrophy and degeneration of smooth muscle cells. Furthermore, cells with ultrastructural feature of apoptosis were noted. In the disorganized and expanded extracellular matrix membrane-bound vesicles, ghost bodies with different size and electron density were observed. Ghost bodies seem to bud off from smooth muscle cells and are likely to be involved in extracellular matrix remodeling as they are seen in close contact with collagen fibers. CONCLUSIONS: The study demonstrates increase of apoptotic cells in the wall of varicose veins along with vein wall structural abnormalities including alterations of smooth muscle cells and decline of elastin and collagen IV expression.

Rosemary extract and rosmarinic acid accelerate elastic fiber formation by increasing the expression of elastic fiber components in dermal fibroblasts.

Ultraviolet (UV)-induced skin photoaging is caused by qualitative and quantitative degradation of dermal extracellular matrix components such as collagen and elastic fibers. Elastic fibers are important for maintaining cutaneous elasticity, despite their small amount in the skin. Previously, microfibril-associated protein 4 (MFAP-4), which is downregulated in photoaging dermis, has been found to be essential for elastic fiber formation by interaction with both fibrillin-1 and elastin, which are core components of elastic fiber. In addition, enhanced cutaneous MFAP-4 expression in a human skin-xenografted murine photoaging model protects against UV-induced photodamage accompanied by the prevention of elastic fiber degradation and aggravated elasticity. We therefore hypothesized that the upregulation of MFAP-4 in dermal fibroblasts may more efficiently accelerate elastic fiber formation. We screened botanical extracts for MFAP-4 expression-promoting activity in normal human dermal fibroblasts (NHDFs). We found that rosemary extract markedly promotes early microfibril formation and mature elastic fiber formation along with a significant upregulation of not only MFAP-4 but also fibrillin-1 and elastin in NHDFs. Furthermore, rosmarinic acid, which is abundant in rosemary extract, accelerated elastic fiber formation via upregulation of transforming growth factor ß-1. This was achieved by the induction of cAMP response element-binding protein phosphorylation, demonstrating that rosmarinic acid represents one of the active ingredients in rosemary extract. Based on the findings in this study, we conclude that rosemary extract and rosmarinic acid represent promising materials that exert a preventive or ameliorative effect on skin photoaging by accelerating elastic fiber formation.

Enhanced Optic Nerve Expansion and Altered Ultrastructure of Elastic Fibers Induced by Lysyl Oxidase Inhibition in a Mouse Model of Marfan Syndrome.

Two major constituents of exfoliation material, fibrillin-1 and lysyl oxidase-like 1 (encoded by FBN1 and LOXL1), are implicated in exfoliation glaucoma, yet their individual contributions to ocular phenotype are minor. To test the hypothesis that a combination of FBN1 mutation and LOXL1 deficiency exacerbates ocular phenotypes, the pan-lysyl oxidase inhibitor ß-aminopropionitrile (BAPN) was used to treat adult wild-type (WT) mice and mice heterozygous for a missense mutation in Fbn1 (Fbn1C1041G/+) for 8 weeks and their eyes were examined. Although intraocular pressure did not change and exfoliation material was not detected in the eyes, BAPN treatment worsened optic nerve and axon expansion in Fbn1C1041G/+ mice, an early sign of axonal damage in rodent models of glaucoma. Disruption of elastic fibers was detected only in Fbn1C1041G/+ mice, which increased with BAPN treatment, as shown by histologic and immunohistochemical staining of the optic nerve pia mater. Transmission electron microscopy showed that Fbn1C1041G/+ mice had fewer microfibrils, smaller elastin cores, and a lower density of elastic fibers compared with WT mice in control groups. BAPN treatment led to elastin core expansion in both WT and Fbn1C1041G/+ mice, but an increase in the density of elastic fiber was confined to Fbn1C1041G/+ mice. LOX inhibition had a stronger effect on optic nerve and elastic fiber parameters in the context of Fbn1 mutation, indicating the Marfan mouse model with LOX inhibition warrants further investigation for exfoliation glaucoma pathogenesis.

Aortic elastic fiber degeneration during acute type a aortic dissection and reverse aortic remodeling.

BACKGROUND: Progression of proximal or distal aortic dilatation is defined as reverse aortic remodeling after surgery for acute type A aortic dissection (ATAAD) that may be dependent on aortic wall degeneration. METHODS: We investigated whether aortic wall degeneration is associated with reverse aortic remodeling leading to aortic reoperation after surgery for ATAAD. Altogether, 141 consecutive patients undergoing surgery for ATAAD at Tampere were evaluated. The resected ascending aortic wall at surgery was processed for 42 degenerative, atherosclerotic and inflammatory histological variables. Patients undergoing aortic reoperations (Redos) were compared with those without aortic reoperations (Controls) during a mean 4.9-year follow-up. RESULTS: Redos were younger than Controls (56 and 66 years, respectively, P < 0.001), and had less frequently previous cardiac surgery prior to ATAAD. Initial surgery encompassed replacement of the ascending aorta in the majority. There were 21 Redos in which one patient died during follow-up as compared with 51 deaths in Controls (log Rank P = 0.002). Histology of the aortic wall revealed increased elastic fiber fragmentation, loss, and disorganization in Redos as compared with Controls (2.1 ± 0.5 vs. 1.9 ± 0.5, Point score unit (PSU), P = 0.043 and 1.7 ± 0.8 vs. 1.2 ± 0.8, PSU, P = 0.016, respectively). Moderate atherosclerosis occurred less often in Redos vs. Controls (9.5% vs. 33%, PSU, P = 0.037, respectively). CONCLUSIONS: According to this exploratory study, histopathology reveals distinctive aortic wall degeneration during ATAAD. Reverse aortic remodeling after ATAAD is associated with the presence of ascending aortic wall elastic fiber fragmentation, loss and disorganization during ATAAD.

The prognostic values of plasma desmosines, crosslinking molecules of elastic fibers, in the disease progression of Moyamoya disease.

Moyamoya disease (MMD) is a cerebrovascular disease which is characterized by the chronic progression of steno-occlusive changes at the terminal portion of internal carotid arteries and the development of "moyamoya vessels." Dysregulation of the extracellular matrix is regarded as a key pathophysiology underlying unique vascular remodeling. Here, we measured the concentration of elastin crosslinkers desmosine and isodesmosine in the plasma of MMD patients. We aimed to reveal its diagnostic values of desmosines in the progression of steno-occlusive lesions. The concentrations of plasma desmosines were determined by liquid chromatography-tandem mass spectrometry. The temporal profiles of steno-occlusive lesions on magnetic resonance angiography were retrospectively evaluated, and the correlation between the progression of steno-occlusive changes in intracranial arteries and plasma desmosines concentrations was further analyzed. Plasma desmosines were significantly higher in MMD patients with disease progression compared to MMD patients without disease progression. Also, the incidence of disease progression was higher in MMD patients with plasma desmosines levels over limit of quantitation (LOQ) than those with plasma desmosines levels below LOQ. In conclusion, plasma desmosines could be potential biomarkers to predict the progression of steno-occlusive changes in MMD patients.

Alterations in epidermal stem cells within the pilosebaceous unit in atrophic acne scars.

BACKGROUND: Atrophic acne scarring is a common sequela of inflammatory acne, causing significant problems for affected patients. Although prolonged inflammation and subsequent aberrant tissue regeneration are considered the underlying pathogenesis, the role of epidermal stem cells, which are crucial to the regeneration of pilosebaceous units, remains unknown. OBJECTIVES: To examine the changes occurring in epidermal stem cells in atrophic acne scars. METHODS: Changes in collagen, elastic fibre and human leukocyte antigen (HLA)-DR expression were analysed in normal skin and inflammatory acne lesions at days 1, 3 and 7 after development. The expression of epidermal stem cell markers and proliferation markers was compared between normal skin and mature atrophic acne scar tissue. RESULTS: In acne lesions, inflammation had invaded into pilosebaceous units over time. Their normal structure had been destructed and replaced with a reduced amount of collagen and elastic fibre. Expression of stem cell markers including CD34, p63, leucine-rich repeat-containing G protein-coupled receptor (LGR)6 and LGR5, which are expressed in the interfollicular epidermis, isthmus and bulge of hair follicles, significantly decreased in atrophic acne scar tissue compared to normal skin. Epidermal proliferation was significantly reduced in scar tissue. CONCLUSIONS: These findings suggest that as inflammatory acne lesions progress, inflammation gradually infiltrates the pilosebaceous unit and affects the resident stem cells. This disruption impedes the normal regeneration of the interfollicular epidermis and adnexal structures, resulting in atrophic acne scars.

Histochemical analysis, smooth muscle immunolocalization and volumetric density of the elastic system fibres of the ostrich (Struthio camelus) phallus.

There are few scientific reports on the histology of the phallus of ratite birds. The aim of this study was to conduct a histochemical analysis to determine the distribution of smooth muscle cells and the volumetric density (Vv) of the fibres of the elastic system in the ostrich phallus. Adult ostriches, 14 months old, were used. The phalluses were fixed in Bouin's solution and then transferred to a buffered formalin solution. They were then processed using standard histological stains for paraffin and slices were obtained. The following techniques were performed: HE, Picrosirius red, Alcian Blue at pH 1.0 and 2.5. The Periodic acid-Schiff reagent and Weigert's Resorcin-Fuchsin with previous oxidation were performed. The M42 test system was used to quantify the elastic system fibres. For immunohistochemical analysis, an anti alpha smooth muscle actin monoclonal antibody was used. The surface of the phallus is covered by a non-keratinized stratified squamous epithelium, which becomes stratified cylindrical in the region of the spermatic sulcus. No glands associated with the connective tissue were observed. The Vv of the elastic system fibres was 4.75%. Smooth muscle cells were visualized only in the walls of blood vessels through immunostaining, with an absence in the lymphatic sinuses. Despite similarities with other birds, such as the presence of a fibrous external axis, a lymphatic core, and a spermatic groove, the ostrich phallus shows marked differences, including the absence of an elastic core, a non-keratinized lining epithelium, and the absence of glands throughout its extension.

Stereological comparison of smooth muscle, collagen, and elastic fibers of the clitoris and glans penis in young adults.

OBJECTIVE: To compare the collagen, elastic fibers, and smooth muscle content of the clitoris and the glans penis in young adults. MATERIALS AND METHODS: The clitoris and the glans penis of six women and six men (mean age 25±3) who died as a result of accidents were excised. The samples were placed under a formaldehyde solution and histologically processed. Masson's trichrome and Weigert's resorcin-fuchsin stain was used to highlight the elastic fibers, smooth muscle, and collagen. Stereological analysis was conducted in 5 random fields of 5 slides for each sample. For statistical analysis, the unpaired t-test was used to compare values between groups, and a value of P<0.05 was considered as significant for all analyses. RESULTS: Stereology revealed a mean smooth muscle content of 35.84±6.46% and 31.64±4.74% for the clitoris and glans penis, respectively, while it also revealed collagen content of 26.11±7.41% and 28.44±3.55% and elastic fibers content of 24.12±4.34% and 30.97±6.13% for the clitoris and glans penis, respectively. The statistical analysis showed no significant differences between them. CONCLUSION: Regardless of anatomical differences, the volumetric density of collagen, elastic fibers, and smooth muscle were similar for the clitoris and glans penis in young adults, a feature possibly explained by their embryology.

Evaluation of microneedling depth of penetration in management of atrophic acne scars: a split-face comparative study.

BACKGROUND: Microneedling is a technique of repeated puncturing or drilling of the skin to induce repair and collagen induction. There are many reported important factors determining the efficacy of microneedling treatment. The extent of injury needed to produce the desired effect in each condition is one of these important factors. OBJECTIVES: We designed the present split-face comparative study to evaluate the use and effectiveness of two different depths of penetration of Dermapen needles in the management of atrophic postacne scars. PATIENTS AND METHODS: The present study involved 14 subjects with atrophic postacne scars. In each patient, both sides of the face were treated with six sessions of microneedling, using Dermapen at 2-week intervals. A split-face study design was performed. The right (Rt) side of the face was treated with Dermapen using 2.5 mm needle length, while the left (Lt) side was treated using 1.5 mm needle length. RESULTS: There was a significantly better percentage of improvement of acne scars on the Rt side of the face compared to the Lt side (P = 0.02) after six sessions. Both sides of the face showed improvement of collagen bundles and elastic fibers characteristics after six sessions. CONCLUSIONS: The use of 2.5 mm depth proved to be more effective both clinically and histologically in the management of atrophic postacne scars.

Remodeling of murine vaginal smooth muscle function with reproductive age and elastic fiber disruption.

Advanced maternal age during pregnancy is associated with increased risk of vaginal tearing during delivery and maladaptive postpartum healing. Although the underlying mechanisms of age-related vaginal injuries are not fully elucidated, changes in vaginal microstructure may contribute. Smooth muscle cells promote the contractile nature of the vagina and contribute to pelvic floor stability. While menopause is associated with decreased vaginal smooth muscle content, whether contractile changes occur before the onset of menopause remains unknown. Therefore, the first objective of this study was to quantify the active mechanical behavior of the murine vagina with age. Further, aging is associated with decreased vaginal elastin content. As such, the second objective was to determine if elastic fiber disruption alters vaginal contractility. Vaginal samples from mice aged 2-14 months were used in maximum contractility experiments and biaxial extension-inflation protocols. To evaluate the role of elastic fibers with age, half of the vaginal samples were randomly allocated to enzymatic elastic fiber disruption. Contractile potential decreased and vaginal material stiffness increased with age. These age-related changes in smooth muscle function may be due, in part, to changes in microstructural composition or contractile gene expression. Furthermore, elastic fiber disruption had a diminished effect on smooth muscle contractility in older mice. This suggests a decreased functional role of elastic fibers with age. Quantifying the age-dependent mechanical contribution of smooth muscle cells and elastic fibers to vaginal properties provides a first step towards better understanding how age-related changes in vaginal structure may contribute to tissue integrity and healing. STATEMENT OF SIGNIFICANCE: Advanced maternal age at the time of pregnancy is linked to increased risks of vaginal tearing during delivery, postpartum hemorrhaging, and the development of pelvic floor disorders. While the underlying causes of increased vaginal injuries with age and associated pathologies remain unclear, changes in vaginal microstructure, such as elastic fibers and smooth muscle cells, may contribute. Menopause is associated with fragmented elastic fibers and decreased smooth muscle content; however, how reproductive aging affects changes in the vaginal composition and the mechanical properties remains unknown. Quantifying the mechanical contribution of smooth muscle cells and elastic fibers to vaginal properties with age will advance understanding of the potential structural causes of age-related changes to tissue integrity and healing.

Pulmonary fibroelastosis - A review.

Elastin is a long-lived fibrous protein that is abundant in the extracellular matrix of the lung. Elastic fibers provide the lung the characteristic elasticity during inhalation with recoil during exhalation thereby ensuring efficient gas exchange. Excessive deposition of elastin and other extracellular matrix proteins reduces lung compliance by impairing ventilation and compromising gas exchange. Notably, the degree of elastosis is associated with the progressive decline in lung function and survival in patients with interstitial lung diseases. Currently there are no proven therapies which effectively reduce the elastin burden in the lung nor prevent dysregulated elastosis. This review describes elastin's role in the healthy lung, summarizes elastosis in pulmonary diseases, and evaluates the current understanding of elastin regulation and dysregulation with the goal of guiding future research efforts to develop novel and effective therapies.

Elastic fibers define embryonic tissue stiffness to enable buckling morphogenesis of the small intestine.

During embryonic development, tissues must possess precise material properties to ensure that cell-generated forces give rise to the stereotyped morphologies of developing organs. However, the question of how material properties are established and regulated during development remains understudied. Here, we aim to address these broader questions through the study of intestinal looping, a process by which the initially straight intestinal tube buckles into loops, permitting ordered packing within the body cavity. Looping results from elongation of the tube against the constraint of an attached tissue, the dorsal mesentery, which is elastically stretched by the elongating tube to nearly triple its length. This elastic energy storage allows the mesentery to provide stable compressive forces that ultimately buckle the tube into loops. Beginning with a transcriptomic analysis of the mesentery, we identified widespread upregulation of extracellular matrix related genes during looping, including genes related to elastic fiber deposition. Combining molecular and mechanical analyses, we conclude that elastin confers tensile stiffness to the mesentery, enabling its mechanical role in organizing the developing small intestine. These results shed light on the role of elastin as a driver of morphogenesis that extends beyond its more established role in resisting cyclic deformation in adult tissues.

Dill Extract Preserves Dermal Elastic Fiber Network and Functionality: Implication of Elafin.

INTRODUCTION: Elastic skin fibers lose their mechanical properties during aging due to enzymatic degradation, lack of maturation, or posttranslational modifications. Dill extract has been observed to increase elastin protein expression and maturation in a 3D skin model, to improve mechanical properties of the skin, to increase elastin protein expression in vascular smooth muscle cells, to preserve aortic elastic lamella, and to prevent glycation. OBJECTIVE: The aim of the study was to highlight dill actions on elastin fibers during aging thanks to elastase digestion model and the underlying mechanism. METHODS: In this study, elastic fibers produced by dermal fibroblasts in 2D culture model were injured by elastase, and we observed the action of dill extract on elastic network by elastin immunofluorescence. Then action of dill extract was examined on mice skin by injuring elastin fibers by intradermal injection of elastase. Then elastin fibers were observed by second harmonic generation microscopy, and their functionality was evaluated by oscillatory shear stress tests. In order to understand mechanism by which dill acted on elastin fibers, enzymatic tests and real-time qPCR on cultured fibroblasts were performed. RESULTS: We evidence in vitro that dill extract is able to prevent elastin from elastase digestion. And we confirm in vivo that dill extract treatment prevents elastase digestion, allowing preservation of the cutaneous elastic network in mice and preservation of the cutaneous elastic properties. Although dill extract does not directly inhibit elastase activity, our results show that dill extract treatment increases mRNA expression of the endogenous inhibitor of elastase, elafin. CONCLUSION: Dill extract can thus be used to counteract the negative effects of elastase on the cutaneous elastic fiber network through modulation of PI3 gene expression.

Association of NOTCH3 With Elastic Fiber Dispersion in the Infrarenal Abdominal Aorta of Cynomolgus Monkeys.

BACKGROUND: The regional heterogeneity of vascular components and transcriptomes is an important determinant of aortic biology. This notion has been explored in multiple mouse studies. In the present study, we examined the regional heterogeneity of aortas in nonhuman primates. METHODS: Aortic samples were harvested from the ascending, descending thoracic, suprarenal, and infrarenal regions of young control monkeys and adult monkeys with high fructose consumption for 3 years. The regional heterogeneity of aortic structure and transcriptomes was examined by histological and bulk RNA sequencing analyses, respectively. RESULTS: Immunostaining of CD31 and αSMA (alpha-smooth muscle actin) revealed that endothelial and smooth muscle cells were distributed homogeneously across the aortic regions. In contrast, elastic fibers were less abundant and dispersed in the infrarenal aorta compared with other regions and associated with collagen deposition. Bulk RNA sequencing identified a distinct transcriptome related to the Notch signaling pathway in the infrarenal aorta with significantly increased NOTCH3 mRNA compared with other regions. Immunostaining revealed that NOTCH3 protein was increased in the media of the infrarenal aorta. The abundance of medial NOTCH3 was positively correlated with the dispersion of elastic fibers. Adult cynomolgus monkeys with high fructose consumption displayed vascular wall remodeling, such as smooth muscle cell loss and elastic fiber disruption, predominantly in the infrarenal region. The correlation between NOTCH3 and elastic fiber dispersion was enhanced in these monkeys. CONCLUSIONS: Aortas of young cynomolgus monkeys display regional heterogeneity of their transcriptome and the structure of elastin and collagens. Elastic fibers in the infrarenal aorta are dispersed along with upregulation of medial NOTCH3.

Qualitative Assessment of Collagen and Elastic Fibers in Oral Submucous Fibrosis (OSMF), OSMF with Dysplasia and Oral Squamous Cell Carcinoma Arising from OSMF: A Histochemical Study.

BACKGROUND: Oral submucous fibrosis (OSMF) is a habit related potentially malignant disorder seen mainly in South Asian people. The malignancy arising from OSMF has been regarded as low grade with better outcome. The present study was orchestrated to histochemically analyze collagen and elastic fibres in OSMF without dysplasia, OSMF with dysplasia and OSMF turning malignant. MATERIALS AND METHODS: 100 cases (80 cases and 20 healthy controls) were included after obtaining clearance from ethical committee. All cases were subjected to Van Gieson staining for collagen and a novel simple method for elastic fibres (Orcein staining). Data were analyzed using SPSS software. RESULTS: Controls showed haphazard arrangement of collagen and elastic fibres. The collagen bundles were parallelly arranged in OSMF without dysplasia and OSMF with dysplasia; the collagen was however haphazard in cases of OSMF turning malignant. As with collagen, elastic fibres were also haphazardly arranged in the control group; in contrast, the elastic fibres were predominantly arranged in a criss-cross pattern in the other study groups. The difference in orientation and density among the groups was statistically significant. CONCLUSION: With advancement of stage there is increased collagenization of OSMF, as the condition acquires dysplastic changes and turns malignant, microenvironment alters resulting in increased activity of collagenases. However, the arrangement of more resistant elastic fibres depicts the better outcome, once OSMF shows malignant transformation, limiting locoregional and distant spread.