An investigation of konjac glucomannan-keratin hydrogel scaffold loaded with Avena sativa extracts for diabetic wound healing.

We have developed a novel hydrogel composed of konjac glucomannan (KGM), human hair proteins (KER), and an ethanolic extract of Avena sativa (OAT) and evaluated its potential as a dressing material for diabetic wounds. KGM is an excellent biocompatible gelling agent that stimulates fibroblast proliferation and immunomodulation. Human hair proteins (KER) are biocompatible, biodegradable, and possess abundant cell adhesion sites. KER also promotes fibroblast attachment and proliferation, keratinocyte migration, and collagen expression, which can accelerate wound healing. OAT consists of oat ß-glucans and several anti-inflammatory and antioxidant moieties that can reduce prolonged inflammation in chronic wounds. SEM images confirm the highly porous architecture of the scaffolds. When immersed in PBS, KGM+KER+OAT hydrogels absorb 7.5 times their dry weight. These hydrogels display a measured rate of degradation in lysozyme. KGM+KER+OAT hydrogels showed no significant cytotoxicity against NIH/3T3 fibroblasts. DAPI and SEM images obtained after 48h of cell culture illustrate the attachment and infiltration of fibroblasts. In vivo studies performed using a diabetic rat excision wound model showed that KGM+KER+OAT hydrogels significantly accelerated wound healing compared to the control and the KGM+KER hydrogels.

Development of reduced graphene oxide (rGO)-isabgol nanocomposite dressings for enhanced vascularization and accelerated wound healing in normal and diabetic rats.

Treatment of chronic non-healing wounds in diabetes is still a major clinical challenge. Here, we have developed reduced graphene oxide (rGO) loaded isabgol nanocomposite scaffolds (Isab + rGO) to treat normal and diabetic wounds. rGO was synthesized by rapid reduction of graphene oxide (GO) under focused solar radiation. Then, rGO was uniformly dispersed into isabgol solution to prepare Isab + rGO nanocomposite scaffolds. These scaffolds were characterized using various physiochemical techniques. Isab + rGO nanocomposite scaffolds showed suitable cell viability, proliferation, and attachment. In vivo experiments were performed using Wistar rats to study the wound healing efficacy of these scaffolds in normal and diabetic rats. Results revealed that rGO stimulated collagen synthesis, collagen crosslinking, wound contraction, and reduced the wound re-epithelialization time significantly compared to control. Histology and immunohistochemistry analyses showed that Isab + rGO scaffold treatment enhanced angiogenesis, collagen synthesis, and deposition in treated wounds. Isab + rGO scaffold treatment also played a major role in shortening the inflammation phase and recruiting macrophages to enhance the early phase of wound healing. Overall, this investigation showed that Isab + rGO scaffold dressing could significantly accelerate the healing of normal and diabetic wounds.

Morin incorporated polysaccharide-protein (psyllium-keratin) hydrogel scaffolds accelerate diabetic wound healing in Wistar rats.

Chronic wounds cost several billion dollars of public healthcare spending annually and continue to be a persistent threat globally. Several treatment methods have been explored, and all of them involve covering up the wound with therapeutic dressings that reduce inflammation and accelerate the healing process. In this present study, morin (MOR) was loaded onto hydrogel scaffolds prepared from psyllium seed husk polysaccharide (PSH), and human hair keratins (KER) crosslinked with sodium trimetaphosphate. ATR-FTIR confirmed the presence of the constituent chemical ingredients. SEM images of the scaffold surface reveal a highly porous architecture, with about 80% porosity measured by liquid displacement measurement, irrespective of the morin concentration. Swelling assays carried out on the scaffolds portray an ability to absorb up to seven times their dry weight of fluids. This makes them attractive for guiding moist wound healing on medium exuding wounds. An Alamar blue assay of NIH/3T3 fibroblast cells shows that cell viability decreases in the first 24 h but recovers to 85% in comparison to a control after 48 h. SEM images of fibroblast cells grown on the scaffolds confirm cellular attachment. An in vivo diabetic wound healing study showed that PSH + KER + MOR scaffold treatment significantly reduced the re-epithelialization time (p < 0.01) and enhanced the rate of wound contraction (p < 0.001), by accelerating collagen synthesis in diabetic rats compared to controls.

Wound healing activity of a collagen-derived cryptic peptide.

Wound healing involves a well-controlled series of interactions among cells and several mediators leading to the restoration of damaged tissue. Degradation of the extracellular matrix (ECM) protein collagen during remodelling of wound tissue leads to the release of bioactive peptides that can possibly influence the healing process. The RGD-containing, antioxidative collagen peptide E1 isolated in an earlier work was screened in this study for its ability to influence multiple steps of the wound healing process. E1 was assayed for and found to be chemotactic. Excision and incision wounds were created on separate groups of rats and E1 was administered topically. The wound tissues were isolated on the 4th and 8th days post-wound and subjected to biochemical and biophysical analysis. A significant decrease in lipid peroxides in the treatment group confirmed the in vivo antioxidant capacity of E1. The treatment group also displayed significant increase in total protein, collagen and amino sugar synthesis indicating faster ECM formation. The significantly increased rate of wound contraction and reepithelialisation along with higher tensile strength of the wound tissue corroborated the results of biochemical analysis. The results confirm the significant role played by collagen peptides in accelerating the healing process and justify their possible use as a pharmaceutical agent.

Efficacy of Annona squamosa L in the synthesis of glycosaminoglycans and collagen during wound repair in streptozotocin induced diabetic rats.

The aim of this work was to find out the effects of Annona squamosa on the formation of glycosaminoglycans and collagen during wound healing in normal and diabetic rats. Diabetes induced rats were segregated into 4 groups, each containing six animals. Groups I and III served as the normal and diabetic control while groups II and IV served as normal and diabetic treated. The animals were treated with 200 µL of Annona squamosa extract topically. The granulation tissues formed were removed on the 8th day and the amount of glycosaminoglycans (GAGs) and collagen formed was evaluated by sequential extraction and SDSPAGE, respectively. Histological evaluation was also carried out using Masson's trichrome stain. In vitro wound healing efficacy of A. squamosa in human dermal fibroblast culture (HDF) was also carried out. The fibroblasts treated with varying concentrations of A. squamosa were examined for proliferation and closure of the wound area and photographed. A. squamosa increased cellular proliferation in HDF culture. The granulation tissues of treated wounds showed increased levels of glycosaminoglycans (P < 0.05) and collagen which were also confirmed by histopathology. The results strongly substantiate the beneficial effects of A. squamosa on the formation of glycosaminoglycans and collagen during wound healing.

Efficacy of Acorus calamus on collagen maturation on full thickness cutaneous wounds in rats.

BACKGROUND: The rhizomes of Acorus calamus and their essential oil are widely used in the flavoring industry and production of alcoholic beverages in Europe. Recent reports have confirmed the presence of several pharmacological components in the rhizomes of A. calamus. OBJECTIVE: The objective of this study was to find out the efficacy of topical administration of ethanolic extract of A. calamus on dermal wound healing in rats. Wound healing is a natural process occurring in living organisms, which results in a complete or partial remodeling of injured tissue and ultimately progresses to the formation of a fibrous scar. Several natural products have been reported to augment the wound healing process. MATERIALS AND METHODS: An ethanolic extract of A. calamus was prepared and its wound-healing efficacy was studied. An excision wound was made on the back of the rat and 200 µL (40 mg/kg body weight) of the A. calamus extract was applied topically once daily for the treated wounds. The control wounds were treated with 200 µL of phosphate buffered saline. RESULTS: The granulation tissues formed were removed at 4, 8 and 12 days and biochemical parameters such as deoxyribonucleic acid, total protein, total collagen, hexosamine and uronic acids were measured. The amount of type I/III collagen formed in control and treated wound tissues was evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The epithelialization time, tensile strength and histological examination of the wounds were also studied. Biochemical analyses of the granulation tissues revealed a significant increase in collagen, hexosamine and uronic acid when compared with the control. The tensile strength of extract treated wounds was found to increase by 112%. A significant reduction in lipid peroxide levels suggested that A. calamus possesses antioxidant components. CONCLUSIONS: The results strongly confirm the beneficial effects of A. calamus in augmenting the wound healing process.

Green synthesis of pullulan stabilized gold nanoparticles for cancer targeted drug delivery.

The aim of this study was to synthesize green chemistry based gold nanoparticles using liver specific biopolymer and to develop a liver cancer targeted drug delivery system with enhanced efficacy and minimal side effects. Pullulan stabilized gold nanoparticles (PAuNPs) were coupled with 5-Fluorouracil (5-Fu) and folic acid (Fa) which could be used as a tool for targeted drug delivery and imaging of cancer. The toxicity of 5-Fu, 5-Fu adsorbed gold nanoparticles (5-Fu@AuNPs), Fa-coupled 5-Fu adsorbed gold nanoparticles (5-Fu@AuNPs-Fa), was studied using zebrafish embryo as an in vivo model. The in vitro cytotoxicity of free 5-Fu, 5-Fu@AuNPs, 5-Fu@AuNPs-Fa against HepG2 cells was studied and found that the amount of 5-Fu required to achieve 50% of growth of inhibition (Ic50) was much lower in 5-Fu@AuNP-Fa than in free 5-Fu, 5-Fu@AuNPs. The in vivo biodistribution of PAuNPs showed that higher amount of gold had been accumulated in liver (54.42±5.96 µg) than in other organs.

An eco-friendly method for short term preservation of skins/hides using Semecarpus anacardium nut extract.

Preservation or curing of hides and skins is performed as the primary step of leather processing. Common salt is employed as the conventional agent for curing purpose. Use of salt enhances the pollution load of tannery effluent which becomes highly contaminated with increased total dissolved solids and chlorides. To overcome this hurdle, researchers are in constant search of alternative preservation techniques which are either totally void of salt or use only a meager amount of salt. In the present study, we had explored the possibility of using Semecarpus anacardium nut extract as an alternative to salt for the curing process by assessing different parameters like hair slip, putrefaction odor, volatile nitrogen content, moisture content, bacterial count, and shrinkage temperature in comparison to the salt curing method. The antibacterial property of the plant extract was also investigated. The results obtained substantiated that the nut extract of S. anacardium effectively could preserve the skins for more than a month, by its antibacterial activity along with the dehydrating property of acetone.

Efficacy of L-proline administration on the early responses during cutaneous wound healing in rats.

Proline (Pro) plays a versatile role in cell metabolism and physiology. Pro and hydroxypro are major imino acids present in collagen, an important connective tissue protein, essential for wound healing, which is a primary response to tissue injury. This study explains the role of L-pro on cutaneous wound healing in rats when administered both topically and orally. Open excision wounds were made on the back of rats, and 200 µl (200 mg) of pro was administered topically and orally once daily to the experimental rats until the wounds healed completely. The control wounds were left untreated. Granulation tissues formed were removed after day 4 and 8 of post excision wounding, and biochemical parameters such as total protein, collagen, hexosamine, and uronic acid were estimated. Levels of enzymatic and non-enzymatic antioxidants such as catalase, superoxide dismutase, glutathione peroxidase, ascorbic acid, and reduced glutathione were evaluated along with lipid peroxides in the granulation tissues. Tensile strength and period of epithelialization were also measured. It was observed that the treated wounds healed very fast as evidenced by augmented rates of epithelialization and wound contraction, which was also confirmed by histological examinations. The results strappingly authenticate the beneficial effects of the topical administration of L-proline in the acceleration of wound healing than the oral administration and control.

Spontaneous ultra fast synthesis of gold nanoparticles using Punica granatum for cancer targeted drug delivery.

Rapid synthesis of mono-dispersed gold nanoparticles through economically feasible green chemistry approach is highly desirable. In this study, we have developed a method to synthesize mono-dispersed gold nanoparticles (PAuNPs) by mixing gold solution with fruit peel extract of Punica granutum without using any surfactant or external energy. In this method, physiologically stable, biocompatible PAuNPs were formed within 60s. Casein, being a biocompatible polymer, is used to couple the prepared PAuNPs for functionalization of folic acid, which is highly expressed in cancer cells. These functionalized PAuNPs could be used for targeted drug delivery for cancer with enhanced therapeutic efficacy and minimal side effects. PAuNPs were characterized by UV, IR, TEM, Particle size analyzer and zeta potential measurement. In vitro stability of the PAuNPs was also analyzed. Hemocompatibility of PAuNPs was evaluated in human blood samples and found that the particles were hemocompatible. The toxicity of the PAuNPs, 5-Fu and 5Fu@PAuNPs was analyzed in zebrafish embryos. The in vitro cytotoxicity of free 5-Fu, 5Fu@PAuNPs-Fa was investigated against MCF-7 cells (breast cancer) and observed that the amount of 5-Fu required to achieve 50% of growth of inhibition (Ic50) was much lower when compared to free 5-Fu.

Preparation of amphiphilic hollow carbon nanosphere loaded insulin for oral delivery.

Hollow amphiphilic carbon nanosphere loaded insulin with biodegradable polymer coating (ACP) for oral delivery, was developed to overcome intestinal epithelial barriers and protect insulin from photolytic enzymes. ACP was characterized by laser diffraction spectroscopy for size, distribution and shape by using transmission electron microscopy. Drug-carrier compatibility was studied individually by FTIR and CD spectroscopy. Insulin encapsulation efficiency and in vitro release were determined by Bradford protein assay. Anti-diabetic activity of ACP loaded insulin was determined using Wistar diabetic rats. The mean size of ACP was ∼330 nm with poly index value of 1. Insulin encapsulation efficiency was 93.35%. We observed that ACP formulation suppressed insulin release in acidic media and promoted a sustained release at near neutral conditions. Hence, ACP insulin could be an alternative for injections to diabetic patients.

Topical application of Acalypha indica accelerates rat cutaneous wound healing by up-regulating the expression of Type I and III collagen.

ETHNOPHARMACOLOGICAL RELEVANCE: Acalypha indica Linn. (Acalypha indica) vernacularly called Kuppaimeni in Tamil, has been used as a folklore medicine since ages for the treatment of wounds by tribal people of Tamil Nadu, Southern India. The present study investigates the biochemical and molecular rationale behind the healing potential of Acalypha indica on dermal wounds in rats. MATERIAL AND METHODS: Acalypha indica extract (40 mg/kg body weight) was applied topically once a day on full-thickness excision wounds created on rats. The wound tissue was removed and used for estimation of various biochemical and biophysical analyses and to observe histopathological changes with and with-out extract treatment. The serum levels of pro-inflammatory cytokine tumor necrosis factor (TNF-α) was measured at 12 h, 24 h, 48 h and 72 h post-wounding using ELISA. Reverse transcription-polymerase chain reaction (RT-PCR) analysis was performed to study the expression pattern of transforming growth factor [TGF-ß1], collagen 1 α (I) [Col 1 α (I)] and collagen 3 α (I) [Col 3 α (I)]. Likewise, linear incision wounds were created and treated with the extract and used for tensile strength measurements. RESULTS: Wound healing in control rats was characterized by less inflammatory cell infiltration, lack of granulation tissue formation, deficit of collagen and significant decrease in biomechanical strength of wounds. Acalypha indica treatment mitigated the oxidative stress and decreased lipid peroxidation with concomitant increase in ascorbic acid levels. It also improved cellular proliferation, increased TNF-α levels during early stages of wound healing, up-regulated TGF-ß1 and elevated collagen synthesis by markedly increasing the expression of Col 1 α (I) and Col 3 α (I). Increased rates of wound contraction, epithelialization, enhanced shrinkage temperature and high tensile strength were observed in the extract treated rats. CONCLUSION: Acalypha indica extract was shown to augment the process of dermal wound healing by its ability to increase collagen synthesis through up-regulation of key players in different phases of wound healing and by its antioxidative potential.

Efficacy of Annona squamosa on wound healing in streptozotocin-induced diabetic rats.

Annona squamosa L. (Annonaceae), commonly known as custard apple, mainly used for its edible fruit, is also recognised with numerous medicinal properties. As there is no report on the efficacy of this plant for wound healing, we examined the efficacy of ethanolic extract of A. squamosa leaves on wound repair in streptozotocin-nicotinamide-induced diabetic rats. Open excision wounds were made on the back of rats. The drug at a dosage of 100 mg/kg body wt was reconstituted in 200 µl of phosphate buffered saline and applied topically once daily for the treated wounds. The control wounds were left untreated. Wound tissues formed on days 4, 8, 12 and 16 (post-wound) were used to estimate DNA, total protein, total collagen, hexosamine and uronic acid. Levels of lipid peroxides were also evaluated along with tensile strength and period of epithelialisation. A. squamosa L. increased cellular proliferation and collagen synthesis at the wound site as evidenced by increase in DNA, protein and total collagen. The treated wounds were observed to heal much faster as proved by enhanced rates of epithelialisation and wound contraction, which was also confirmed by histopathological examinations. The results strongly substantiate the beneficial effects of the topical application of A. squamosa L. in the acceleration of normal and diabetic wound healing.

Phyllanthin of Standardized Phyllanthus amarus Extract Attenuates Liver Oxidative Stress in Mice and Exerts Cytoprotective Activity on Human Hepatoma Cell Line.

BACKGROUND: Phyllanthus amarus, a traditional herbal liver-protecting medicine, is known to contain an active ingredient phyllanthin. Many research studies and clinical trials performed in the past using this plant have given contentious results which clearly accentuates the need for the standardization of the extracts. AIM: In this study, P. amarus extract was standardized for phyllanthin content by high performance thin layer chromatography (HPTLC) and high performance liquid chromatography (HPLC) analysis. The preventive role of a standardized extract of P. amarus against CC14-induced hepatotoxicity in vivo and in vitro using mice model and human hepatoma HepG2 cell line, respectively, was investigated. METHODS: Phyllanthin was used as a marker phytochemical for the standardization of P. amarus extract. The extracts were verified for phyllanthin content by HPTLC and HPLC. Female mice were orally administered with CCl4 either with or without standardized P. amarus extract in three different doses. Similarly, the cytoprotective role of the standardized extract in vitro was studied in HepG2 cell line. RESULTS: Oral administration of CCl4 resulted in increased oxidative stress, decreased antioxidative defense, and liver injury. Treatment with P. amarus along with CCl4 significantly mitigated the increase in activities of liver marker enzymes, lipid peroxidation, and bilirubin content. It also increased the antioxidant enzymatic and non-enzymatic defense parameter levels. The results of the in vitro study conducted in HepG2 cells indicated that the hepatotoxin lowered 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (Mil) metabolism and increased the release of transaminases which were corrected with co-incubation with P. amarus. CONCLUSION: The study established a significant liver-protecting role of standardized P. amarus extract due to the presence of active ingredient phyllanthin.

Emblica officinalis exerts wound healing action through up-regulation of collagen and extracellular signal-regulated kinases (ERK1/2).

During wound healing, the wound site is rich in oxidants, such as hydrogen peroxide, mostly contributed by neutrophils and macrophages. Ascorbic acid and tannins of low molecular weight, namely emblicanin A (2,3-di-O-galloyl-4,6-(S)-hexahydroxydiphenoyl-2-keto-glucono-delta-lactone) and emblicanin B (2,3,4,6-bis-(S)-hexahydroxydiphenoyl-2-keto-glucono-delta-lactone) present in Emblica officinalis (emblica), have been shown to exhibit a very strong antioxidant action. We proposed that addition of these antioxidants to the wound microenvironment would support the repair process. The present investigation was undertaken to determine the efficacy of emblica on dermal wound healing in vivo. Full-thickness excision wounds were made on the back of the rat and topical application of emblica accelerated wound contraction and closure. Emblica increased cellular proliferation and cross-linking of collagen at the wound site, as evidenced by an increase in the activity of extracellular signal-regulated kinase 1/2, along with an increase in DNA, type III collagen, acid-soluble collagen, aldehyde content, shrinkage temperature and tensile strength. Higher levels of tissue ascorbic acid, alpha-tocopherol, reduced glutathione, superoxide dismutase, catalase, and glutathione peroxidase support the fact that emblica application promotes antioxidant activity at the wound site. In summary, this study provides firm evidence to support that topical application of emblica represents a feasible and productive approach to support dermal wound healing.

Curcumin improves wound healing by modulating collagen and decreasing reactive oxygen species.

Wound healing consists of an orderly progression of events that re-establish the integrity of the damaged tissue. Several natural products have been shown to accelerate the healing process. The present investigation was undertaken to determine the role of curcumin on changes in collagen characteristics and antioxidant property during cutaneous wound healing in rats. Full-thickness excision wounds were made on the back of rat and curcumin was administered topically. The wound tissues removed on 4th, 8th and 12th day (post-wound) were used to analyse biochemical and pathological changes. Curcumin increased cellular proliferation and collagen synthesis at the wound site, as evidenced by increase in DNA, total protein and type III collagen content of wound tissues. Curcumin treated wounds were found to heal much faster as indicated by improved rates of epithelialisation, wound contraction and increased tensile strength which were also confirmed by histopathological examinations. Curcumin treatment was shown to decrease the levels of lipid peroxides (LPs), while the levels of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), activities were significantly increased exhibiting the antioxidant properties of curcumin in accelerating wound healing. Better maturation and cross linking of collagen were observed in the curcumin treated rats, by increased stability of acid-soluble collagen, aldehyde content, shrinkage temperature and tensile strength. The results clearly substantiate the beneficial effects of the topical application of curcumin in the acceleration of wound healing and its antioxidant effect.

The effect of nerve growth factor on the early responses during the process of wound healing.

In this study, we investigated the role of nerve growth factor (NGF)-incorporated collagen on wound healing in rats. Full-thickness excision wounds were made on the back of female rats weighing about 150-160 g. Topical application of NGF-incorporated collagen, at a concentration of 1 microg/1.2 mg collagen/cm(2), once a day, for 10 days resulted in complete healing of wounds on the 15th day. The concentrations of collagen, hexosamine and uronic acid in the granulation tissue were determined. The NGF-incorporated collagen-treated rats required shorter duration for the healing with an increased rate of wound contraction. Histological and electron microscopical evaluations were also performed, which reveal the activation of fibroblasts and endoplasmic reticulum and therefore increased level of collagen synthesis due to NGF application. These results clearly indicate that the topical application of NGF-incorporated collagen enhanced the rate of healing of excision wounds.

Modulatory effect of Coccinia indica on aortic collagen in streptozotocin-induced diabetic rats.

1. The effects of Coccinia indica, an indigenous plant used in Ayurvedic medicine in India, on aortic collagen content and its characteristics were assessed in streptozotocin (STZ) diabetic rats. 2. Rats were made diabetic with a single intraperitoneal injection of STZ (45 mg/kg). Blood glucose, hydroxyproline, collagen, extent of glycation, collagen-linked fluorescence, soluble pattern of pepsin-soluble collagen, shrinkage temperature, alpha/beta ratio of type I collagen and type I/type III collagen ratio were determined in rats treated with C. indica leaf extract (CLEt; 200 mg/kg for 45 days using an oral intragastric tube). 3. In diabetic rats, the collagen content, as well as the degree of cross-linking, was increased, as evidenced by increased shrinkage temperature and decreased pepsin solubility. The alpha/beta ratio of type I collagen and the type I/type III collagen ratio of pepsin-soluble collagen were significantly decreased in STZ diabetic rats. 4. In conclusion, administration of CLEt for 45 days to STZ diabetic rats significantly reduced the accumulation and cross-linking of collagen. The effects of C. indica (collagen content 23.87 +/- 1.52 mg/100 mg tissue (t value = 6.80), extent of cross-linking 0.893 +/- 0.072 mg hydroxyproline/100 mg tissue (t value = 9.0)) were comparable with those of glibenclamide (collagen content 26.18 +/- 1.65 mg/100 mg tissue (t value = 4.58), extent of cross-linking 0.787 +/- 0.057 mg hydroxyproline/100 mg tissue (t value = 7.1)), a reference drug.

Similarity between the major collagens of cuttlefish cranial cartilage and cornea.

Invertebrates possess unique collagen-containing connective tissue elements, the biochemistry of which is not clearly understood. We previously reported the occurrence of a novel heterotrimeric type V/XI like collagen in the cranial cartilage of the cuttlefish Sepia officinalis. We report here the purification of the three chains by ion exchange chromatography and the physicochemical characteristics of this collagen. This collagen shared substantial similarity to the collagen purified from the cornea of S. officinalis, with respect to chain composition, cyanogen bromide peptide profile and amino acid composition. The mobility of the C3 chain was retarded in the corneal collagen, which also had an increased glycine content and a smaller ratio of hydroxylysine to lysine, together with a reduction in bound carbohydrates. The cartilage collagen had a higher denaturation temperature than corneal collagen. As observed by transmission electron microscopy of reconstituted fibrils, the heterotrimeric invertebrate collagen formed fibrils of no apparent periodicities as opposed to the regular 64-nm banding pattern of milk shark (Rhizoprionodon acutus) cartilage collagen. This is also the first report on the molecular species of collagen in an invertebrate cornea. Our results strongly support the functioning of minor vertebrate collagens as major collagens in some invertebrates, close similarity of collagens in two tissues with different functions and would hold significance to our understanding of collagen polymorphism and the evolution of the extracellular matrix.