Denudation of human amniotic membrane by a novel process and its characterisations for biomedical applications R. Sripriya, R. Kumar Progress in Biomaterials, 2016 This study was aimed to investigate the suitability of a modified method to get decellularised human amniotic membrane (DHAM). The obtained membrane was subjected to physico-chemical and biological evaluations to validate its potential for biomedical applications. The human amniotic membrane was processed with detergent and alkali followed by enzymatic treatments. Hematoxylin and eosin (H&E) and Masson’s trichrome staining of membrane were in accordance with conjectures: the decellularised membrane stained for extracellular matrix is rich in collagen. Scanning electron micrograph also showed the denudation in the processed membrane with the cellular impressions on the basement membrane. Physical characteristics namely the differential scanning calorimetric, tensile, shrinkage behaviour and the Fourier transform infrared spectra of decellularised membrane showed its stability and intact structure similar to the unprocessed membrane. In the visible range of light, the membrane was found to be transparent from 90 to 98 %. Proliferation rate of fibroblasts, keratinocytes, myoblasts and hepatocytes were significantly upregulated compared to the control. The cell morphologies were normal and differentiation of myoblasts into myotubes were more pronounced in decellularised membrane. Proliferation of corneal limbal cells on decellularised membrane showed 92–100 % confluency on day 21 and the migrated cells displayed a spindle shape and changing later to a more cuboidal appearance.
Preparation and comparative characterization of keratin-chitosan and keratin-gelatin composite scaffolds for tissue engineering applications S. Balaji, Ramadhar Kumar, R. Sripriya, Prachi Kakkar, D. Vijaya Ramesh, P. Neela Kanta Reddy, P.K. Sehgal Materials Science and Engineering C, 2012 We report fabrication of three dimensional scaffolds with well interconnected matrix of high porosity using keratin, chitosan and gelatin for tissue engineering and other biomedical applications. Scaffolds were fabricated using porous Keratin–Gelatin (KG), Keratin–Chitosan (KC) composites. The morphology of both KG and KC was investigated using SEM. The scaffolds showed high porosity with interconnected pores in the range of 20–100 μm. They were further tested by FTIR, DSC, CD, tensile strength measurement, water uptake and swelling behavior. In vitro cell adhesion and cell proliferation tests were carried out to study the biocompatibility behavior and their application as an artificial skin substitute. Both KG and KC composite scaffolds showed similar properties and patterns for cell proliferation. Due to rapid degradation of gelatin in KG, we found that it has limited application as compared to KC scaffold. We conclude that KC scaffold owing to its slow degradation and antibacterial properties would be a better substrate for tissue engineering and other biomedical application.
Physical characterization of succinylated type I collagen by Raman spectra and MALDI-TOF/MS and in vitro evaluation for biomedical applications Ramadhar Kumar, R. Sripriya, S. Balaji, M. Senthil Kumar, P.K. Sehgal Journal of Molecular Structure, 2011 In this study, we report on physical and in vitro biological characterization of succinylated collagen (SC). SC was prepared by succinylation of type I bovine tendon collagen. SC swells and dissolves in physiological pH buffers (pH 7.4) Biocompatibility of SC to collagen for fibroblasts was comparable but L6 myoblasts showed pronounced proliferation and differentiation with SC. Using the MALDI-TOF/MS technique, SC was found with increased molecular mass by 16,359 Da per molecule which corresponds to about 54 succinyl groups covalently linked to the collagen strand. Raman spectroscopy revealed the retention of triple helical structure conformation in the presence of linked succinyl groups. New peaks near 1737, 1675 and 1420 cm −1 and decreased intensities near 2440 and 488 cm −1 provides the most convenient marker bands for succinylation of collagen. The intense band regions near 2856–2934, 2724, and 1445 cm −1 also confirms the existence of succinyl groups.
Characterizations of polyanionic collagen prepared by linking additional carboxylic groups R. Sripriya, Ramadhar Kumar, S. Balaji, M. Senthil Kumar, P.K. Sehgal Reactive and Functional Polymers, 2011 We have successfully attempted to characterize the polyanionic collagen (PAC) intended for biomaterial and tissue engineering applications. PAC was obtained from type I collagen by chemically linking additional carboxylic groups to the reactive free amino groups. These changes do not affect the α chains morphology and conformation in collagen. Pure type I collagen obtained from bovine Achilles tendons was allowed to react slowly at pH 9 with succinic anhydride to attain 96% conversion into PAC confirmed by trinitrobenzenesulfonic acid (TNBS) assay. Electrophoretic mobility pattern of α, β and γ components of PAC shifted to higher molecular weight by 16,300 Da indicating modification of collagen. Similarly, FTIR, CD and DSC indicated retention of triple helical conformation of PAC. Novelty of PAC lies in its swelling behavior resulting in dissolution thus offering transparency in neutral pH buffer. The results obtained here give us an opportunity to use PAC as an injectable device for tissue engineering applications as its up-regulatory efficiency for cytoproliferation validates its biocompatibile potentials.
In vitro evaluation of antioxidants of fruit extract of Momordica charantia L. on fibroblasts and keratinocytes Ramadhar Kumar, S. Balaji, R. Sripriya, N. Nithya, T. S. Uma, P. K. Sehgal Journal of Agricultural and Food Chemistry, 2010 The antioxidant activity of the total aqueous extract (TAE) and total phenolic extract (TPE) of Momordica charantia fruits was assayed by radical-scavenging methods and cytoprotective effects on hydrogen peroxide (H(2)O(2))- and hypoxanthin-xanthin oxidase (HX-XO)-induced damage to rat cardiac fibroblasts (RCFs), NIH 3T3, and keratinocyte (A431). Cell viability was monitored by a 3-[4,5-dimethyltriazol-2-yl]-2,5-diphenyltretrazolium (MTT) assay. For fibroblasts, TPE at 200 and 300 microg/mL showed maximum and consistent cytoprotection against oxidants. The extract at 50 microg/mL also had significant and slightly protective effects on fibroblasts against H(2)O(2)- and HX-XO-induced damage, respectively. RCF was more tolerant toward the damage. For keratinocytes, a dose-dependent relationship of oxidant toxicity was only seen with H(2)O(2) but the protective action of the extract correlated with oxidant dosage. At 200 and 300 microg/mL TPE, cytoprotection was dose-dependent against oxidants. Extracts had no effect on HX-XO toxicity at 50 microg/mL. Pretreatment with both the extracts did not show any cytoprotection.
Triphala Incorporated Collagen Sponge-A Smart Biomaterial for Infected Dermal Wound Healing Muthusamy Senthil Kumar, Shanmugam Kirubanandan, Ramasamy Sripriya, Praveen Kumar Sehgal Journal of Surgical Research, 2010 BACKGROUND Wound infection is a major problem in the medical community since many types of wounds are more prone to microbial contamination leading to infection. Triphala (a traditional ayurvedic herbal formulation) incorporated collagen sponge was investigated for its healing potential on infected dermal wound in albino rats. MATERIALS AND METHODS Methanol extract of triphala was prepared and analyzed for the presence of catechin by high-pressure liquid chromatography analysis. Collagen sponge was prepared by incorporating triphala into collagen sponge. The triphala incorporated collagen was characterized by Fourier transform infrared spectroscopy, differential scanning calorimetry, and water uptake analysis. Infected wound was dressed with triphala incorporated collagen sponge. Wound reduction rate, collagen content, and matrix metalloproteinases in the granulation tissue, histology, and Fourier transform electron microscopy analysis were done to obtain the healing pattern. RESULTS High-pressure liquid chromatography analysis showed the presence of (-)epigallocatechin gallate. FT-IR spectroscopy study revealed the interaction of polyphenols with the collagen. Triphala incorporated collagen sponge has shown to increase thermal stability and water uptake capability, faster wound closure, improved tissue regeneration, collagen content at the wound site, and supporting histopathological parameters pertaining to wound healing. Matrix metalloproteinases expression was correlated well with reduction in the inflammatory phase, thus confirming efficacy of the dressing. CONCLUSIONS Better healing efficacy of triphala incorporated collagen sponge may provides a scientific rationale for the use of this dressing as an effective wound cover in the management of infected dermal wound.
Drug delivery dressings P.K. Sehgal, R. Sripriya, M. Senthilkumar Advanced Textiles for Wound Care A Volume in Woodhead Publishing Series in Textiles, 2009 A description of wound types is presented and the wounds that need drug delivery dressings are identified. Both acute and chronic wounds require drugs for controlling infection during healing. Healing of such wounds needs proper management mediated through the concept of delivering drug to the wound site. The properties of drug delivery dressings are described. Some dressings control the transport of biological fluids, whereas others offer haemostatic properties leading to control of infection and improved healing. With the advent of tissue engineering technology and advances in micro and nanotechnology, gene delivery and stem cell research, it is possible to design and construct an environment-sensitive dressing tailor-made to the specific wound type leading to its complete healing without any adverse reactions.
Triphala Promotes Healing of Infected Full-Thickness Dermal Wound Muthusamy Senthil Kumar, Shanmugam Kirubanandan, Ramasamy Sripriya, Praveen Kumar Sehgal Journal of Surgical Research, 2008 BACKGROUND Infection is a major problem in the management of wounds. Even though the development of synthetic antimicrobial agents persists, drug resistance and toxicity hinder their way. Many plants with multi-potent pharmaceutical activities may offer better treatment options, and Triphala (dried fruits of Terminalia chebula, Terminalia bellirica, and Phyllanthus emblica) are potential formulations evaluated for healing activity on infected wound as it possesses numerous activities. MATERIALS AND METHODS Alcoholic extract of Triphala has shown in vitro antimicrobial activity against wound pathogens such as Staphylococcus aureus, Pseudomonas aeruginosa, and Streptococcus pyogenes. An ointment was prepared from the Triphala extract (10% w/w) and assessed for in vivo wound healing on infected rat model by rate of healing, bacterial count, biochemical analysis, and expression of matrix metalloproteinases. RESULTS The treated group has shown significantly improved wound closure. Assessment of granulation tissue on every fourth day showed significant reduction in bacterial count with significant level of collagen, hexosamine, uronic acid, and superoxide dismutase in the treated group (P < 0.01). Reduction of matrix metalloproteinase expression observed in the treated group by gelatin zymography and immunoblotting confirms our in vivo assessment. CONCLUSIONS The above results showed the antibacterial, wound healing, and antioxidant activities of Triphala ointment, necessary for the management of infected wounds. Active principles of the Triphala may be further evaluated and used as an excellent therapeutic formulation for infected wounds.
Modified type I collagen - A barrier membrane in human gingival recession management Trends in Biomaterials and Artificial Organs, 2008
