Praveen Kumar Sehgal

Formulation and evaluation of quercetin polycaprolactone microspheres for the treatment of rheumatoid arthritis

Quercetin had been shown to be effective in the management of arthritis. However, bioavailability of quercetin is a concern for such treatment. This work aims at the development of intra-articular drug delivery system by controlled release of quercetin (loaded in microspheres) for the management of rheumatoid arthritis. Polycaprolactone has been used for the preparation of microspheres (with quercetin) using the solvent evaporation method. The physio-chemical characterisation of polycaprolactone-loaded quercetin microspheres was carried out to obtain information about particle size distribution, drug loading efficiency, morphology, thermal properties, polymorphism and release trends in phosphate-buffered saline at pH 7.4 and 37°C. Quercetin-loaded polycaprolactone microspheres were found to be biocompatible as evidenced from in vitro and in vivo studies using a rabbit synovial cells and Wistar rats, respectively. Quercetin release from microspheres of selected formulations showed biphasic nature due to initial burst effect followed by a controlled release. These results suggest that optimised quercetin-loaded polycaprolactone microspheres may be the viable strategy for controlled release of quercetin in the joint cavity for more than 30 days by intra-articular injection to treat rheumatoid arthritis.

Preparation and properties of tannic acid cross-linked collagen scaffold and its application in wound healing.

A biodurable porous scaffold of collagen with good biocompatibility and enhanced wound healing potential is prepared through casting technique using tannic acid (TA) as crosslinker. The morphological analysis of the tannic acid cross-linked collagen scaffold (TCCs) distinctively shows scaly interlinks with large pores. The enzymatic stability of the scaffold is characterized in vitro to detail the role of TA in stabilization of collagen matrix against collagenolytic degradation. TCCs shows more stability (>54%) against collagenase than that of the collagen scaffolds (Cs). The attenuated total reflectance Fourier transform infrared analysis of the TCCs confirms the noncovalent interaction between collagen and TA. The biocompatibility of the scaffold (TCCs) in vitro has been established using 3T3 fibroblasts. Therapeutic and wound healing potential of the TCCs has been studied in vivo using excision wound model in rats. The results clearly indicates that the TCCs has greater and significant effect in wound closure and increased the wound healing rate compared with native Cs. This biocompatible and biodurable scaffold may find broad applications in the tissue engineering and drug delivery applications.

Fruit extracts of Momordica charantia potentiate glucose uptake and up-regulate Glut-4, PPARγ and PI3K

ETHNOPHARMACOLOGICAL RELEVANCE Momordica charantia fruit is a widely used traditional medicinal herb as, anti-diabetic, anti-HIV, anti-ulcer, anti-inflammatory, anti-leukemic, anti-microbial, and anti-tumor. AIMS OF STUDY The present study is undertaken to investigate the possible mode of action of fruit extracts derived from Momordica charantia (MC) and study its pharmacological effects for controlling diabetic mellitus. Effects of aqueous and chloroform extracts of Momordica charantia fruit on glucose uptake and up-regulation of glucose transporter (Glut-4), peroxisome proliferator activator receptor gamma (PPAR gamma) and phosphatidylinositol-3 kinase (PI3K), were investigated to show its efficacy as a hypoglycaemic agent. MATERIALS AND METHODS Dose dependent glucose uptake assay was performed on L6 myotubes using 2-deoxy-D-[1-(3)H] glucose. Up-regulatory effects of the extracts on the mRNA expression level of Glut-4, PPAR gamma and PI3K have been studied. RESULTS The association of Momordica charantia with the aqueous and chloroform extracts of Momordica charantia fruit at 6 microg/ml has shown significant up-regulatory effect, respectively, by 3.6-, 2.8- and 3.8-fold on the battery of targets Glut-4, PPAR gamma and PI3K involved in glucose transport. The up-regulation of glucose uptake was comparable with insulin and rosiglitazone which was approximately 2-fold over the control. Moreover, the inhibitory effect of the cyclohexamide on Momordica charantia fruit extract mediated glucose uptake suggested the requirement of new protein synthesis for the enhanced glucose uptake. CONCLUSION This study demonstrated the significance of Glut-4, PPAR gamma and PI3K up-regulation by Momordica charantia in augmenting the glucose uptake and homeostasis.

Green chemistry approaches to leather tanning process for making chrome-free leather by unnatural amino acids

In the present study, green and sustainable method or eco-friendly approaches to tanning process based on unnatural D-amino acids (D-AA)-aldehyde (Ald) as a substitute for chrome-free tanning has been attempted. The distribution of optically active D-AA in tanned leather, the hydrothermal stability, the mechanical properties and resistance to collagenolytic activity of tanned leather, the evaluation of eco-friendly characteristics were investigated. Scanning electron microscopic (SEM) and Atomic force microscopic (AFM) analyses indicate the surface morphology and roughness, respectively, of the tanned leather collagen matrix. Shrinkage and Differential scanning calorimetric (DSC) analyses shows that the shrinkage temperature (T(s)) and denaturation temperature (T(d)) of tanned leather are related to the content of D-AA+Ald present in the leather matrix. It has been found that the T(s) of D-AA tanned leather is more than that of Ald tanned leather and also more or less equal to chrome tanned leather. Environmental impact assessment (EIA) shows that the developed process results in significant reduction in total solids content (TSC) and improves better biodegradability of organic compound present in the effluent compared to chrome tanning.

Synthesis and Characterization of Hybrid Biodegradable Films From Bovine Hide Collagen and Cellulose Derivatives for Biomedical Applications

Biomaterials are used as wound dressings and scaffolds, which are generally required to disperse by resorption into the body after tissue generation. Most methods employ relatively toxic cross-linking agent. In this study, an attempt has been made to prepare hybrid biodegradable films from bovine collagen by combining with cellulose derivatives without employing a cross-linking agent. The bovine hide trimmings were converted into soluble collagen and blended with 25, 50, 75, and 100% (wt/wt) 2-hydroxyethyl cellulose. These blends were cast into thin films and were analyzed for structure, thermal, mechanical, and biostability properties. It was found that the dry hybrid films have improved tensile strength in the range of 22 ± 4.6 to 58.9 ± 4.3 MPa, compared to the pure collagen films. On the other hand, hydrated films exhibited reduced strength and improved elongation compared to dry hybrid films. The structure of select films was investigated using scanning electron microscope (SEM) both in wet and dry state and found that the pore distribution is more in the wet state with high cellulose concentration. Differential scanning calorimetric (DSC) analysis showed that the thermal stability of dry hybrid films was improved moderately while the hydrated hybrid films exhibited enhanced hydrothermal stability compared to pure collagen films. The infrared (IR) spectral analysis of hybrid films reveals the presence of functional groups from both collagen and cellulose, especially the signature peaks of collagen triple helical structure. The equilibrium swelling and in-vitro biodegradability analysis proved that the presence of cellulose derivative could noticeably improve the biostability of the hybrid films leading to controlled degradation compared to the pure collagen films. Cell interaction studies show good biocompatibility and increased cell growth on the hybrid films as a function of cellulose concentration, although collagen is sourced from bovine hide trimmings of leather industry.

Protective Effect of Withania somnifera and Cardiospermum halicacabum Extracts Against Collagenolytic Degradation of Collagen

The irreversible destruction of extracellular matrix (ECM) such as cartilage, tendon, and bone that comprise synovial joints is the hallmark of both rheumatoid arthritis and osteoarthritis by over-expression of matrix metalloproteinase (MMP)-collagenases. We report herein the detailed study on the inhibitory effects of Withania somnifera extract (WSE) and Cardiospermum halicacabum extract (CHE) on Clostridium histolyticum collagenase (ChC) activity against the degradation of the ECM component of bovine Achilles tendon type I collagen by hydroxyproline assay method. Interaction of WSE and CHE with ChC exhibited 71% and 88% inhibition, respectively, to the collagenolytic activity of ChC against collagen degradation, and the inhibition was found to be concentration-dependent. The inhibition kinetics of ChC by both the extracts has been deduced from the extent of hydrolysis of N-[3-(2-furyl) acryloyl]-Leu-Gly-Pro-Ala. Both WSE and CHE are provided competitive and mixed type inhibition on ChC activity, respectively. Circular dichroism studies of ChC on treatment with WSE and CHE revealed changes in the secondary structure of collagenase. These results suggest that the WSE and CHE facilitated collagen stabilization through collagenase inhibition.

In Vitro Evaluation of Antioxidants of Fruit Extract of Momordica charantia L. on Fibroblasts and Keratinocytes

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.

Novel collagen scaffolds prepared by using unnatural D-amino acids assisted EDC/NHS crosslinking

This work discusses the preparation and characterization of novel collagen scaffolds by using unnatural D-amino acids (Coll-D-AAs)-assisted 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC)/N-hydroxyl succinimide(NHS)-initiated crosslinking. The mechanical strength, hydrothermal and structural stability, resistance to biodegradation and the biocompatibility of Coll-D-AAs matrices were investigated. The results from Thermo mechanical analysis, Differential scanning calorimetric analysis and Thermo gravimetric analysis of the Coll-D-AAs matrices indicate a significant increase in the tensile strength (TS, 180 ± 3), % elongation (% E, 80 ± 9), elastic modulus (E, 170 ± 4) denaturation temperature (T d, 108 ± 4) and a significant decrease in decomposition rate (T g, 64 ± 6). Scanning electron microscopic and Atomic force microscopic analyses revealed a well-ordered with properly oriented and well-aligned structure of the Coll-D-AAs matrices. FT-IR results suggest that the incorporation of D-AAs favours the molecular stability of collagen matrix. The D-AAs stabilizing the collagen matrices against degradation by collagenase would have been brought about by protecting the active sites in collagen. The Coll-D-AAs matrices have good biocompatibility when compared with native collagen matrix. Molecular docking studies also indicate better understanding of bonding pattern of collagen with D-AAs. These Coll-D-AAs matrices have been produced in high mechanical strength, thermally and biologically stable, and highly biocompatible forms that can be further manipulated into the functional matrix suitable in designing scaffolds for tissue engineering and regenerative medical applications.

Development of D-lysine-assisted 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide/N-hydroxysuccinimide-initiated cross linking of collagen matrix for design of scaffold†

This work discusses the preparation and characterization of collagen scaffold with presence of D-Lysine (Coll-D-Lys)-assisted 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC)/N-hydroxysuccinimide (NHS)-initiated cross linking. The mechanical strength, thermal and structural stability, resistance to biodegradation and cell viability of this scaffold was investigated. The results of the Coll-D-Lys-EDC/NHS scaffold also indicate an increase in the tensile strength (T(S)), percentage of elongation (% E), denaturation temperature (T(d)), and decrease the decomposition rate. Scanning electron microscopic (SEM) and atomic force microscopic (AFM) analyses revealed a well ordered with properly oriented and well-aligned structure of scaffold. The D-Lys stabilizes the scaffold against degradation by collagenase than L-Lys. The cell assay showed more than 98 ± 2% fibroblast viability (NIH 3T3) after 72 h of culture Coll-D-Lys-scaffold when compared with native Coll and Coll-L-Lys-scaffold. The proteolytic machinery is not well equipped to deal with Coll-D-Lys-scaffold than Coll-L-Lys-scaffold. Incorporating D-Lys in scaffold design has the potential to improve existing collagen stability and create new topologies inaccessible to homochiral molecules. This method may assist in the functionalization of the scaffold for regenerative applications.

Studies on collagen-tannic acid-collagenase ternary system: inhibition of collagenase against collagenolytic degradation of extracellular matrix component of collagen

We report the detailed studies on the inhibitory effect of tannic acid (TA) on Clostridium histolyticum collagenase (ChC) activity against degradation of extracellular matrix component of collagen. The TA treated collagen exhibited 64% resistance against collagenolytic hydrolysis by ChC, whereas direct interaction of TA with ChC exhibited 99% inhibition against degradation of collagen and the inhibition was found to be concentration dependant. The kinetic inhibition of ChC has been deduced from the extent of hydrolysis of N-[3-(2-furyl) acryloyl]-Leu-Gly-Pro-Ala (FALGPA). This data provides a selective competitive mode of inhibition on ChC activity seems to be influenced strongly by the nature and structure of TA. TA showed inhibitor activity against the ChC by molecular docking method. This result demonstrated that TA containing digalloyl radical possess the ability to inhibit the ChC. The inhibition of ChC in gaining new insight into the mechanism of stabilization of collagen by TA is discussed.