Pradeep Srivastava

5‐Florouracil‐loaded poly(lactic acid)‐poly(caprolactone) hybrid scaffold: Potential chemotherapeutic implant

Hybrid nanofibers of poly(lactic acid) and polycaprolactone have been developed by embedding cancerous drug through electrospinning technique. The composition of polymer has been varied to check the compositional effect on properties. The quality of nanofibers has been testified through surface morphology, wetting properties using contact angle and mechanical strength under uniaxial elongation. The compatibility of drug (5-fluorourasil) with matrix fiber has been verified using Fourier transform infrared, X-ray diffraction, Raman spectroscopy, and differential scanning calorimetry. The drug release study has been performed showing greater release in hybrid fibers when compared with pure polymers as a result of synergism of two immiscible polymers and quasi-Fickian diffusion mechanism in hybrid nanofiber as implants showing compositional effect on drug release. A model has been proposed showing faster release of drugs in hybrid systems. Biological responses through fluorescence imaging and MTT assay confirm the release of drug from hybrid nanofibers showing potential use of hybrid scaffolds as chemotherapeutic implant.

Evaluation of poly(L-lactide) and chitosan composite scaffolds for cartilage tissue regeneration

Abstract The present study delineates the development of chitosan and poly(L-lactide) (PLLA) scaffolds cross-linked using a mixture of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC), n-hydroxysuccinimide (NHS), and chondroitin sulfate (CS) for cartilage tissue engineering applications. Chitosan and PLLA were varied in concentration for developing scaffolds and prepared by freeze-drying method. The various scaffolds were studied using scanning electron microscopy (SEM), porosity by mercury intrusion porosimeter, and the molecular interactions among polymers using Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD) studies. Differential scanning calorimetry was used to predict the thermal properties of the scaffolds. The mechanical properties of the scaffolds were studied using static mechanical tester. The ability of the scaffolds to support chondrocyte proliferation was also studied. The microscopy suggests that the pore size of the scaffolds varied with the composition in the range of 38–172 μm and the porosities in the range of 73–93%. The XRD and the FTIR studies suggested that an alternation in the composition of the scaffolds altered the molecular interactions among the scaffold components. An increase in the chitosan content enhanced the swelling property. The degradation of the scaffolds was least when the proportion of chitosan and PLLA was in the ratio of 70:30. The in vitro cell proliferation study suggested that the developed scaffolds were able to support chondrogenesis, the glycosaminoglycan (GAG) content of the mature chondrocyte was 40 μg/ml and the viability was approximately 90%. Hence, the so designed scaffolds may be tried for cartilage tissue engineering applications.

Studies on process optimization methods for rapamycin production using Streptomyces hygroscopicus ATCC 29253

Rapamycin is a high-value product finding immense use as a drug, in organ transplantation, and as a potential immunosuppressant. Optimization of fermentation parameters of rapamycin production by Streptomyceshygroscopicus NRRL 5491 has been carried out. The low titer value of rapamycin in the original producer strain limits its applicability at industrial level. This study aims at improving the production of rapamycin by optimizing the nutrient requirements. Addition of l-lysine increased the production of rapamycin up to a significant level which supports the fact that it acts as precursor for rapamycin production, as found in previous studies. Effect of optimized medium on the Streptomyces growth rate as well as rapamycin production has been studied. The optimization study incorporates one at a time parameter optimization studies followed by tool-based hybrid methodology. This methodology includes the Plackett–Burman design (PBD) method, artificial neural networks (ANN), and genetic algorithms (GA). PBD screened mannose, soyabean meal, and l-lysine concentrations as significant factors for rapamycin production. ANN was used to construct rapamycin production model. This strategy has led to a significant increase of rapamycin production up to 320.89xa0mg/L at GA optimized concentrations of 25.47, 15.39, and 17.48xa0g/L for mannose, soyabean meal, and l-lysine, respectively. The present study must find its application in scale-up study for industrial level production of rapamycin.

Design and evaluation of chitosan/poly(l-lactide)/pectin based composite scaffolds for cartilage tissue regeneration

Poor regenerative potential of cartilage tissue due to the avascular nature and lack of supplementation of reparative cells impose an important challenge in recent medical practice towards development of artificial extracellular matrix with enhanced neo-cartilage tissue regeneration potential. Chitosan (CH), poly (l-lactide) (PLLA), and pectin (PC) compositions were tailored to generate polyelectrolyte complex based porous scaffolds using freeze drying method and crosslinked by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), N-hydroxysuccinimide (NHS) solution containing chondroitin sulfate (CS) to mimic the composition as well as architecture of the cartilage extracellular matrix (ECM). The physical, chemical, thermal, and mechanical behaviors of developed scaffolds were done. The scaffolds were porous with homogeneous pore structure with pore size 49-170μm and porosities in the range of 79 to 84%. Fourier transform infrared study confirmed the presence of polymers (CH, PLLA and PC) within the scaffolds. The crystallinity of the scaffold was examined by the X-ray diffraction studies. Furthermore, scaffold shows suitable swelling property, moderate biodegradation and hemocompatibility in nature and possess suitable mechanical strength for cartilage tissue regeneration. MTT assay, GAG content, and attachment of chondrocyte confirmed the regenerative potential of the cell seeded scaffold. The histopathological analysis defines the suitability of scaffold for cartilage tissue regeneration.

Cytotoxic T- Lymphocyte Antigen-4 (CTLA4) Gene Expression and Urinary CTLA4 Levels in Idiopathic Nephrotic Syndrome

ObjectivesTo detect Cytotoxic T- Lymphocyte Antigen-4 (CTLA4) single nucleotide polymorphisms (SNPs) at +49A/G (rs231775) and -318C/T (rs5742909) positions in children with idiopathic nephrotic syndrome (INS) and also assay urinary soluble CTLA4 (sCTLA4) levels in children with minimal change disease (MCD), focal segmental glomerulosclerosis (FSGS) and steroid sensitive nephrotic syndrome (SSNS) in remission.MethodsThe study included 59 patients of INS (MCD-23, FSGS-15 and SSNS in remission-21) and 35 healthy controls. The CTLA4 SNPs profiling was done in peripheral blood mononuclear cells and urinary sCTLA4 level was assayed by ELISA kit.ResultsAlthough frequency of homozygous +49 GG (rs4553808) genotype (26.3% vs. 11.4%; pu2009=u20090.231) and G allele (52.6% vs. 40%; pu2009=u20090.216) were found to be higher in INS as compared to controls, the differences were statistically non-significant. Genotypes GG, AG, AA and alleles A and G frequencies were comparable among MCD, FSGS and controls. SNP at −318 C/T (rs5742909) did not show homozygous TT genotype both in INS as well as controls. Median urinary sCTLA4/creatinine level was significantly higher in MCD as compared to FSGS (pu2009=u20090.027), SSNS in remission (pu2009=u20090.001) and controls (pu2009=u20090.003).ConclusionsThe positive associations of +49 GG genotype and G allele in patients with nephrotic syndrome were not observed. The frequencies did not differ significantly among MCD, FSGS and controls. Urinary sCTLA4 level was significantly increased in MCD; suggesting its possible role in the pathogenesis of disease.

Strategies on process engineering of chondrocyte culture for cartilage tissue regeneration

The current work is an attempt to study the strategies for cartilage tissue regeneration using porous scaffold in wavy walled airlift bioreactor (ALBR). Novel chitosan, poly (l-lactide) and hyaluronic acid based composite scaffold were prepared. The scaffolds were cross-linked with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide, N-hydroxysuccinimide and chondroitin sulfate to obtain interconnected 3D microstructure showing excellent biocompatibility, higher cellular differentiation and increased stability. The surface morphology and porosity of the scaffolds were analyzed using scanning electron microscopy (SEM) and mercury intrusion porosimeter and optimized for chondrocyte regeneration. The study shows that the scaffolds were highly porous with pore size ranging from 48 to 180xa0µm and the porosities in the range 80–92%. Swelling and in vitro degradation studies were performed for the composite scaffolds; by increasing the chitosan: HA ratio in the composite scaffolds, the swelling property increases and stabilizes after 24xa0h. There was controlled degradationxa0of composite scaffolds for 4 weeks. The uniform chondrocyte distribution in the scaffold using various growth modes in the shake flask and ALBR was studied by glycosaminoglycans (GAG) quantification, MTT assay and mixing time evaluation. The cell culture studies demonstrated that efficient designing ofxa0ALBR increases the cartilage regeneration as compared to using a shake flask. The free chondrocyte microscopy and cell attachment were performed by inverted microscope and SEM, and from the study it was confirmed that the cells uniformly attached to the scaffold. This study focuses on optimizing strategies for the culture of chondrocyte using suitable scaffold for improved cartilage tissue regeneration.

Combined 3D QSAR Based Virtual Screening and Molecular Docking Study of Some Selected PDK-1 Kinase Inhibitors

Phosphoinositide-dependent kinase-1 (PDK-1) is an important therapeutic target for the treatment of cancer. In order to identify the important chemical features of PDK-1 inhibitors, a 3D QSAR pharmacophore model was developed based on 21 available PDK-1 inhibitors. The best pharmacophore model (Hypo1) exhibits all the important chemical features required for PDK-1 inhibitors. The correlation coefficient, root mean square deviation (RMSD), and cost difference were 0.96906, 1.0719, and 168.13, respectively, suggesting a good predictive ability of the model (Hypo1) among all the ten pharmacophore models that were analyzed. The best pharmacophore model (Hypo1) was further validated by Fisher’s randomization method (95%), test set method , and the decoy set with the goodness of fit (0.73). Further, this validated pharmacophore model Hypo1 was used as a 3D query to screen the molecules from databases like NCI database and Maybridge. The resultant hit compounds were subsequently subjected to filtration by Lipinski’s rule of five as well as the ADMET study. Docking study was done to refine the retrieved hits and as a result to reduce the rate of false positive. Best hits will further be subjected to in vitro study in future.