Vellore Institute of Technology (VIT) is a private university located in&nbsp;Tamil Nadu, India. Founded in 1984, as Vellore Engineering College, the institution offers 20 undergraduate, 34 postgraduate, four integrated and four research programs. It has campuses in Vellore, Amravati, Bhopal and Chennai.

VIT is one of the top ranked private universities in India according to NIRF, THE and QS Rankings.&nbsp;Govt. of India has recognized&nbsp;VIT, Vellore as an&nbsp;Institution of Eminence. This has allowed VIT to take independent quality initiatives and move up in world ranking.

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VIT University

Department of Bio-Sciences

Biphasic calcium phosphate (BCP) ceramics consisting of hydroxyapatite (HA) and tricalcium phosphate (TCP) has been used as a bone graft material during the last decade. In this paper, we report the bone in-growth induced by BCP ceramic in the experimentally created circular defects in the femur of dogs. This BCP ceramic consists of 55% hydroxyapatite (HA) and 45% β-tricalcium phosphate (TCP) prepared in situ by the microwave method. The defects were created as 4-mm holes on the lateral aspect of the femur of dogs and the holes were packed with the implant material. The defective sites were radiographed at a period of 4, 8, and 12 weeks postoperatively. The radiographical results showed that the process of ossification started after 4 weeks and the defect was completely filled with new woven bone after 12 weeks. Histological examination of the tissue showed the formation of osteoblast inducing the osteogenesis in the defect. The collageneous fibrous matrix and the complete Haversian system were observed after 12 weeks. The blood serum was collected postoperatively and biochemical assays for alkaline phostaphase activity were carried out. The measurement of alkaline phostaphase activity levels also correlated with the formation of osteoblast-like cells. This microwave-prepared BCP ceramic has to proved to be a good biocompatible implant as well as osteoconductive and osteoinductive materials to fill bone defects. © 2005 Sage Publications.

School of Bio Sciences and Technology

Vellore Campus

World University root node

Journal of Biomaterials Applications

Bone in-growth induced by biphasic calcium phosphate ceramic in femoral defect of dogs

Background: With microarray technology the expression of thousands of genes can be measured simultaneously. It is well known that the expression levels of genes of interacting proteins are correlated significantly more strongly in Saccharomyces cerevisiae than those of proteins that are not interacting. The objective of this work is to investigate whether this observation extends to the human genome. Results: We investigated the quantitative relationship between expression levels of genes encoding interacting proteins and genes encoding random protein pairs. Therefore we studied 1369 interacting human protein pairs and human gene expression levels of 155 arrays. We were able to establish a statistically significantly higher correlation between the expression levels of genes whose proteins interact compared to random protein pairs. Additionally we were able to provide evidence that genes encoding proteins belonging to the same GO-class show correlated expression levels. Conclusion: This finding is concurrent with the naive hypothesis that the scales of production of interacting proteins are linked because an efficient interaction demands that involved proteins are available to some degree. The goal of further research in this field will be to understand the biological mechanisms behind this observation. © 2005 Hahn et al; licensee BioMed Central Ltd.

Fulltext

BMC Bioinformatics

Confirmation of human protein interaction data by human expression data

Bone and cartilage generation by three-dimensional scaffolds is one of the promising techniques in tissue engineering. One approach is to generate histologically and functionally normal tissue by delivering healthy cells in biocompatible scaffolds. These scaffolds provide the necessary support for cells to proliferate and maintain their differentiated function, and their architecture defines the ultimate shape. Rapid prototyping (RP) is a technology by which a complex 3-dimensional (3D) structure can be produced indirectly from computer aided design (CAD). The present study aims at developing a 3D organic-inorganic composite scaffold with defined internal architecture by a RP method utilizing a 3D printer to produce wax molds. The composite scaffolds consisting of chitosan and hydroxyapatite were prepared using soluble wax molds. The behaviour and response of MC3T3-E1 pre-osteoblast cells on the scaffolds was studied. During a culture period of two and three weeks, cell proliferation and in-growth were observed by phase contrast light microscopy, histological staining and electron microscopy. The Giemsa and Gömöri staining of the cells cultured on scaffolds showed that the cells proliferated not only on the surface, but also filled the micro pores of the scaffolds and produced extracellular matrix within the pores. The electron micrographs showed that the cells covering the surface of the struts were flattened and grew from the periphery into the middle region of the pores. © 2005 Springer Science + Business Media, Inc.

Journal of Materials Science: Materials in Medicine

Biomimetic mineral-organic composite scaffolds with controlled internal architecture

The material bone consists of a biopolymer matrix (collagen) reinforced with mineral nanoparticles (carbonated hydroxylapatite), forming a natural composite which builds up a dense shell on the exterior and a network of struts with a mean diameter of 200μm in the core of many bones. The architecture of the foamy inner part of bones (spongiosa) is determined by loading conditions. The architecture strongly influences the mechanical properties of cellular solids together with the apparent density and the material it consists of. In addition, the ingrowth of bone cells into porous implants depends on pore size, size distribution and interconnectivity. From this it is clear that the possibility to design the architecture of a bone replacement material is beneficial from a biological as well as a mechanical point of view. Our approach uses rapid prototyping methods, ceramic gelcasting and sintering to produce cellular structures with designed architecture from hydroxylapatite and other bioceramics. The influence of sintering temperature and atmosphere on the physical properties of these scaffolds was investigated with x-ray diffraction and scanning electron microscopy. Furthermore, the cell ingrowth behaviour was determined in cell culture experiments, using the praeosteoblastic cell line MC3T3-E1, derived from mouse calvariae. The cell ingrowth behaviour was evaluated during a culture period of two and three weeks, by light microscopy and afterwards by histology after embedding and Giemsa-staining. The phase composition of the material was found to change with increasing sintering temperature and its surface characteristics was influenced by the sintering atmosphere. These changes also affected the cell ingrowth behaviour. In some experiments, the osteoblasts-like cells were found to cover the whole external and internal surface of the scaffold. The cells produced extracellular matrix consisting of collagen, which eventually filled nearly all the pores. In particular, the cells had the tendency to fill any crack or opening in the scaffolds, and to generally smooth the surfaces. In conclusion, rapid prototyping and ceramic gelcasting allows the freeform fabrication of porous bioceramics with controlled architecture. Such structures made of hydroxylapatit were found to support the growth of mouse osteoblasts. © 2005 Materials Research Society.

Materials Research Society Symposium Proceedings

The influence of the thermal treatment of hydroxylapatite scaffolds on the physical properties and the bone cell ingrowth behaviour

Nitrosative stress plays a role in calcium oxalate stone formation, as nitrosated proteins have been identified in stone formers. Nitric oxide (NO •), the common precursor for reactive nitrogen species, is synthesized in the juxtaglomerular apparatus of the kidneys. The present study is aimed to determine the role of nitric oxide synthase (NOS) in an experimental hyperoxaluric condition by histological and biochemical techniques. Hyperoxaluria was induced by 0.75% ethylene glycol in drinking water. L-arginine (L-arg) was supplemented at a dose of 1.25 g/kg body weight orally for 28 days. Nitric oxide metabolites (NOx), protein content in the urine and lipid peroxidation in the kidney were determined at the end of the experimental period. Histopathological examination of the rat kidneys was then carried out. NADPH-diaphorase and eNOS expression studies were carried out in control and hyperoxaluric rat kidneys using histochemical and immunohistochemical techniques. Significant amounts of NOx were present in the urine of hyperoxaluric animals when compared to control rats. Histopathological examinations revealed membrane injury, tubular dilatation and edema in the hyperoxaluric rats, whereas co-supplementation of L-arg to the hyperoxaluric rats significantly reduced these changes. The results of histochemical analysis for NADPH-diaphorase staining demonstrate the role of NOS in hyperoxaluric rats. Hyperoxaluric rats showed intense staining for NADPH-diaphorase when compared to control and L-arg co-supplemented hyperoxaluric rats. Immunohistochemical demonstration confirmed that eNOS expression was markedly increased in L-arg supplemented rats, when compared to EG treated rat kidney sections. Thus, from the present study, we conclude that supplementation of L-arg to the hyperoxaluric animals minimizes the cellular injury mediated by ethylene glycol, prevents oxidative/nitrosative damage to the membranes and reduces the incidence of calcium oxalate stone formation. © Springer-Verlag 2005.

Urological Research

Detection of endothelial nitric oxide synthase and NADPH-diaphorase in experimentally induced hyperoxaluric animals

Background: Free radical induced renal damage leads to crystal retention and formation of large stones. However, the scenario behind uric acid (UA) stone formation is still a mystery, as uric acid, a risk factor of stone formation, seems to be a potent antioxidant that can protect cells from damage by reactive oxygen species. This study was intended to evaluate the role of uric acid in stone formers by assessing the oxidative stress status of the stone patients. Methods: Determination of urinary stone forming risk factors and oxidative stress factors like plasma lipid peroxidation, protein carbonyls of stone formers and histopathological changes and uric acid deposition in stone patients kidney biopsy were studied. Results: Increased concentrations of urinary uric acid and oxalate in both uric acid as well as calcium oxalate stone formers were observed, whereas calcium is increased in calcium stone formers and not in the uric acid stone patients. Inhibitors such as citrate and glycosaminoglycans (GAGs) were found to be significantly decreased in all the stone patients. Histopathological studies confirmed the deposition of crystals in the damaged tubules and De Galantha staining authenticates that the damage is caused due to uric acid crystals. Increased oxidative stress is dictated by the concentrations of lipid peroxidation and protein carbonyls in stone formers. Moreover, increased activities of urinary marker enzymes substantiate the tubular damage. Conclusion: We speculated that uric acid acts as a calculi forming salt rather than an antioxidant and it has no role in preventing oxidative stress pertaining to urolithiasis. © 2004 Elsevier B.V. All rights reserved.

Clinica Chimica Acta

Uric acid: An abettor or protector in calcium oxalate urolithiasis? Biochemical study in stone formers

Myosin 10 contains a region of predicted coiled coil 120 residues long. However, the highly charged nature and pattern of charges in the proximal 36 residues appear incompatible with coiled-coil formation. Circular dichroism, NMR, and analytical ultracentrifugation show that a synthesized peptide containing this region forms a stable single α-helix (SAH) domain in solution and does not dimerize to form a coiled coil even at millimolar concentrations. Additionally, electron microscopy of a recombinant myosin 10 containing the motor, the three calmodulin binding domains, and the full-length predicted coiled coil showed that it was mostly monomeric at physiological protein concentration. In dimers the molecules were joined only at their extreme distal ends, and no coiled-coil tail was visible. Furthermore, the neck lengths of both monomers and dimers were much longer than expected from the number of calmodulin binding domains. In contrast, micrographs of myosin 5 heavy meromyosin obtained under the same conditions clearly showed a coiled-coil tail, and the necks were the predicted length. Thus the predicted coiled coil of myosin 10 forms a novel elongated structure in which the proximal region is a SAH domain and the distal region is a SAH domain (or has an unknown extended structure) that dimerizes only at its end. Sequence comparisons show that similar structures may exist in the predicted coiled-coil domains of myosins 6 and 7a and MyoM and could function to increase the size of the working stroke.

Journal of Biological Chemistry

The predicted coiled-coil domain of myosin 10 forms a novel elongated domain that lengthens the head

Background: Oral supplementation of L-arginine (L-arg) is found to be beneficial in many kidney disorders. We determined whether L-arg supplementation safeguards the renal epithelial cell damage induced by hyperoxaluria with excretion of urinary marker enzymes and lithogenic salts with special reference to Tamm-Horsfall glycoprotein (THP). Methods: Hyperoxaluria was induced by 0.75% ethylene glycol (EG) in drinking water. L-Arg was co-supplemented at the dose of 1.25 g/kg b.w. orally for 28 days. At the end of experimental period, 24-h urine samples were collected in all the experimental groups. Isolation and purification of THP was carried in rat urine and were subjected to spectrophotometric crystallization assay and calcium-14C-oxalate binding studies. Determination of the lithogenic risk factors like calcium, oxalate, phosphorus, citrate, and marker enzymes such as lactate dehydrogenase (LDH) and γ-glutamyltransferase (γ-GT) were carried out in the collected urine sample. Results: Urinary excretion of calcium and oxalate was significantly increased in EG-treated rats. In L-arg supplemented hyperoxaluric rats, these concentrations were significantly (p &lt; 0.001) decreased when compared to that of hyperoxaluric rats, and were moderately elevated from that of control rats. The activities of urinary marker enzymes, both LDH and γ-GT were 2-fold increased in EG-treated rats, when compared to control rats, but these values were maintained near normal in L-arg supplemented EG-treated rats. Citrate excretion was enhanced in the L-arg co-supplemented hyperoxaluric rats. In spectrophotometric crystallization assay system, L-arg supplemented rat THP showed inhibition in nucleation and aggregation phases, whereas EG-treated rat THP showed promotion of both calcium oxalate nucleation and aggregation phases. In calcium-14C-oxalate binding assay, THP derived from hyperoxaluric rats exhibited 2-fold increase (p &lt; 0.001) in the Ca* Ox binding when compared to control and L-arg supplemented animals. Conclusions: L-Arg could act as a potent antilithic agent, by increasing the level of citrate in the hyperoxaluria-induced rats and decreasing calcium oxalate binding to the THP. L-Arg also effectively prevents the deposition of calcium oxalate crystals by curtailing the renal epithelial damage and protein oxidation as evidenced by the normal activities of urinary marker enzymes in L-arg supplemented hyperoxaluric rats. © 2005 Elsevier B.V. All rights reserved.

Oral L-arginine supplementation ameliorates urinary risk factors and kinetic modulation of Tamm-Horsfall glycoprotein in experimental hyperoxaluric rats

Background. This study aimed to assess the therapeutic efficacy of oral vitamin E supplementation on the biochemical and kinetic properties of Tamm-Horsfall glycoprotein (THP) in hypertensive and hyperoxaluric patients. Methods. Newly detected hypertensives (n = 200) and stone formers (n = 200) were each subdivided into two groups. One group (n = 100) was administered the antioxidant vitamin E at 400 mg/day given as an oral supplement along with standard therapeutic drugs for hypertension and hyperoxaluria and the patients were followed for a period of 9 months. The other group (n = 100) did not receive vitamin E (placebo controls). Age and sex-matched controls (n = 100) were monitored simultaneously. THP was isolated from 24 h urine samples before and at the end of every third month during a period of 9 months from the vitamin E-treated hypertensive and hyperoxaluric groups. THP samples were also collected from control subjects, and at the end of the ninth month from placebo controls. The isolated protein was assessed for purity by SDS-PAGE. The purity-checked proteins were subjected to spectrophotometric crystallization assay, calcium oxalate (CaOx) crystal interaction studies, and biochemical analysis of sialic acid, thiol and carbonyl content. Plasma superoxide, hydroxyl radical, hydrogen peroxide and vitamin E levels as well as superoxide dismutase and catalase activities were also monitored. Results. The THP from the hypertensive and hyperox aluric subjects exhibited a significant promoting effect on the nucleation and aggregation phases and caused a concomitant increase in CaOx crystal interaction. The altered kinetic properties of THP in these subjects were strongly associated with increased carbonyl content and with decreased thiol and sialic acid contents. Oral administration of vitamin E to these patients caused near normalization of these biochemical alterations and satisfactorily restored the kinetic properties of THP to near normal activity. At the end of 9 months, THP isolated from placebo controls (hypertensive and hyperoxaluric) showed highly aggregated calcium oxalate monohydrate crystals as observed by light microscopy. In contrast, vitamin E-supplemented patients showed CaOx dihydrate crystals that were similar to control THP. There was an imbalance in the oxidant and antioxidant levels. For the oxidants, superoxide, hydrogen peroxide and hydroxyl radical levels were increased, and for the antioxidants, there was loss of antioxidant enzyme activities and a decline in plasma vitamin E level in both hypertensive and hyperoxaluric patients. Supplementary antioxidant (vitamin E) corrected this imbalance to near normal conditions. Conclusion. We hypothesize that the loss of THP inhibitory activity in the hypertensive and hyperoxaluric patients in a crystallizing medium is mediated primarily by oxidative damage to this protein. The possible occurrence of renal stones in essential hypertensive subjects, and the risk of recurrence in hyperoxaluric subjects, may be explained by oxidative damage to renal tissues that remained unchecked by standard drug therapies. The normalization of the kinetic properties of THP following vitamin E supplementation is in support of our hypothesis. © The Author [2005]. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.

Nephrology Dialysis Transplantation

Beneficial effect of vitamin E supplementation on the biochemical and kinetic properties of Tamm-Horsfall glycoprotein in hypertensive and hyperoxaluric patients

Background: Our understanding of nitrosative stress in the process of urolithiasis is far from complete. Earlier studies carried out in our laboratory demonstrate the presence of nitrated THP in stone formers, l-arginine (l-arg) a precursor of nitric oxide (NO), attenuates the endothelial dysfunction caused by reactive nitrogen species. We investigated the role of l-arg in ethylene glycol (EG)-induced urolithic rat model and observed its antilithic and antioxidative properties. Methods: Hyperoxaluria was induced using 0.75% EG in drinking water. l-arg [1.25 g/kg body weight] was given orally for a period of 28 days. Results: EG-treated rats showed significant loss in body weight and increase in the activities of oxalate synthesizing enzymes such as glycollic acid oxidase in liver. Lactate dehydrogenase activity in liver and kidney was increased. The activity of the free radical producing enzyme xanthine oxidase, tissue oxalate and calcium levels were significantly increased in EG-treated rats. Depletion in the antioxidant enzymes, membrane bound ATPases and thiol status was observed in these rats. l-arg co-supplementation to EG-treated rats maintained the activities of the oxalate synthesizing enzymes and free radical producing enzymes with in the normal range. Tissue oxalate and calcium levels were also maintained near normal in l-arg treated hyperoxaluric rats. l-arg, by its cytoprotective effect, maintained the thiol status, thereby preserving the activities of the membrane bound ATPases and preventing proteinuria and subsequent weight loss in EG-treated rats. Conclusion: l-arg feeding prevents the retention of calcium oxalate crystals in hyperoxaluric rats by way of protecting the renal cells from oxidative injury and also by providing a second line of defense through the normalization of the oxalate metabolism. It reduces the risk of stone formation, by curtailing free radicals and hyperoxaluria as both of them have to work in close association to form stones. © 2004 Elsevier B.V. All rights reserved.