Venkat Raghavan Krishnaswamy

A novel FRET ‘off–on’ fluorescent probe for the selective detection of Fe3+, Al3+ and Cr3+ ions: Its ultrafast energy transfer kinetics and application in live cell imaging

A rhodamine-naphthalimide dyad probe, 1, that selectively responds to the addition of trivalent metal ions (Fe(3+) or Al(3+) or Cr(3+)) via ultrafast Förster resonance energy transfer (FRET) from naphthalimide to rhodamine is designed and synthesized. 1 is highly selective to the trivalent metal ions and the presence of other monovalent or divalent metal ions do not affect its detection ability. The probe is highly sensitive and it can respond to the presence of trivalent metal ions even at sub-micromolar levels. 1 is stable over a broad range of pH, non-toxic under experimental conditions and suitable to the fluorescence bio-imaging of live cells exposed to trivalent metal ions. The trivalent metal ion induced ultrafast energy transfer kinetics of 1 is explored using time resolved fluorescence experiments.

Fabrication of highly aligned fibrous scaffolds for tissue regeneration by centrifugal spinning technology.

Centrifugal spinning (C-Spin) is an emerging technology which uses centrifugal force to produce ultrafine fibers. Being a voltage free technique it can overcome the limitations of electrospinning. Owing to the unique characteristic features such as high surface area to volume ratio, porosity, mechanical strength and fiber alignment, centrifugal spun (C-spun) fibrous mat has a wide range of scope in various biomedical applications. Higher degree of fiber alignment can be effortlessly achieved by the C-Spin process. In order to prove the versatility of C-Spin system with respect to fiber alignment, Polycaprolactone (PCL) and gelatin were spun taking them as model polymers. The morphological analysis revealed that highly aligned ultrafine fibers with smooth surface are achieved by C-Spinning. Hydrophilicity, porosity and mechanical property results confirm that the C-spun mat is more suitable for tissue engineering applications. In vitro and in vivo experiments proved that the scaffolds are biocompatible and can be efficiently used as a wound dressing material.

Recent advancements in nanotechnological strategies in selection, design and delivery of biomolecules for skin regeneration

Skin is a very complex organ and hence designing a bioengineered skin model replicating the essential physiological characteristics for replacing the diseased or damaged parts has been a challenging goal for many. Newer technologies for satisfying most of the criteria are being attempted with the copious efforts of biologists, engineers, physiologists, using multitude of features in combination. Amongst them nanotechnology based biomaterials have gained prominence owing to the enhanced pharmacokinetics, bio-distribution profile, extended half-life and reduced side effects. Designing a matrix that can be assimilated into the body during the regeneration and delivering the essential pharmacological agents in a temporal and spatially specific manner is a tremendous goal. This review essentially deals with the various approaches for designing a multidisciplinary translational smart matrix for addressing the various skin related ailments.

Role of Dermatopontin in re-epithelialization: Implications on keratinocyte migration and proliferation

Re-epithelialization is a key event in wound healing and any impairment in that process is associated with various pathological conditions. Epidermal keratinocyte migration and proliferation during re-epithelialization is largely regulated by the cytokines and growth factors from the provisional matrix and dermis. Extracellular matrix consists of numerous growth factors which mediate cell migration via cell membrane receptors. Dermatopontin (DPT), a non-collagenous matrix protein highly expressed in dermis is known for its striking ability to promote cell adhesion. DPT also enhances the biological activity of transforming growth factor beta 1 which plays a central role in the process of wound healing. This study was designed to envisage the role of DPT in keratinocyte migration and proliferation along with its mRNA and protein expression pattern in epidermis. The results showed that DPT promotes keratinocyte migration in a dose dependant fashion but fail to induce proliferation. Further, PCR and immunodetection studies revealed that the mRNA and protein expression of DPT is considerably negligible in the epidermis in contrast to the dermis. To conclude, DPT has a profound role in wound healing specifically during re-epithelialization by promoting keratinocyte migration via paracrine action from the underlying dermis.

Expression and integrity of dermatopontin in chronic cutaneous wounds: a crucial factor in impaired wound healing

Chronic cutaneous wound (CCW) is a major health care burden wherein the healing process is slow or rather static resulting in anatomical and functional restriction of the damaged tissue. Dysregulated expression and degradation of matrix proteins, growth factors and cytokines contribute to the disrupted and uncoordinated healing process of CCW. Therefore, therapeutic approaches for effective management of CCW should be focused towards identifying and manipulating the molecular defects, such as reduced bioavailability of the pro-healing molecules and elevated activity of proteases. This study essentially deals with assessing the expression and integrity of an extracellular matrix protein, Dermatopontin (DPT), in CCW using real-time quantitative reverse transcriptase PCR and immunological techniques. The results indicate that, despite DPT’s high mRNA expression, the protein levels are markedly reduced in both CCW tissue and its exudate. To elucidate the cause for this contradiction in mRNA and protein levels, the stability of DPT is analyzed in the presence of wound exudates and various proteases that are naturally elevated in CCW. DPT was observed to be degraded at higher rates when incubated with certain recombinant proteases or chronic wound exudate. In conclusion, the susceptibility of DPT protein to specific proteases present at high levels in the wound milieu resulted in the degradation of DPT, thus leading to impaired healing response in CCW.

Keloid collagen–cell interactions: structural and functional perspective

Keloids are a benign dermal proliferative disorder characterised by dense fibrotic tissue developing due to abnormal wound healing. Clinically, keloids extend beyond the margin of the original wound, do not undergo spontaneous regression and also recur after excision. The biochemical and cellular composition of keloids largely differ from that of normal dermis and mature scars plaguing physicians and patients for several decades. Keloids are rich in densely packed fibrillar collagen paralleled by up regulation of fibrogenic cytokines and growth factors. Adhesion of cells to the extracellular matrix (ECM) is a fundamental process during the formation and maintenance of animal tissue and therefore, aberrant cell–ECM interactions result in a number of diseases. There are numerous reports on the signalling mechanisms responsible for excessive production of collagen in keloids. However, the structure–function correlation of collagen fibres in keloids is not understood properly. This paper is the first report towards the understanding of the influence of keloid collagen on the behaviour of fibroblasts with reference to cell adhesion, spreading and growth that translates the adhesion mediated signalling response in keloid pathogenesis. Cells grown on normal collagen were well spread, adhered and proliferated while those on keloid collagen exhibited weak cell–matrix interactions and hence exhibited a lower degree of proliferation. The differences observed in the cell behaviour could be attributed to the altered structural and thermal properties of the keloid collagen.

Dermatopontin augments angiogenesis and modulates the expression of transforming growth factor beta 1 and integrin alpha 3 beta 1 in endothelial cells

Dermatopontin (DPT) is a matricellular protein with cardinal roles in cutaneous wound healing. The protein is also reported to be altered in various anomalies including cancer. The present study is aimed to unravel the role of DPT in angiogenesis which is imperative in many physiological and pathological processes. DPTs capabilities on promoting angiogenesis were assessed using various in vitro and ex vivo systems. The results indicated that DPT enhances cell motility and induces lamellipodia formation in endothelial cells. Additionally, we noticed that DPT stimulates tube formation in endothelial cells when plated on a matrigel substrate. However, it was observed that DPT had no effect on the proliferation of endothelial cells even at higher concentrations and prolonged treatment periods. Additional experiments on CAM and aortic arch assays apparently depicted that DPT promotes neovascularisation and tube sprouting, thus unraveling its prominent role in angiogenesis. Further, PCR analysis revealed that endothelial cells are devoid of DPT expression, but when exogenously supplied, modulates the expression of transforming growth factor β1 and integrin α3β1 which are reported to have crucial roles in endothelial cell behaviour during angiogenesis. In conclusion, DPT possess vital pro-angiogenic properties and thus retains promising therapeutic values in managing chronic wounds and cancer.