Luna Goswami

In situ FT-IR microscopic study on enzymatic treatment of poplar wood cross-sections

The feasibility of Fourier transform infrared (FT-IR) microscopy to monitor in situ the enzymatic degradation of wood was investigated. Cross-sections of poplar wood were treated with cellulase Onozuka RS within a custom-built fluidic cell. Light-optical micrographs and FT-IR spectra were acquired in situ from normal and tension wood fibers. Light-optical micrographs showed almost complete removal of the gelatinous (G) layer in tension wood. No structural and spectral changes were observed in the lignified cell walls. The accessibility of cellulose within the lignified cell wall was found to be the main limiting factor, whereas the depletion of the enzyme due to lignin adsorption could be ruled out. The fast, selective hydrolysis of the crystalline cellulose in the G-layer, even at room temperature, might be explained by the gel-like structure and the highly porous surface. Young plantation grown hardwood trees with a high proportion of G-fibers thus represent an interesting resource for bioconversion to fermentable sugars in the process to bioethanol.

Stress generation in the tension wood of poplar is based on the lateral swelling power of the G-layer

The mechanism of active stress generation in tension wood is still not fully understood. To characterize the functional interdependency between the G-layer and the secondary cell wall, nanostructural characterization and mechanical tests were performed on native tension wood tissues of poplar (Populus nigra x Populus deltoids) and on tissues in which the G-layer was removed by an enzymatic treatment. In addition to the well-known axial orientation of the cellulose fibrils in the G-layer, it was shown that the microfibril angle of the S2-layer was very large (about 36 degrees). The removal of the G-layer resulted in an axial extension and a tangential contraction of the tissues. The tensile stress-strain curves of native tension wood slices showed a jagged appearance after yield that could not be seen in the enzyme-treated samples. The behaviour of the native tissue was modelled by assuming that cells deform elastically up to a critical strain at which the G-layer slips, causing a drop in stress. The results suggest that tensile stresses in poplar are generated in the living plant by a lateral swelling of the G-layer which forces the surrounding secondary cell wall to contract in the axial direction.

Polysaccharide-capped silver Nanoparticles inhibit biofilm formation and eliminate multi-drug-resistant bacteria by disrupting bacterial cytoskeleton with reduced cytotoxicity towards mammalian cells

Development of effective anti-microbial therapeutics has been hindered by the emergence of bacterial strains with multi-drug resistance and biofilm formation capabilities. In this article, we report an efficient green synthesis of silver nanoparticle (AgNP) by in situ reduction and capping with a semi-synthetic polysaccharide-based biopolymer (carboxymethyl tamarind polysaccharide). The CMT-capped AgNPs were characterized by UV, DLS, FE-SEM, EDX and HR-TEM. These AgNPs have average particle size of ~20–40 nm, and show long time stability, indicated by their unchanged SPR and Zeta-potential values. These AgNPs inhibit growth and biofilm formation of both Gram positive (B. subtilis) and Gram negative (E. coli and Salmonella typhimurium) bacterial strains even at concentrations much lower than the minimum inhibitory concentration (MIC) breakpoints of antibiotics, but show reduced or no cytotoxicity against mammalian cells. These AgNPs alter expression and positioning of bacterial cytoskeletal proteins FtsZ and FtsA. CMT-capped AgNPs can effectively block growth of several clinical isolates and MDR strains representing different genera and resistant towards multiple antibiotics belonging to different classes. We propose that the CMT-capped AgNPs can have potential bio-medical application against multi-drug-resistant microbes with minimal cytotoxicity towards mammalian cells.

Acrylic acid grafted guargum–nanosilica membranes for transdermal diclofenac delivery

Green, hydrophobic device for controlled transdermal release of diclofenac sodium was designed from in situ nanosilica/acrylic acid grafted guargum membranes. Best grafting condition was assigned and nanocomposites were formed in situ using varying proportions of aqueous nanosilica sol. Nanocomposite/drug conjugates were formed by bringing down the medium pH from 9.0 to 7.0. The conjugates were characterized through infrared and solid state NMR spectroscopy, electron microscopy, hydro-swelling, surface contact angle, viscometry and biocompatibility. Most balanced property was exhibited by the membrane containing 1wt% nanosilica. It also had shown the highest encapsulation efficacy vis-à-vis slowest release as compared to others during experimentation in a Franz diffusion cell.

Thermosensitive ion channel TRPV1 is endogenously expressed in the sperm of a fresh water teleost fish (Labeo rohita) and regulates sperm motility

Sperm cells exhibit extremely high sensitivity in response to slight changes in temperature, osmotic pressure and/or presence of various chemical stimuli. In most cases throughout the evolution, these physico-chemical stimuli trigger Ca2+-signaling and subsequently alter structure, cellular function, motility and survival of the sperm cells. Few reports have recently demonstrated the presence of Transient Receptor Potential (TRP) channels in the sperm cells from higher eukaryotes, mainly from higher mammals. In this work, we have explored if the sperm cells from lower vertebrates can also have thermo-sensitive TRP channels. In this paper, we demonstrate the endogenous presence of one specific thermo-sensitive ion channel, namely Transient Receptor Potential Vanilloid family member sub type 1 (TRPV1) in the sperm cells collected from fresh water teleost fish, Labeo rohita. By using western blot analysis, fluorescence assisted cell sorting (FACS) and confocal microscopy; we confirm the presence of this non-selective cation channel. Activation of TRPV1 by an endogenous activator NADA significantly increases the quality as well as the duration of fish sperm movement. The sperm cell specific expression of TRPV1 matches well with our in silico sequence analysis. The results demonstrate that TRPV1 gene is conserved in various fishes, ranging from 1–3 in copy number, and it originated by fish-specific duplication events within the last 320 million years (MY). To the best of our knowledge, this is the first report demonstrating the presence of any thermo-sensitive TRP channels in the sperm cells of early vertebrates as well as of aquatic animals, which undergo external fertilization in fresh water. This observation may have implications in the aquaculture, breeding of several fresh water and marine fish species and cryopreservation of fish sperms.

Fabrication of acrylic acid grafted guar gum-multiwalled carbon nanotube hydrophobic membranes for transdermal drug delivery

Environmentally stable acrylic acid (AA) grafted guar gum (GG)-carboxy functionalized multiwalled carbon nanotube (f-MWCNT) in situ composite membranes for the sustained release of a hydrophobic drug, namely, diclofenac sodium, have been developed and characterized. Absolute matrix–filler interaction, particularly up to 1 wt% f-MWCNT concentration, instigated finer dispersion of the filler and subsequent increase in the water resistance and drug retention properties of the composites compared to the 2 and 3 wt% levels. The latter showed non-uniform filler networking and eventually resulted in poor water resistance and drug retention behaviors. The slowest drug release was achieved at the 1 wt% f-MWCNT level and fastest was observed at 3 wt% (21.5% vs. 48.5%). The releases followed a non-Fickian mechanism with the greater influence of viscoelastic relaxation at 0.5 and 1 wt% compositions compared to 2 and 3 wt% compositions, respectively.

A transdermal device from 2-hydroxyethyl methacrylate grafted carboxymethyl guar gum–multi-walled carbon nanotube composites

A hydrophobic and biocompatible transdermal device has been developed from 2-hydroxyethyl methacrylate (HEMA) grafted carnboxymethyl guar gum (CMG)-functionalized multi-walled carbon nanotube (MWCNT) in situ composite membranes. The developed composites may be used for sustained delivery of diclofenac sodium. Supreme matrix–MWCNT interaction at 0.5 and 1 wt% MWCNT concentrations induces excellent copolymer-adsorbed fibrillar orientation of MWCNT compared to that at 2 and 3 wt%. It successively leads to efficient encapsulation and more sustained release of the drug molecules at these compositions (65% vs. 17% at 1 wt%). Analyses show that the releases are dominated by the viscoelastic relaxation behavior (power law indices 0.75 and 0.80 and extremely high Deborah numbers) of the devices. Conversely, for the latter, the releases are comparatively less sustained and more swelling controlled (indices are 0.5 and 0.6 and low Deborah numbers). However, all devices have significantly increased the half life period of the drug molecules (2.5 h vs. 47 h at 1 wt%).

A carboxy methyl tamarind polysaccharide matrix for adhesion and growth of osteoclast-precursor cells

Remodeling of bone by tissue engineering is a realistic option for treating several bone-related pathophysiological ailments such as osteoporosis, bone tumor, bone cancer or abnormal bone development. But, these possibilities are hindered due to lack of proper natural and biodegradable surface on which bone precursor cells can adhere efficiently and grow further. Here we describe the synthesis and characterization of a new hydrogel as an effective surface which can acts as a material for bone tissue engineering. This hydrogel has been prepared by chemically grafting a semi-synthetic polymer with a synthetic monomer, namely hydroxyethyl methacrylate (HEMA). Carboxy methyl tamarind (CMT) was selected as the semi-synthetic polymer. The hydrogel was prepared at different mole ratios and at the ratio of 1:10 (CMT:HEMA) yielded the best hydrogel as characterized by several physico-chemical analysis such as UV spectroscopy, FT-IR spectroscopy and swelling properties. We further demonstrate that this material is suitable for effective adhesion, growth and further clustering of bone precursor cells (RAW 264.7). This material is also compatible for growing other sensitive cells such as neuronal cells (Neuro2a) and human umbilical vein endothelial cells (HUVEC) demonstrating that this surface does not possess any cytotoxicity and is compatible for primary human cells too. We conclude that the hydrogel made of CMT:HEMA at a ratio of 1:10 can be suitable for bone tissue engineering and thus may have clinical as well as commercial application in future.

Expression of temperature-sensitive ion channel TRPM8 in sperm cells correlates with vertebrate evolution.

Transient Receptor Potential cation channel, subfamily Melastatin, member 8 (TRPM8) is involved in detection of cold temperature, different noxious compounds and in execution of thermo- as well as chemo-sensitive responses at cellular levels. Here we explored the molecular evolution of TRPM8 by analyzing sequences from various species. We elucidate that several regions of TRPM8 had different levels of selection pressure but the 4th–5th transmembrane regions remain highly conserved. Analysis of synteny suggests that since vertebrate origin, TRPM8 gene is linked with SPP2, a bone morphogen. TRPM8, especially the N-terminal region of it, seems to be highly variable in human population. We found 16,656 TRPM8 variants in 1092 human genomes with top variations being SNPs, insertions and deletions. A total of 692 missense mutations are also mapped to human TRPM8 protein of which 509 seem to be delateroiours in nature as supported by Polyphen V2, SIFT and Grantham deviation score. Using a highly specific antibody, we demonstrate that TRPM8 is expressed endogenously in the testis of rat and sperm cells of different vertebrates ranging from fish to higher mammals. We hypothesize that TRPM8 had emerged during vertebrate evolution (ca 450 MYA). We propose that expression of TRPM8 in sperm cell and its role in regulating sperm function are important factors that have guided its molecular evolution, and that these understandings may have medical importance.

A Modified Polysaccharide-Based Hydrogel for Enhanced Osteogenic Maturation and Mineralization Independent of Differentiation Factors.

Bone related problems are increasing as a consequence of increased life expectancy, disorders in life style, and other medical conditions enforcing the need for functional bones prepared in vitro at affordable cost. Lack of suitable surface which promotes growth of both osteogenic and nonosteogenic cells is a major limitation. Here a novel biomaterial is reported that is synthesized from natural polysaccharide, namely, tamarind kernel polysaccharide (TKP), which is grafted with hydrophilic acrylic acid (AA) by radical polymerization. Modification in surface functionality removes unwanted proteins and alters hydrophilic/hydrophobic balance. TKP-AA is suitable for the growth of different nonosteogenic and osteogenic cells. This material is suitable for osteoblasts and promotes in vitro mineralization and differentiation without the addition of exogenous growth factors. TKP-AA can be used for the growth of mesenchymal stem cell-derived osteoblasts. It is suggested that TKP-AA can potentially be used as a scaffold for diverse cell types and particularly for bone tissue engineering at low cost.