Thiolated Polymeric Hydrogels for Biomedical Applications: A Review.

Hydrogels are a three-dimensional (3D) network of hydrophilic polymers. The physical and chemical crosslinking of polymeric chains maintains the structure of the hydrogels even when they are swollen in water. They can be modified with thiol by thiol epoxy, thiol-ene, thiol-disulfide, or thiol-one reactions. Their application as a matrix for protein and drug delivery, cellular immobilization, regenerative medicine, and scaffolds for tissue engineering was initiated in the early 21st century. This review focuses on the ingredients, classification techniques, and applications of hydrogels, types of thiolation by different thiol-reducing agents, along with their mechanisms. In this study, different applications for polymers used in thiolated hydrogels, including dextran, gelatin, polyethylene glycol (PEG), cyclodextrins, chitosan, hyaluronic acid, alginate, poloxamer, polygalacturonic acid, pectin, carrageenan gum, arabinoxylan, carboxymethyl cellulose (CMC), gellan gum, and polyvinyl alcohol (PVA) are reviewed.

Pretreatment with troxerutin protects/improves neurological deficits in a mouse model of traumatic brain injury.

Traumatic brain injury (TBI) is the major and leading cause of mortality and an alarming public health challenge. TBI leads to permanent cognitive, motor, sensory and psychotic disabilities. Patients suffering from the various and long-term repercussions of TBI currently have limited therapy choices. The current research work was designed to evaluate the beneficial and neuroprotective role of Troxerutin (Trox) (a natural flavonoid) in a closed brain injury mouse model. The male BALB/c 8-weeks old mice (n꞊150) were randomly distributed in three experimental groups. Control group of mice (n꞊50), TBI group (n꞊50) and Trox pre-treated mice group (Trox + TBI, n꞊50). The mice in Trox + TBI were pre-treated with Trox (150 mg/kg, 7 days) before TBI. The weight-drop mechanism was used to induce mild-moderate injury in mice in both the groups. Our results showed that the mice pre-treated with troxerutin significantly improved neurological severity score, blood glucose level, food intake and brain edema as compared to the mice in the TBI group. Furthermore, compared to the TBI group, the mice treated with troxerutin improved cognitive behavior as evaluated by Open field test, Shallow Water Maze and Y-Maze, decreased brain-infarct volume and blood-brain barrier (BBB) permeability, significantly decreased Reactive Oxygen Species (ROS), improved neuronal morphology and survival in the brain regions such as cortex and hippocampus. In summary, our data provided evidence that pre-treatment with troxerutin improved neurological functions, decreased the BBB permeability, improved behavior, reduced ROS and increased neuronal survival in the weight-drop close head traumatic injury mouse model.

Impact of an indigenously produced multi-enzyme complex from Bacillus subtilis KT004404 on growth and blood parameters in broiler chicken.

A 42-days experiment was conducted on a day old birds (n = 400) to evaluate the effect of enzyme supplements in feed on the growth, blood parameters, phosphorous content in bones, and nitrogen retention. Different treatments included: control (C) without enzyme supplement, while the other three groups included enzyme mixture T1 and T2 with two commercially available enzyme mix, and T3 with indigenously produced multi-enzyme complex from Bacillus subtilis KT004404. Birds that were fed with indigenously produced multi-enzyme complex showed significant weight gain as compared to other groups. The total feed intake of the birds fed with enzyme supplements was higher than the birds in the control group. The feed conversion ratio was significantly improved (p < 0.05) in treatment groups (T1, T2, T3) as compared to the control. The blood parameters which were analyzed included uric acid, triglycerides, total cholesterol, and serum proteins i.e. globulin and albumin. Birds fed with the enzyme in the group T1, T2 and T3 exhibited higher (p < 0.05) body weight gain. Tibia ash content was significantly higher (p < 0.05) in T1, T2, and T3 as compared to the control. The results of the current study indicate that supplementing poultry feed with the exogenous multi-enzyme produced from Bacillus subtilis KT004404 improved the growth of the birds, feed utilization, and exhibited beneficial effects on the blood parameters, phosphorous and nitrogen retention in broiler chicken.

Microplastic pollution in the surface water and sediments from Kallar Kahar wetland, Pakistan: occurrence, distribution, and characterization by ATR-FTIR.

This study reports the distribution of microplastics in surface water and sediments collected from Kallar Kahar wetland, Punjab, Pakistan, which is a game reserve and hosts migratory birds during winter season. Microplastics were extracted using density separation and wet oxidation method. The microplastics identification was done under a stereo-microscope, and their polymer compositions were characterized using an attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. The average abundance of microplastics in water and sediment samples was 88 ± 14.5 items/L and 5720 ± 2580 items/kg, respectively. The dominant shape groups of microplastics in water were fiber (58.7%), irregular fragments (32.4%), and beads (8.7%) with dominant colors as transparent > black > yellow ≈ white > red > green > pink > blue. Similar distribution in sediments was found, i.e., fiber (61.2%), irregular fragments (28.4%), and beads (10.3%) with dominant colors as transparent > pink > white > red ≈ black > blue > brown > green ≈ yellow. The ATR-FTIR spectra of visible microplastics were identified to be polypropylene (PP), high density polyethylene (HDPE), ethylene vinyl acetate (EVA), low density polyethylene (LDPE), nitrile, polymethyl methacrylate (PMMA), latex, and polyethylene terephthalate (PETE). In the study area, recreational activities, improper waste disposal, and runoff from catchment areas are the main reasons for the contamination of microplastics in the Lake. The pollution load can be minimized by taking measures such as creating awareness, promotion of ecotourism, and reducing plastic use.

Effect of exogenous alpha-tocopherol on physio-biochemical attributes and agronomic performance of lentil (Lens culinaris Medik.) under drought stress.

Water being a vital part of cell protoplasm plays a significant role in sustaining life on earth; however, drastic changes in climatic conditions lead to limiting the availability of water and causing other environmental adversities. α-tocopherol being a powerful antioxidant, protects lipid membranes from the drastic effects of oxidative stress by deactivating singlet oxygen, reducing superoxide radicals, and terminating lipid peroxidation by reducing fatty acyl peroxy radicals under drought stress conditions. A pot experiment was conducted and two groups of lentil cultivar (Punjab-2009) were exposed to 20 and 25 days of drought induced stress by restricting the availability of water after 60th day of germination. Both of the groups were sprinkled with α-tocopherol 100, 200 and 300 mg/L. Induced water deficit stress conditions caused a pronounced decline in growth parameters including absolute growth rate (AGR), leaf area index (LAI), leaf area ratio (LAR), root shoot ratio (RSR), relative growth rate (RGR), chlorophyll a, b, total chlorophyll content, carotenoids, and soluble protein content (SPC) which were significantly enhanced by exogenously applied α-tocopherol. Moreover, a significant increase was reported in total proline content (TPC), soluble sugar content (SSC), glycine betaine (GB) content, endogenous tocopherol levels, ascorbate peroxidase (APX), catalase (CAT) peroxidase (POD) and superoxide dismutase (SOD) activities. On the contrary, exogenously applied α-tocopherol significantly reduced the concentrations of malondialdehyde (MDA) and hydrogen peroxide (H2O2). In conclusion, it was confirmed that exogenous application of α-tocopherol under drought induced stress regimes resulted in membrane protection by inhibiting lipid peroxidation, enhancing the activities of antioxidative enzymes (APX, CAT, POD, and SOD) and accumulation of osmolytes such as glycine betaine, proline and sugar. Consequently, modulating different growth, physiological and biochemical attributes.

The major outer membrane protein of Legionella pneumophila Lpg1974 shows pore-forming characteristics similar to the human mitochondrial outer membrane pore, hVDAC1.

Legionella pneumophila is an aerobic and nonspore-forming pathogenic Gram-negative bacterium of the genus Legionella. It is the causative agent of Legionnaires' disease, also known as Legionellosis. The hosts of this organism are diverse, ranging from simple water borne protozoans such as amoebae to more complex hosts such as macrophages in humans. Genome analyses have shown the presence of genes coding for eukaryotic like proteins in several Legionella species. The presence of these proteins may assist L. pneumophila in its adaptation to the eukaryotic host. We studied the characteristics of a protein (Lpg1974) of L. pneumophila that shows remarkable homologies in length of the primary sequence and for the identity/homology of many amino acids to the voltage dependent anion channel (human VDAC1, Porin 31HL) of human mitochondria. Two different forms of Lpg1974 were overexpressed in Escherichia coli and purified to homogeneity: the one containing a putative N-terminal signal sequence and one without it. Reconstituted protein containing the signal sequence formed ion-permeable pores in lipid bilayer membranes with a conductance of approximately 5.4 nS in 1 M KCl. When the predicted N-terminal signal peptide of Lpg1974 comprising an α-helical structure similar to that at the N-terminus of hVDAC1 was removed, the channels formed in reconstitution experiments had a conductance of 7.6 nS in 1 M KCl. Both Lpg1974 proteins formed pores that were voltage-dependent and anion-selective similar to the pores formed by hVDAC1. These results suggest that Lpg1974 of L. pneumophila is indeed a structural and functional homologue to hVDAC1.

Identification and characterization of smallest pore-forming protein in the cell wall of pathogenic Corynebacterium urealyticum DSM 7109.

BACKGROUND: Corynebacterium urealyticum, a pathogenic, multidrug resistant member of the mycolata, is known as causative agent of urinary tract infections although it is a bacterium of the skin flora. This pathogenic bacterium shares with the mycolata the property of having an unusual cell envelope composition and architecture, typical for the genus Corynebacterium. The cell wall of members of the mycolata contains channel-forming proteins for the uptake of solutes. RESULTS: In this study, we provide novel information on the identification and characterization of a pore-forming protein in the cell wall of C. urealyticum DSM 7109. Detergent extracts of whole C. urealyticum cultures formed in lipid bilayer membranes slightly cation-selective pores with a single-channel conductance of 1.75 nS in 1 M KCl. Experiments with different salts and non-electrolytes suggested that the cell wall pore of C. urealyticum is wide and water-filled and has a diameter of about 1.8 nm. Molecular modelling and dynamics has been performed to obtain a model of the pore. For the search of the gene coding for the cell wall pore of C. urealyticum we looked in the known genome of C. urealyticum for a similar chromosomal localization of the porin gene to known porH and porA genes of other Corynebacterium strains. Three genes are located between the genes coding for GroEL2 and polyphosphate kinase (PKK2). Two of the genes (cur_1714 and cur_1715) were expressed in different constructs in C. glutamicum ΔporAΔporH and in porin-deficient BL21 DE3 Omp8 E. coli strains. The results suggested that the gene cur_1714 codes alone for the cell wall channel. The cell wall porin of C. urealyticum termed PorACur was purified to homogeneity using different biochemical methods and had an apparent molecular mass of about 4 kDa on tricine-containing sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). CONCLUSIONS: Biophysical characterization of the purified protein (PorACur) suggested indeed that cur_1714 is the gene coding for the pore-forming protein in C. urealyticum because the protein formed in lipid bilayer experiments the same pores as the detergent extract of whole cells. The study is the first report of a cell wall channel in the pathogenic C. urealyticum.

Single Residue Acts as Gate in OccK Channels.

The OccK protein subfamily located in the outer membrane of Pseudomonas aeruginosa contains dynamic channels with several conformational states that range from open to closed forms. The molecular determinants of the OccK channels that contribute to the diverse gating have, however, remained elusive so far. Performing molecular dynamics (MD) simulations on OccK5 (OpdH) as an example, local fluctuations of loop L7 mediated by a single residue were identified that effectively gate the channel. The features of this gate residue were studied by single-channel electrophysiology and site-directed mutagenesis demonstrating that this gate residue indeed confers unique gating properties to the OccK channels. In support of these functional measurements, MD simulations highlight the correlations between the size of the side-chain belonging to the gate residue on one side and the pore size as well as the L7 flexibility on the other side.

OmpW of Caulobacter crescentus Functions as an Outer Membrane Channel for Cations.

Caulobacter crescentus is an oligotrophic bacterium that lives in dilute organic environments such as soil and freshwater. This bacterium represents an interesting model for cellular differentiation and regulation because daughter cells after division have different forms: one is motile while the other is non-motile and can adhere to surfaces. Interestingly, the known genome of C. crescentus does not contain genes predicted to code for outer membrane porins of the OmpF/C general diffusion type present in enteric bacteria or those coding for specific porins selective for classes of substrates. Instead, genes coding for 67 TonB-dependent outer membrane receptors have been identified, suggesting that active transport of specific nutrients may be the norm. Here, we report that high channel-forming activity was observed with crude outer membrane extracts of C. crescentus in lipid bilayer experiments, indicating that the outer membrane of C. crescentus contained an ion-permeable channel with a single-channel conductance of about 120 pS in 1M KCl. The channel-forming protein with an apparent molecular mass of about 20 kDa was purified to homogeneity. Partial protein sequencing of the protein indicated it was a member of the OmpW family of outer membrane proteins from Gram-negative bacteria. This channel was not observed in reconstitution experiments with crude outer membrane extracts of an OmpW deficient C. crescentus mutant. Biophysical analysis of the C. crescentus OmpW suggested that it has features that are special for general diffusion porins of Gram-negative outer membranes because it was not a wide aqueous channel. Furthermore, OmpW of C. crescentus seems to be different to known OmpW porins and has a preference for ions, in particular cations. A putative model for OmpW of C. crescentus was built on the basis of the known 3D-structures of OmpW of Escherichia coli and OprG of Pseudomonas aeruginosa using homology modeling. A comparison of the two known structures with the model of OmpW of C. crescentus suggested that it has a more hydrophilic interior and possibly a larger diameter.