Controlled Delivery of Pan-PAD-Inhibitor Cl-Amidine Using Poly(3-Hydroxybutyrate) Microspheres.

This study deals with the process of optimization and synthesis of Poly(3-hydroxybutyrate) microspheres with encapsulated Cl-amidine. Cl-amidine is an inhibitor of peptidylarginine deiminases (PADs), a group of calcium-dependent enzymes, which play critical roles in a number of pathologies, including autoimmune and neurodegenerative diseases, as well as cancer. While Cl-amidine application has been assessed in a number of in vitro and in vivo models; methods of controlled release delivery remain to be investigated. P(3HB) microspheres have proven to be an effective delivery system for several compounds applied in antimicrobial, wound healing, cancer, and cardiovascular and regenerative disease models. In the current study, P(3HB) microspheres with encapsulated Cl-amidine were produced in a size ranging from ~4-5 µm and characterized for surface morphology, porosity, hydrophobicity and protein adsorption, in comparison with empty P(3HB) microspheres. Cl-amidine encapsulation in P(3HB) microspheres was optimized, and these were found to be less hydrophobic, compared with the empty microspheres, and subsequently adsorbed a lower amount of protein on their surface. The release kinetics of Cl-amidine from the microspheres were assessed in vitro and expressed as a function of encapsulation efficiency. There was a burst release of ~50% Cl-amidine in the first 24 h and a zero order release from that point up to 16 days, at which time point ~93% of the drug had been released. As Cl-amidine has been associated with anti-cancer effects, the Cl-amidine encapsulated microspheres were assessed for the inhibition of vascular endothelial growth factor (VEGF) expression in the mammalian breast cancer cell line SK-BR-3, including in the presence of the anti-proliferative drug rapamycin. The cytotoxicity of the combinatorial effect of rapamycin with Cl-amidine encapsulated P(3HB) microspheres was found to be 3.5% more effective within a 24 h period. The cells treated with Cl-amidine encapsulated microspheres alone, were found to have 36.5% reduction in VEGF expression when compared with untreated SK-BR-3 cells. This indicates that controlled release of Cl-amidine from P(3HB) microspheres may be effective in anti-cancer treatment, including in synergy with chemotherapeutic agents. Using controlled drug-delivery of Cl-amidine encapsulated in Poly(3-hydroxybutyrate) microspheres may be a promising novel strategy for application in PAD-associated pathologies

Enhanced production of biobased, biodegradable, Poly(3-hydroxybutyrate) using an unexplored marine bacterium Pseudohalocynthiibacter aestuariivivens, isolated from highly polluted coastal environment.

The production and disposal of plastics from limited fossil reserves, has prompted research for greener and sustainable alternatives. Polyhydroxyalkanoates (PHAs) are biocompatible, biodegradable, and thermoprocessable polyester produced by microbes. PHAs found several applications but their use is limited due to high production cost and low yields. Herein, for the first time, the isolation and characterization of Pseudohalocynthiibacter aestuariivivens P96, a marine bacterium able to produce surprising amount of PHAs is reported. In the best growth condition P96 was able to reach a maximum production of 4.73 g/L, corresponding to the 87 % of total cell dry-weight. Using scanning and transmission microscopy, lab-scale fermentation, spectroscopic techniques, and genome analysis, the production of thermoprocessable polymer Polyhydroxybutyrate P(3HB), a PHAs class, endowed with mechanical and thermal properties comparable to that of petroleum-based plastics was confirmed. This study represents a milestone toward the use of this unexplored marine bacterium for P(3HB) production. [Abstract copyright: Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Preclinical study of peripheral nerve regeneration using nerve guidance conduits based on polyhydroxyalkanaotes

Nerve guidance conduits (NGCs) are used as an alternative to the “gold standard” nerve autografting, preventing the need for surgical intervention required to harvest autologous nerves. However, the regeneration outcomes achieved with the current NGCs are only comparable with autografting when the gap is short (less than 10 mm). In the present study, we have developed NGCs made from a blend of Polyhydroxyalkanoates, a family of natural resorbable polymers. Hollow NGCs made from a 75:25 Poly(3‐hydroxyoctanoate)/Poly(3‐hydroxybutyrate) blend (PHA‐NGCs) were manufactured using dip‐moulding. These PHA‐NGCs showed appropriate flexibility for peripheral nerve regeneration. In vitro cell studies performed using RT4‐D6P2T rat Schwann cell line confirmed that the material is capable of sustaining cell proliferation and adhesion. PHA‐NGCs were then implanted in vivo to repair 10 mm gaps of the median nerve of female Wistar rats for 12 weeks. Functional evaluation of the regenerated nerve using the grasping test showed that PHA‐NGCs displayed similar motor recovery as the autograft, starting from week 7. Additionally, nerve cross‐sectional area, density and number of myelinated cells, as well as axon diameter, fibre diameter, myelin thickness and g‐ratio obtained using the PHA‐NGCs were found comparable to an autograft. This preclinical data confirmed that the PHA‐NGCs are indeed highly promising candidates for peripheral nerve regeneration

Dual production of polyhydroxyalkanoates and antibacterial/antiviral gold nanoparticles

Gold nanoparticles (AuNPs) have been explored for their use in medicine. Here, we report a sustainable, and cost-effective method to produce AuNPs using a bacterial strain such as Pseudomonas mendocina CH50 which is also known to be a polyhydroxyalkanoate (PHA) producer. A cell-free bacterial supernatant, which is typically discarded after PHA extraction, was used to produce spherical AuNPs of 3.5 ± 1.5 nm in size as determined by Transmission Electron Microscopy (TEM) analysis. The AuNPs/PHA composite coating demonstrated antibacterial activity against Staphylococcus aureus 6538P, and antiviral activity, with a 75% reduction in viral infectivity against SARS-CoV-2 pseudotype virus

Antibacterial Composite Materials Based on the Combination of Polyhydroxyalkanoates With Selenium and Strontium Co-substituted Hydroxyapatite for Bone Regeneration

Due to the threat posed by the rapid growth in the resistance of microbial species to antibiotics, there is an urgent need to develop novel materials for biomedical applications capable of providing antibacterial properties without the use of such drugs. Bone healing represents one of the applications with the highest risk of postoperative infections, with potential serious complications in case of bacterial contaminations. Therefore, tissue engineering approaches aiming at the regeneration of bone tissue should be based on the use of materials possessing antibacterial properties alongside with biological and functional characteristics. In this study, we investigated the combination of polyhydroxyalkanoates (PHAs) with a novel antimicrobial hydroxyapatite (HA) containing selenium and strontium. Strontium was chosen for its well-known osteoinductive properties, while selenium is an emerging element investigated for its multi-functional activity as an antimicrobial and anticancer agent. Successful incorporation of such ions in the HA structure was obtained. Antibacterial activity against Staphylococcus aureus 6538P and Escherichia coli 8739 was confirmed for co-substituted HA in the powder form. Polymer-matrix composites based on two types of PHAs, P(3HB) and P(3HO-co-3HD-co-3HDD), were prepared by the incorporation of the developed antibacterial HA. An in-depth characterization of the composite materials was conducted to evaluate the effect of the filler on the physicochemical, thermal, and mechanical properties of the films. In vitro antibacterial testing showed that the composite samples induce a high reduction of the number of S. aureus 6538P and E. coli 8739 bacterial cells cultured on the surface of the materials. The films are also capable of releasing active ions which inhibited the growth of both Gram-positive and Gram-negative bacteria

Nanofibrous poly(3-hydroxybutyrate)/poly(3-hydroxyoctanoate) scaffolds provide a functional microenvironment for cartilage repair

Articular cartilage defects, when repaired ineffectively, often lead to further deterioration of the tissue, secondary osteoarthritis and, ultimately, joint replacement. Unfortunately, current surgical procedures are unable to restore normal cartilage function. Tissue engineering of cartilage provides promising strategies for the regeneration of damaged articular cartilage. As yet, there are still significant challenges that need to be overcome to match the long-term mechanical stability and durability of native cartilage. Using electrospinning of different blends of biodegradable poly(3-hydroxybutyrate)/poly(3-hydroxyoctanoate), we produced polymer scaffolds and optimised their structure, stiffness, degradation rates and biocompatibility. Scaffolds with a poly(3-hydroxybutyrate)/poly(3-hydroxyoctanoate) ratio of 1:0.25 exhibit randomly oriented fibres that closely mimic the collagen fibrillar meshwork of native cartilage and match the stiffness of native articular cartilage. Degradation of the scaffolds into products that could be easily removed from the body was indicated by changes in fibre structure, loss of molecular weight and a decrease in scaffold stiffness after one and four months. Histological and immunohistochemical analysis after three weeks of culture with human articular chondrocytes revealed a hyaline-like cartilage matrix. The ability to fine tune the ultrastructure and mechanical properties using different blends of poly(3-hydroxybutyrate)/poly(3-hydroxyoctanoate) allows to produce a cartilage repair kit for clinical use to reduce the risk of developing secondary osteoarthritis. We further suggest the development of a toolbox with tailor-made scaffolds for the repair of other tissues that require a ‘guiding’ structure to support the body’s self-healing process

Natural Biomaterials for Cardiac Tissue Engineering: A Highly Biocompatible Solution.

Cardiovascular diseases (CVD) constitute a major fraction of the current major global diseases and lead to about 30% of the deaths, i.e., 17.9 million deaths per year. CVD include coronary artery disease (CAD), myocardial infarction (MI), arrhythmias, heart failure, heart valve diseases, congenital heart disease, and cardiomyopathy. Cardiac Tissue Engineering (CTE) aims to address these conditions, the overall goal being the efficient regeneration of diseased cardiac tissue using an ideal combination of biomaterials and cells. Various cells have thus far been utilized in pre-clinical studies for CTE. These include adult stem cell populations (mesenchymal stem cells) and pluripotent stem cells (including autologous human induced pluripotent stem cells or allogenic human embryonic stem cells) with the latter undergoing differentiation to form functional cardiac cells. The ideal biomaterial for cardiac tissue engineering needs to have suitable material properties with the ability to support efficient attachment, growth, and differentiation of the cardiac cells, leading to the formation of functional cardiac tissue. In this review, we have focused on the use of biomaterials of natural origin for CTE. Natural biomaterials are generally known to be highly biocompatible and in addition are sustainable in nature. We have focused on those that have been widely explored in CTE and describe the original work and the current state of art. These include fibrinogen (in the context of Engineered Heart Tissue, EHT), collagen, alginate, silk, and Polyhydroxyalkanoates (PHAs). Amongst these, fibrinogen, collagen, alginate, and silk are isolated from natural sources whereas PHAs are produced via bacterial fermentation. Overall, these biomaterials have proven to be highly promising, displaying robust biocompatibility and, when combined with cells, an ability to enhance post-MI cardiac function in pre-clinical models. As such, CTE has great potential for future clinical solutions and hence can lead to a considerable reduction in mortality rates due to CVD

Dual production of polyhydroxyalkanoates and antibacterial/antiviral gold nanoparticles

Gold nanoparticles (AuNPs) have been explored for their use in medicine. Here, we report a sustainable, and cost-effective method to produce AuNPs using a bacterial strain such as Pseudomonas mendocina CH50 which is also known to be a polyhydroxyalkanoate (PHA) producer. A cell-free bacterial supernatant, which is typically discarded after PHA extraction, was used to produce spherical AuNPs of 3.5 ± 1.5 nm in size as determined by Transmission Electron Microscopy (TEM) analysis. The AuNPs/PHA composite coating demonstrated antibacterial activity against Staphylococcus aureus 6538P, and antiviral activity, with a 75% reduction in viral infectivity against SARS-CoV-2 pseudotype virus

Biosynthesis and characterization of a novel, biocompatible medium chain length polyhydroxyalkanoate by Pseudomonas mendocina CH50 using coconut oil as the carbon source

This study validated the utilization of triacylglycerides (TAGs) by Pseudomonas mendocina CH50, a wild type strain, resulting in the production of novel mcl-PHAs with unique physical properties. A PHA yield of 58% dcw was obtained using 20g/L of coconut oil. Chemical and structural characterisation confirmed that the mcl-PHA produced was a terpolymer comprising of three different repeating monomer units, 3-hydroxyoctanoate, 3-hydroxydecanoate and 3-hydroxydodecanoate or P(3HO-3HD-3HDD). Bearing in mind the potential of P(3HO-3HD-3HDD) in biomedical research, especially in neural tissue engineering, in vitro biocompatibility studies were carried out using NG108-15 (neuronal) cells. Cell viability data confirmed that P(3HO-3HD-3HDD) supported the attachment and proliferation of NG108-15 and was therefore, confirmed to be biocompatible in nature and suitable for neural regeneration

"Coverage of Northeast India in the Indian Mainstream Media "

The research is on Coverage of Northeast India in the Indian Mainstream Media: A Study of the Perception of Northeast Indians Living in Bangalore. Northeast 'refers to the eastern most region of India consisting of Arunachal Pradesh, Assam, Manipur, Meghalaya, Mizoram, Nagaland, Tripura and Sikkim. By media the focus here is not just on the printed press but also on news channels. Mainstream media refers to national newspapers and news channels in either Hindi or English language that circulates or is available for viewing across the country. This study used a quantitative method and data was collected with the help of the research tool, questionnaire. The study was conducted in Bangalore in the year 2009 - 2010. Since this is a public perception of Northeast people residing in Bangalore on the coverage of Northeast India, it is subjective with respect to people's opinion.The motivation to conduct this research came from a viewable communication gap about the Northeast public in the mainstream or the national media. Irrespective of the varied socio-politico-economic dynamics of all northeastern states, this is one of the common problems faced by each of the northeastern states. The hypothesis for this paper was media is not successful in giving the right picture of Northeast India to the rest of the country thereby making people from the Northeast unsatisfied with the amount of media coverage or the kind of media coverage they receive