Finosh G. Thankam

Growth and survival of cells in biosynthetic poly vinyl alcohol–alginate IPN hydrogels for cardiac applications

Biosynthetic hydrogels of poly vinyl alcohol-calcium alginate were prepared as semi (semi-IPN hydrogel, PAH) and full interpenetrating polymeric network (IPN hydrogel, PAHG) for tissue engineering of cardiac tissue. The biological response of these hydrogels was studied. The IPN hydrogel exhibits amphiphilic nature and moderate equilibrium water content. The IPN hydrogel inherits the water as structured one along with more free water. The IPN hydrogel has adequate mechanical strength and fatigue life and stability in physiological media over the semi-IPN hydrogel. The structured water along with more free water in IPN hydrogel promotes blood compatibility, cell adhesion and proliferation and maintains three dimensional growths of L929 fibroblast and H9C2 cardiomyoblasts. IPN hydrogel maintain long term cell viability and infiltration with exchange of nutrients in the interstices of the hydrogel. The IPN hydrogel, PAHG is a promising material for engineering cardiac tissue.

Influence of plasma protein–hydrogel interaction moderated by absorption of water on long-term cell viability in amphiphilic biosynthetic hydrogels

Studies on the effect of plasma protein binding on biosynthetic hydrogels on the long term cell viability and infiltration onto the hydrogel scaffolds were carried out. Three hydrogels, PALG–P (PALG-co-PEGDA), PALG–PA (PALG-co-PEGDA-co-AA) and PALG–PB (PALG-co-PEGDA-co-BMA) were prepared using a copolymer of poly(propylene fumarate)-co-alginate (PALG) and cross-linker PEGDA and vinyl monomers. The nature of vinyl monomer largely influences the nature of water (structured bound water/freezing free water) present in the hydrogel and also the adsorption of protein, cell growth and infiltration. The extensively bound structured water as observed with butyl methacrylate based poly(propylene fumarate)-co-alginate-PEGDA hydrogel (PALG–PB) do not favour absorption of proteins and sustain cell growth and infiltration for long duration. Though moderately bound structured water favours absorption of protein moderately as observed with poly(propylene fumarate)-co-alginate-PEGDA hydrogel (PALG–PB), it does not sustain the cell growth. However, the minimally bound structured water favours absorption of protein extensively as observed with acrylic acid based poly(propylene fumarate)-co-alginate-PEGDA hydrogel (PALG–PA) and sustains the cell growth and infiltration for long duration.

TREM-1, HMGB1 and RAGE in the Shoulder Tendon: Dual Mechanisms for Inflammation Based on the Coincidence of Glenohumeral Arthritis

Rotator cuff injury (RCI) is a major musculoskeletal disorder in the adult population where inflammation and pain are major contributing factors. Coincidence of other clinical conditions like glenohumeral arthritis aggravates inflammation and delays the healing response. The mechanism and signaling factors underlying the sustenance of inflammation in the rotator cuff joint are largely unknown. The present article aims to elucidate the involvement of inflammatory molecule, TREM-1 (Triggering Receptors Expressed on Myeloid cells-1), and danger-associated molecular patterns (DAMPs), including high mobility group protein 1 (HMGB-1) and RAGE (receptor for advanced glycation end products), in the setting of RCI with respect to the severity of glenohumeral arthritis. Biceps tendons (15 specimens) from the shoulder and blood (11 samples) from patients with glenohumeral arthritis (Group-1, n = 4) and without glenohumeral arthritis (Group-2, n = 11) after RCI surgery were obtained for the study. Molecular and morphological alterations between the groups were compared using histology, immunofluorescence, RT-PCR and flow cytometry. MRI and histomorphology assessment revealed severe inflammation in Group-1 patients while in Group-2 ECM disorganization was prominent without any hallmarks of inflammation. A significant increase in TREM-1 expression in circulating neutrophils and monocytes was observed. Elevated levels of TREM-1, HMGB-1 and RAGE in Group-1 patients along with CD68+ and CD16+ cells confirmed DAMP-mediated inflammation. Expression of TREM-1 in the tendon of Group-2 patients even in the absence of immune cells presented a new population of TREM-expressing cells that were confirmed by real-time PCR analysis and immunofluorescence. Expression of HMGB-1 and RAGE in the biceps tendon from the shoulder of patients without glenohumeral arthritis implied TREM-1-mediated inflammation without involving immune cells, whereas in patients with glenohumeral arthritis, infiltration and the activation of the immune cells, primarily macrophages, release mediators to induce inflammation. This could be the reason for ECM disorganization without the classical signs of inflammation in patients without glenohumeral arthritis.

Influence of physical and mechanical properties of amphiphilic biosynthetic hydrogels on long-term cell viability.

Maintaining the mechanical properties of biofunctional hydrogels of natural resources for tissue engineering and biomedical applications for an intended period of duration is a challenge. Though anionic polysaccharide alginate has been hailed for its excellent biomimetic characters for tissue engineering, it usually fails in load bearing and other dynamic mechanical environment. In this paper this issue was addressed by copolymerizing alginate with the biocompatible and mechanically robust synthetic biodegradable polyester and crosslinking with polyethylene glycol diacrylate (PEGDA) and vinyl co-monomers, 2-hydroxy ethyl methacrylate (HEMA), methyl methacrylate (MMA) and N N׳ methylene bis acrylamide (NMBA) to form three hydrogels. All three hydrogels were amphiphilic, hemocompatible and non-cytotoxic. These hydrogels exhibited appreciable water holding capacity. Comparatively, hydrogel prepared with PEGDA-NMBA crosslinkers displayed larger pore size, increased crosslinking, higher tensile strength and controlled degradation. With appreciable swelling and EWC, this hydrogel elicited better biological responses with long-term cell viability for cardiac tissue engineering.

Biosynthetic hydrogels—Studies on chemical and physical characteristics on long-term cellular response for tissue engineering

Biosynthetic hydrogels can meet the drawbacks caused by natural and synthetic ones for biomedical applications. In the current article we present a novel biosynthetic alginate-poly(propylene fumarate) copolymer based chemically crosslinked hydrogel scaffolds for cardiac tissue engineering applications. Partially crosslinked PA hydrogel and fully cross linked PA-A hydrogel scaffolds were prepared. The influence of chemical and physical (morphology and architecture of hydrogel) characteristics on the long term cellular response was studied. Both these hydrogels were cytocompatible and showed no genotoxicity upon contact with fibroblast cells. Both PA and PA-A were able to resist deleterious effects of reactive oxygen species and sustain the viability of L929 cells. The hydrogel incubated oxidative stress induced cells were capable of maintaining the intra cellular reduced glutathione (GSH) expression to the normal level confirmed their protective effect. Relatively the PA hydrogel was found to be unstable in the cell culture medium. The PA-A hydrogel was able to withstand appreciable cyclic stretching. The cyclic stretching introduced complex macro and microarchitectural features with interconnected pores and more structured bound water which would provide long-term viability of around 250% after the 24th day of culture. All these qualities make PA-A hydrogel form a potent candidate for cardiac tissue engineering.

Immunobiological factors aggravating the fatty infiltration on tendons and muscles in rotator cuff lesions

Rotator cuff lesions (RCLs) are a common cause of shoulder pain and dysfunction. The rotator cuff tendons can degenerate and/or tear from the greater tuberosity of the humerus, which is associated with several anatomical, physiological, biochemical, and molecular changes in tendon and muscle. In this article, these pathways are critically reviewed and discussed with various management strategies of RCLs. The article also highlights the immunobiological responses following the RCL and the inherent repair mechanisms elicited by the body. The greatest difficulty in treating this pathology is that the muscle can undergo irreversible fatty infiltration in the setting of chronic tears that is associated with poor surgical outcomes. The article also investigates the key molecular pathways of the muscle homeostasis (mTOR, Rho kinase, AMPK, and Ca2+) with the energy metabolism to propose a possible mechanism for fatty infiltration. Future research is warranted to target the key players of these pathways in the management of fatty infiltration and thus RCL.

Triggering receptor expressed on myeloid cells and 5’adenosine monophosphate-activated protein kinase in the inflammatory response: a potential therapeutic target

ABSTRACT Introduction: The events in the cellular and molecular signaling triggered during inflammation mitigate tissue healing. The metabolic check-point control mediated by 5ʹ-adenosine monophosphate-activated protein kinase (AMPK) is crucial for switching the cells into an activated state capable of mediating inflammatory events. The cell metabolism involved in the inflammatory response represents a potential therapeutic target for the pharmacologic management of inflammation. Areas covered: In this article, a critical review is presented on triggering receptor expressed on myeloid cell (TREM) receptors and their role in the inflammatory responses, as well as homeostasis between different TREM molecules and their regulation. Additionally, we discussed the relationship between TREM and AMPK to identify novel targets to limit the inflammatory response. Literature search was carried out from the National Library of Medicine’s Medline database (using PubMed as the search engine) and Google Scholar and identified relevant studies up to 30 March 2016 using inflammation, TREM, AMPK, as the key words. Expert commentary: The prevention of phenotype switching of immune cells during inflammation by targeting AMPK and TREM-1 could be beneficial for developing novel management strategies for inflammation and associated complications.

Influence of matrix and bulk behaviour of an injectable hydrogel on the survival of encapsulated cardiac cells

Cytocompatibility, suitable porosity, higher equilibrium water content and tissue like elasticity are the demanding criteria required to design a hydrogel for cell encapsulation and delivery. Here a mechanically stable cell supporting synthetic hydrogel was fabricated from poly(propylene fumarate-co-ethylene glycol)/PEGDA by redox initiating polymerisation for cell encapsulation. A hydrogel prepared with 93.5% poly(propylene fumarate-co-ethylene glycol) and 6.5% PEGDA acquired matrix and bulk characteristics of equilibrium water content (EWC) 84.45 ± 0.80%, freezable water content 67.93%, Young modulus 212.2 ± 0.02 kPa and pore diameter 88.64 ± 18.96 μm. This hydrogel with higher free water content, favourable pore dimensions and mechanical strength was used to encapsulate cardiomyoblasts. The encapsulated cardiomyoblasts were showing increasing viability from 3–30 days with viable green fluorescence. The matrix and bulk characteristics of the hydrogel are favourable and elicited uniform, green fluorescing, live cardiomyoblasts (H9c2) inside with 150% cell viability (MTT assay) and uniform ECM protein distribution after 30 days. The slow in vitro degradation of the hydrogel in physiological-like conditions is favourable for the delivery and retention of the encapsulated cells at the injection site.

Infiltration and sustenance of viability of cells by amphiphilic biosynthetic biodegradable hydrogels.

Amphiphilic biosynthetic hydrogels comprising natural polysaccharide alginate (I) and synthetic polyester polypropylene fumarate (II) units were prepared by crosslinking the copolymer of I and II with calcium ion and vinyl monomers viz, 2-hydroxyethyl methacrylate (HEMA), methyl methacrylate (MMA), butyl methacrylate (BMA) and N,N′-methylene bisacrylamide (NMBA). Three fast degradable hydrogels, ALPF-MMA, ALPF-HEMA and ALPF-BMA and one slow degradable hydrogel ALPF-NMBA were prepared. These hydrogels are amphiphilic and able to hold sufficient amount of proteins on their surfaces. All these hydrogels are found to be hemocompatible, cytocompatible and genocompatible. ALPF-NMBA promotes infiltration of L929 fibroblasts and 3D growth of H9c2 cardiomyoblasts and long-term viability.

MicroRNAs Associated with Shoulder Tendon Matrisome Disorganization in Glenohumeral Arthritis.

The extracellular matrix (ECM) provides core support which is essential for the cell and tissue architectural development. The role of ECM in many pathological conditions has been well established and ECM-related abnormalities leading to serious consequences have been identified. Though much has been explored in regards to the role of ECM in soft tissue associated pathologies, very little is known about its role in inflammatory disorders in tendon. In this study, we performed microRNA (miRNA) expression analysis in the long head of the human shoulder biceps tendon to identify key genes whose expression was altered during inflammation in patients with glenohumeral arthritis. We identified differential regulation of matrix metalloproteinases (MMPs) that could be critical in collagen type replacement during tendinopathy. The miRNA profiling showed consistent results between the groups and revealed significant changes in the expression of seven different miRNAs in the inflamed tendons. Interestingly, all of these seven miRNAs were previously reported to have either a direct or indirect role in regulating the ECM organization in other pathological disorders. In addition, these miRNAs were also found to alter the expression levels of MMPs, which are the key matrix degrading enzymes associated with ECM-related abnormalities and pathologies. To our knowledge, this is the first report which identifies specific miRNAs associated with inflammation and the matrix reorganization in the tendons. Furthermore, the findings also support the potential role of these miRNAs in altering the collagen type ratio in the tendons during inflammation which is accompanied with differential expression of MMPs.