Mahmut Parmaksiz

Clinical applications of decellularized extracellular matrices for tissue engineering and regenerative medicine

Decellularization is the process of removing the cellular components from tissues or organs. It is a promising technology for obtaining a biomaterial with a highly preserved extracellular matrix (ECM), which may also act as a biological scaffold for tissue engineering and regenerative therapies. Decellularized products are gaining clinical importance and market space due to their ease of standardized production, constant availability for grafting and mechanical or biochemical superiority against competing clinical options, yielding clinical results ahead of the ones with autografts in some applications. Current drawbacks and limitations of traditional treatments and clinical applications can be overcome by using decellularized or acellular matrices. Several companies are leading the market with versatile acellular products designed for diverse use in the reconstruction of tissues and organs. This review describes ECM-based decellularized and acellular products that are currently in use for different branches of clinic.

Decellularization of bovine small intestinal submucosa and its use for the healing of a critical‐sized full‐thickness skin defect, alone and in combination with stem cells, in a small rodent model

In this study, we initially described an efficient decellularization protocol for bovine‐derived small intestinal submucosa (bSIS), involving freeze–thaw cycles, acid/base treatment and alcohol and buffer systems. We compared the efficacy of our protocol to some previously established ones, based on DNA content and SEM and histochemical analyses. DNA content was reduced by ~89.4%, significantly higher than compared protocols. The sulphated GAG content of the remaining interconnected fibrous structure was 5.738 ± 0.207 µg/mg (55% retained). An in vitro study was performed to evaluate whether rat bone marrow mesenchymal stem cells (MSCs) could attach and survive on bSIS membranes. Our findings revealed that MSCs can preserve their viability and proliferate on bSIS for > 2 weeks in culture. We conducted in vivo applications for the treatment of an experimental rat model of critical sized (7 cm2) full‐thickness skin defect. The wound models treated with either MSCs‐seeded (1.5 × 106 cells/cm2) or non‐seeded bSIS membranes were completely closed by week 7 without significant differences in closure time; on the other hand, the open wound control was closed at ~47% at this time point. Immunohistopathology results revealed that the group which received MSCs‐seeded bSIS had less scarring at the end of the healing process and was in further stages of appendage formation in comparison with the non‐seeded bSIS group. Copyright

Extracellular Matrix and Regenerative Therapies from the Cardiac Perspective.

Cardiovascular diseases are the leading cause of death and a major cause of financial burden. Regenerative therapies for heart diseases bring the promise of alternative treatment modalities for myocardial infarction, ischemic heart disease, and congestive heart failure. Although, clinical trials attest to the safety of stem cell injection therapies, researchers need to overcome the underlying mechanisms that are limiting the success of future regenerative options. This article aims to review the basic scientific concepts in the field of mechanobiology and the effects of extracellular functions on stem cell fate.

Time-Resolved Fluorescence Resonance Energy Transfer [TR-FRET] Assays for Biochemical Processes

Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) is a fluorescence based technique which enables the analysis of molecular interactions in biochemical processes. Principle of TR-FRET is based on time-resolved fluorescence (TRF) measurement and fluorescence resonance energy transfer (FRET) between donor and acceptor molecules. To generate FRET signal, donor and acceptor molecules must show spectral overlap and should be in close proximity to each other and display suitable dipole orientation. The specific signal is acquired from molecules of interest via interactions of donor and acceptor molecules. TR-FRET technique is widely used for studying kinase assays, cellular signaling pathways, protein-protein interactions, DNA-protein interactions, and receptor-ligand binding. There are various propriety applications of TR-FRET. Two different sample protocols are summarized in this review.

Differential gene expression profiling of human adipose stem cells differentiating into smooth muscle-like cells by TGFβ1/BMP4

ABSTRACT Regenerative repair of the vascular system is challenging from the perspectives of translational medicine and tissue engineering. There are fundamental hurdles in front of creating bioartificial arteries, which involve recaputilation of the three‐layered structure under laboratory settings. Obtaining and maintaining smooth muscle characteristics is an important limitation, as the transdifferentiated cells fail to display mature phenotype. This study aims to shed light on the smooth muscle differentiation of human adipose stem cells (hASCs). To this end, we first acquired hASCs from lipoaspirate samples. Upon characterization, the cells were induced to differentiate into smooth muscle (SM)‐like cells using a variety of inducer combinations. Among all, TGF&bgr;1/BMP4 combination had the highest differentiation efficiency, based on immunohistochemical analyses. hSM‐like cell samples were compared to hASCs and to the positive control, human coronary artery‐smooth muscle cells (hCA‐SMCs) through gene transcription profiling. Microarray findings revealed the activation of gene groups that function in smooth muscle differentiation, signaling pathways, extracellular modeling and cell proliferation. Our results underline the effectiveness of the growth factors and suggest some potential variables for detecting the SM‐like cell characteristics. Evidence in transcriptome level was used to evaluate the TGF&bgr;1/BMP4 combination as a previously unexplored effector for the smooth muscle differentiation of adipose stem cells. HighlightsHuman adipose stem cells (hASCs) were isolated, characterized and cultured.Growth factor combinations were evaluated for their effectiveness in differentiation using IHC.hASCs were differentiated into smooth muscle (SM)‐like cells using TGF‐&bgr;1 and BMP4 combination.Microarray analysis was performed for hASCs, SM‐like cells and coronary artery‐SMCs.Microarray data was used to perform hierarchical clustering and interpretation of activated pathways.

Decellularization of Bovine Small Intestinal Submucosa

Decellularization technology promises to overcome some of the significant limitations in the regenerative medicine field by providing functional biocompatible grafts. The technique involves removal of the cells from the biological tissues or organs for further use in tissue engineering and clinical interventions. There are significant differences between decellularization protocols due to the intrinsic properties of different tissue types and purpose of use. This multistep, chemical-solution-based protocol is optimized for the preparation of decellularized bovine small intestinal submucosa (SIS).

Decellularized bSIS-ECM as a Regenerative Biomaterial for Skin Wound Repair

Tissue engineering-based regenerative applications can involve the use of stem cells for the treatment of non-healing wounds. Multipotent mesenchymal stem cells have become a focus of skin injury treatments along with many other injury types owing to their unprecedented advantages. However, there are certain limitations concerning the solo use of stem cells in skin wound repair. Natural bioactive extracellular matrix-based scaffolds have great potential for overcoming these limitations by supporting the regenerative activity and localization of stem cells. This chapter describes the use of bone marrow mesenchymal stem cells together with decellularized bovine small intestinal submucosa (SIS), for the treatment of a critical-sized full-thickness skin defect in a small animal model.

Investigations on clonazepam-loaded polymeric micelle-like nanoparticles for safe drug administration during pregnancy

Abstract Medication during pregnancy is often a necessity for women to treat their acute or chronic diseases. The goal of this study is to evaluate the potential of micelle-like nanoparticles (MNP) for providing safe drug usage in pregnancy and protect both foetus and mother from medication side effects. Clonazepam-loaded MNP were prepared from copolymers [polystyrene-poly(acrylic acid) (PS-PAA), poly(ethylene glycol)-b-poly(lactic acid) (PEG-PLA) and distearyl-sn-glycero-3-phosphoethanolamine-N-[methoxy-poly(ethylene glycol) (PEG-DSPE)] with varying monomer ratios and their drug-loading efficiency, drug release ratio, particle size, surface charge and morphology were characterised. The cellular transport and cytotoxicity experiments were conducted on clonazepam and MNP formulations using placenta-choriocarcinoma-BeWo and brain-endothelial-bEnd3 cells. Clonazepam-loaded PEG5000-PLA4500 MNP reduced the drug transport through BeWo cells demonstrating that MNP may lower foetal drug exposure, thus reduce the drug side effects. However, lipofectamine modified MNP improved the transport of clonazepam and found to be promising for brain and in-utero-specific drug treatment.