Cardiolipin clustering promotes mitochondrial membrane dynamics.

Cardiolipin (CL) is a mitochondria-specific phospholipid that forms heterotypic interactions with membrane-shaping proteins and regulates the dynamic remodeling and function of mitochondria. However, the precise mechanisms through which CL influences mitochondrial morphology are not well understood. In this study, employing molecular dynamics (MD) simulations, we observed CL localize near the membrane-binding sites of the mitochondrial fusion protein Optic Atrophy 1 (OPA1). To validate these findings experimentally, we developed a bromine-labeled CL probe to enhance cryoEM contrast and characterize the structure of OPA1 assemblies bound to the CL-brominated lipid bilayers. Our images provide direct evidence of interactions between CL and two conserved motifs within the paddle domain (PD) of OPA1, which control membrane-shaping mechanisms. We further observed a decrease in membrane remodeling activity for OPA1 in lipid compositions with increasing concentrations of monolyso-cardiolipin (MLCL). Suggesting that the partial replacement of CL by MLCL accumulation, as observed in Barth syndrome-associated mutations of the tafazzin phospholipid transacylase, compromises the stability of protein-membrane interactions. Our analyses provide insights into how biological membranes regulate the mechanisms governing mitochondrial homeostasis.

Fatty acyl-coenzyme A activates mitochondrial division through oligomerization of MiD49 and MiD51.

Mitochondrial fission occurs in many cellular processes, but the regulation of fission is poorly understood. We show that long-chain acyl-coenzyme A (LCACA) activates two related mitochondrial fission proteins, MiD49 and MiD51, by inducing their oligomerization, which activates their ability to stimulate the DRP1 GTPase. The 1:1 stoichiometry of LCACA:MiD in the oligomer suggests interaction in the previously identified nucleotide-binding pocket, and a point mutation in this pocket reduces LCACA binding and LCACA-induced oligomerization for MiD51. In cells, this LCACA binding mutant does not assemble into puncta on mitochondria or rescue MiD49/51 knockdown effects on mitochondrial length and DRP1 recruitment. Furthermore, cellular treatment with BSA-bound oleic acid, which causes increased LCACA, promotes mitochondrial fission in an MiD49/51-dependent manner. These results suggest that LCACA is an endogenous ligand for MiDs, inducing mitochondrial fission and providing a potential mechanism for fatty-acid-induced mitochondrial division. Finally, MiD49 or MiD51 oligomers synergize with Mff, but not with actin filaments, in DRP1 activation, suggesting distinct pathways for DRP1 activation.

Effects of liquid-to-solid ratio and gamma irradiation on the rheology and cytocompatibility of a beta-tricalcium phosphate-based injectable bone substitute.

Injectable bone substitute (IBS) materials are commonly used to fill irregular-shaped bone voids in non-load-bearing areas and can offer greater utility over those which are in prefabricated powder, granule, or block forms. This work investigates the impact of liquid-to-solid ratio (LSR) on the rheology and cytocompatibility of IBSs formulated from bioactive glass particles and ß-tricalcium phosphate (ß-TCP) in glycerol and poly(ethylene glycol) (PEG). IBS formulations of varying LSR were prepared and packed in 3 cc open-bore syringes and sterilized via gamma irradiation (10 kGy, 25 kGy). Gamma-irradiated formulations with high PEG content required the highest (73 N) mechanical force for injection from syringes. Oscillatory viscosity measurements revealed that the viscosity of samples was directly proportional to glycerol content. PEG and glycerol displayed competing effects on the washout resistance and cohesiveness of samples, which were based on total weight loss in media and Ca2+ ion release, respectively. Cell viability in 24-h extracts of 10 kGy gamma-sterilized and 25 kGy gamma-irradiated samples were 22.94% and 56.53%, respectively. The research highlights the complex interplay of IBS components on IBS rheology and, moreover, the cytotoxicity behaviors of beta-tricalcium phosphate-based injectable bone substitutes by in vitro experiments.

Extracellular Matrix-Based Conductive Composites for Myocardial Tissue Regeneration.

Myocardial tissue engineering strategies such as fabrication of cardiac patches for tissue regeneration offer various solutions for the loss of function developed due to myocardial infarction. Here, we combined the hybrid structure (previously obtained and combined decellularized myocardium grafts with poly(glycerol-sebacate) polymer) with multiwalled carbon nanotubes (MWCNTs) to provide the essential characteristics for cardiac tissue regeneration. MWCNTs were doped in the cross-linked structure, and the conductivity and Young's modulus of the composite elastomer were found as 5 × 10-3 ± 1 × 10-3 S/m and 374 ± 75.8 kPa, respectively. The cell-material interaction was evaluated, and composite structures supported cell adhesion and showed no cytotoxic effect.

Structural mechanism of mitochondrial membrane remodelling by human OPA1.

Distinct morphologies of the mitochondrial network support divergent metabolic and regulatory processes that determine cell function and fate1-3. The mechanochemical GTPase optic atrophy 1 (OPA1) influences the architecture of cristae and catalyses the fusion of the mitochondrial inner membrane4,5. Despite its fundamental importance, the molecular mechanisms by which OPA1 modulates mitochondrial morphology are unclear. Here, using a combination of cellular and structural analyses, we illuminate the molecular mechanisms that are key to OPA1-dependent membrane remodelling and fusion. Human OPA1 embeds itself into cardiolipin-containing membranes through a lipid-binding paddle domain. A conserved loop within the paddle domain inserts deeply into the bilayer, further stabilizing the interactions with cardiolipin-enriched membranes. OPA1 dimerization through the paddle domain promotes the helical assembly of a flexible OPA1 lattice on the membrane, which drives mitochondrial fusion in cells. Moreover, the membrane-bending OPA1 oligomer undergoes conformational changes that pull the membrane-inserting loop out of the outer leaflet and contribute to the mechanics of membrane remodelling. Our findings provide a structural framework for understanding how human OPA1 shapes mitochondrial morphology and show us how human disease mutations compromise OPA1 functions.

The Co-Use of Stromal Vascular Fraction and Bone Marrow Concentrate for Tendon Healing.

OBJECTIVE: The Achilles tendon is the most frequently injured tendon in the human body, despite being the strongest. Many conventional treatments including medication, surgical interventions, and physical therapy are available, however, the desired results are often not achieved. Stromal vascular fraction (SVF) and bone marrow concentrate (BMC) are two additional cellular treatment options. The purpose of this study is to evaluate the effect of SVF and BMC, used as a combination, for the treatment of Achilles tendon injuries. METHODS: Five male New Zealand rabbits were used for each of the 6 study groups. A 3-mm of SVF and BMC were injected on the Achilles tendons at certain ratios. The histological results were classified by the Movin grading system for tendon healing. The collagen type-I and type-III structures in the tendons were examined by immunohistochemical evaluation. The expressions of tendon-specific genes were also examined by using the RT-PCR method to analyze tendon healing. RESULTS: Histological and immunohistochemical evaluation indicated that tendons receiving the SVF and BMAC mixture performed better than control and individual groups (p < 0.05). Moreover, RT-PCR evaluation showed that mixture-receiving groups were the closest similar to the uninjured group (p < 0.05). CONCLUSION: The combined use of BMC and SVF improved Achilles tendon healing when compared to the individual use of each mixture.

Preparation of hybrid meniscal constructs using hydrogels and acellular matrices.

To search for a suitable meniscus repair material, acellular hybrid scaffolds consisting of in situ cross-linkable 3-D interpenetrating network structures were obtained by decellularization of the meniscus tissues followed by integration of the gel system. Decellularization efficiency was confirmed using a DNA quantification assay (82% decrease in DNA content) and histological stainings. In the second part of the study, the gelatin molecule was functionalized by adding methacrylic anhydride and the degree of functionalization was found to be 75% by (Proton-Nuclear Magnetic Resonance) 1H-NMR. Using this, a series of hybrid constructs named GelMA-Hybrid (G-Hybrid), GELMA/PEGDMA-Hybrid (PG-Hybrid), and GelMA/PEGDMA/HAMA-Hybrid (PGH-Hybrid) were prepared by cross-linking with UVA. Changes in the chemical structure were determined with Fourier Transform Infrared Spectrophotometer (FTIR). Water uptake capacities of cross-linked hybrid structures were measured in swelling studies, and it was found that hybrid scaffolds showed similar swelling properties compared to native counterparts. By compressive mechanical tests, enhanced mechanical properties were revealed in cross-linked scaffolds with PGH-Hybrid having the highest cross-link density. Protein denaturation and decomposition transition temperatures were improved by adding hydrogels to acellular scaffolds according to thermal gravimetric analyses (TGA). Cross-linked acellular scaffolds have exhibited a behavior close to native tissues with below 25% mass loss in phosphate buffer saline (PBS) and enzymatic solution. Cell viability was examined through Alamar Blue on the first day and cell viability in hybrid constructs was found to be above 80% while it was closer to the control group on the 7th day. It was concluded that the developed biomaterials could be used in meniscus tissue engineering with their tunable physicochemical and mechanical properties.

The structure of the human LACTB filament reveals the mechanisms of assembly and membrane binding.

Mitochondria are complex organelles that play a central role in metabolism. Dynamic membrane-associated processes regulate mitochondrial morphology and bioenergetics in response to cellular demand. In tumor cells, metabolic reprogramming requires active mitochondrial metabolism for providing key metabolites and building blocks for tumor growth and rapid proliferation. To counter this, the mitochondrial serine beta-lactamase-like protein (LACTB) alters mitochondrial lipid metabolism and potently inhibits the proliferation of a variety of tumor cells. Mammalian LACTB is localized in the mitochondrial intermembrane space (IMS), where it assembles into filaments to regulate the efficiency of essential metabolic processes. However, the structural basis of LACTB polymerization and regulation remains incompletely understood. Here, we describe how human LACTB self-assembles into micron-scale filaments that increase their catalytic activity. The electron cryo-microscopy (cryoEM) structure defines the mechanism of assembly and reveals how highly ordered filament bundles stabilize the active state of the enzyme. We identify and characterize residues that are located at the filament-forming interface and further show that mutations that disrupt filamentation reduce enzyme activity. Furthermore, our results provide evidence that LACTB filaments can bind lipid membranes. These data reveal the detailed molecular organization and polymerization-based regulation of human LACTB and provide new insights into the mechanism of mitochondrial membrane organization that modulates lipid metabolism.

Three-Country Snapshot of Ornithine Transcarbamylase Deficiency.

X-linked ornithine transcarbamylase deficiency (OTCD) is the most common urea cycle defect. The disease severity ranges from asymptomatic carrier state to severe neonatal presentation with hyperammonaemic encephalopathy. We audited the diagnosis and management of OTCD, using an online 12-question-survey that was sent to 75 metabolic centres in Turkey, France and the UK. Thirty-nine centres responded and 495 patients were reported in total. A total of 208 French patients were reported, including 71 (34%) males, 86 (41%) symptomatic and 51 (25%) asymptomatic females. Eighty-five Turkish patients included 32 (38%) males, 39 (46%) symptomatic and 14 (16%) asymptomatic females. Out of the 202 UK patients, 66 (33%) were male, 83 (41%) asymptomatic and 53 (26%) symptomatic females. A total of 19%, 12% and 7% of the patients presented with a neonatal-onset phenotype in France, Turkey and the UK, respectively. Vomiting, altered mental status and encephalopathy were the most common initial symptoms in all three countries. While 69% in France and 79% in Turkey were receiving protein restriction, 42% were on a protein-restricted diet in the UK. A total of 76%, 47% and 33% of patients were treated with ammonia scavengers in Turkey, France and the UK, respectively. The findings of our audit emphasize the differences and similarities in manifestations and management practices in three countries.

Chronic enteropathy associated with SLCO2A1 gene and hereditary fructose intolerance: A coincidence of two rare diseases.

Chronic enteropathy associated with SLCO2A1 gene (CEAS) is a rare disorder characterized by multiple small intestine ulcers. Patients with CEAS typically present with chronic anemia and gastrointestinal bleeding. Besides CEAS, SLCO2A1 mutations cause primary hypertrophic osteoarthropathy (PHO) which is considered as an extraintestinal manifestation in CEAS patients. Since CEAS and Crohn's disease are clinically indistinguishable, patients are often misdiagnosed with Crohn's disease. Herein, we describe a 4-year-old Turkish girl with CEAS due to homozygous pathogenic variant (c.656C > T) in SLCO2A1 with concomitant hereditary fructose intolerance (HFI) caused by homozygous pathogenic variant (c.1005C > G) in ALDOB. Prompt restriction of fructose, sucrose and sorbitol resulted in hepatomegaly regression and mild amelioration of patient's symptoms. Despite budesonide and azathioprine treatments, patient's protein losing enteropathy and chronic anemia did not improve. Although previous CEAS cases were reported from East Asian countries, it is likely to occur in people from other geographic areas. CEAS seems to be underdiagnosed and high index of suspicion is required for the diagnosis of this rare entity. Patients with prior diagnosis of Crohn's disease with no response to immunosuppressive treatment or anti-TNF therapy should be re-evaluated for possible CEAS diagnosis.

Structure and functionality of a multimeric human COQ7:COQ9 complex.

Coenzyme Q (CoQ) is a redox-active lipid essential for core metabolic pathways and antioxidant defense. CoQ is synthesized upon the mitochondrial inner membrane by an ill-defined "complex Q" metabolon. Here, we present structure-function analyses of a lipid-, substrate-, and NADH-bound complex comprising two complex Q subunits: the hydroxylase COQ7 and the lipid-binding protein COQ9. We reveal that COQ7 adopts a ferritin-like fold with a hydrophobic channel whose substrate-binding capacity is enhanced by COQ9. Using molecular dynamics, we further show that two COQ7:COQ9 heterodimers form a curved tetramer that deforms the membrane, potentially opening a pathway for the CoQ intermediates to translocate from the bilayer to the proteins' lipid-binding sites. Two such tetramers assemble into a soluble octamer with a pseudo-bilayer of lipids captured within. Together, these observations indicate that COQ7 and COQ9 cooperate to access hydrophobic precursors within the membrane and coordinate subsequent synthesis steps toward producing CoQ.

Development of Antithrombogenic ECM-Based Nanocomposite Heart Valve Leaflets.

Thrombogenicity, which is commonly encountered in artificial heart valves after replacement surgeries, causes valvular failure. Even life-long anticoagulant drug use may not be sufficient to prevent thrombogenicity. In this study, it was aimed to develop a heart valve construct with antithrombogenic properties and suitable mechanical strength by combining multiwalled carbon nanotubes within a decellularized bovine pericardium. In this context, the decellularization process was performed by using the combination of freeze-thawing and sodium dodecyl sulfate (SDS). Evaluation of decellularization efficiency was determined by histology (Hematoxylin and Eosin, DAPI and Masson's Trichrome) and biochemical (DNA, sGAG and collagen) analyses. After the decellularization process of the bovine pericardium, composite pericardial tissues were prepared by incorporating -COOH-modified multiwalled carbon nanotubes (MWCNTs). Characterization of MWCNT incorporation was performed by ATR-FTIR, TGA, and mechanical analysis, while SEM and AFM were used for morphological evaluations. Thrombogenicity assessments were studied by platelet adhesion test, Calcein-AM staining, kinetic blood clotting, hemolysis, and cytotoxicity analyses. As a result of this study, the composite pericardial material revealed improved mechanical and thermal stability and hemocompatibility in comparison to decellularized pericardium, without toxicity. Approximately 100% success is achieved in preventing platelet adhesion. In addition, kinetic blood-coagulation analysis demonstrated a low rate and slow coagulation kinetics, while the hemolysis index was below the permissible limit for biomaterials.

A novel mutation in TRMT5 associated with idiopathic non-cirrhotic portal hypertension and hepatopulmonary syndrome: Case report of two siblings.

Non-cirrhotic portal hypertension (NCPH) is a rare clinical entity in children. Familial clusters of idiopathic non-cirrhotic portal hypertension (INCPH) were previously reported in cases with deoxyguanosine kinase (DGOUK) and potassium calcium-activated channel subfamily N member 3 (KCNN3) mutations. Herein, we report two siblings who had a novel mutation in mitochondrial tRNA methyltransferase 5 (TRMT5) gene and presented with hepatopulmonary syndrome and later diagnosed as INCPH. Autosomal recessive inheritance of this mutation may suggest a role of TRMT5 mutations in the development of NCPH. Screening of TRMT5 mutations could be considered when familial INCPH is suspected.

Surface channel patterned and endothelialized poly(glycerol sebacate) based elastomers.

Prevascularization of tissue equivalents is critical for fulfilling the need for sufficient vascular organization for nutrient and gas transport. Hence, endothelial cell culture on biomaterials is of great importance for researchers. Numerous alternate strategies have been suggested in this sense, with cell-based methods being the most commonly employed. In this study, poly (glycerol sebacate) (PGS) elastomers with varying crosslinking ratios were synthesized and their surfaces were patterned with channels by using laser ablation technique. In order to determine an ideal material for cell culture studies, the elastomers were subsequently mechanically, chemically, and biologically characterized. Following that, human umbilical vein endothelial cells (HUVECs) were seeded into the channels established on the PGS membranes and cultured under various culture conditions to establish the optimal culture parameters. Lastly, the endothelial cell responses to the synthesized PGS elastomers were evaluated. Remarkable cell proliferation and impressive cellular organizations were noticed on the constructs created as part of the investigation. On the concrete output of this research, arrangements in various geometries can be created by laser ablation method and the effects of various molecules, drugs or agents on endothelial cells can be evaluated. The platforms produced can be employed as an intermediate biomaterial layer containing endothelial cells for vascularization of tissue-engineered structures, particularly in layer-by-layer tissue engineering approaches.

Bi-allelic loss-of-function OBSCN variants predispose individuals to severe recurrent rhabdomyolysis.

Rhabdomyolysis is the acute breakdown of skeletal myofibres in response to an initiating factor, most commonly toxins and over exertion. A variety of genetic disorders predispose to rhabdomyolysis through different pathogenic mechanisms, particularly in patients with recurrent episodes. However, most cases remain without a genetic diagnosis. Here we present six patients who presented with severe and recurrent rhabdomyolysis, usually with onset in the teenage years; other features included a history of myalgia and muscle cramps. We identified 10 bi-allelic loss-of-function variants in the gene encoding obscurin (OBSCN) predisposing individuals to recurrent rhabdomyolysis. We show reduced expression of OBSCN and loss of obscurin protein in patient muscle. Obscurin is proposed to be involved in sarcoplasmic reticulum function and Ca2+ handling. Patient cultured myoblasts appear more susceptible to starvation as evidenced by a greater decreased in sarcoplasmic reticulum Ca2+ content compared to control myoblasts. This likely reflects a lower efficiency when pumping Ca2+ back into the sarcoplasmic reticulum and/or a decrease in Ca2+ sarcoplasmic reticulum storage ability when metabolism is diminished. OSBCN variants have previously been associated with cardiomyopathies. None of the patients presented with a cardiomyopathy and cardiac examinations were normal in all cases in which cardiac function was assessed. There was also no history of cardiomyopathy in first degree relatives, in particular in any of the carrier parents. This cohort is relatively young, thus follow-up studies and the identification of additional cases with bi-allelic null OBSCN variants will further delineate OBSCN-related disease and the clinical course of disease.

Fabrication of a multi-layered decellularized amniotic membranes as tissue engineering constructs.

As a promising approach in tissue engineering, decellularization has become one of the mostly-studied research areas in tissue engineering thanks to its potential to bring about several advantages over synthetic materials since it can provide a 3-dimensional ECM structure with matching biomechanical properties of the target tissue. Amniotic membranes are the tissues that nurture the embryos during labor. Similarly, these materials have also been proposed for tissue regeneration in several applications. The main drawback in using amniotic membranes is the limited thickness of these materials since most tissues require a 3D matrix for an enhance regeneration. In order to prevent this limitation, here we report a facile fabrication methodology for multilayered amniotic membrane-based tissue constructs. The amniotic membranes of Wistar albino rats were first decellularized with the physical and chemical methods and utilized as scaffolds. Secondly, the prepared decellularized membranes were sutured to form a multilayered 3D structure. Within the study, 7 groups including control (PBS), were prepared based on physical and chemical decellularization methods. UV exposure and freezing techniques were used as a physical decellularization methods while hypertonic medium and SDS (sodium dodecyl sulfate) protocols were used as chemical decellularization methods. The combinations of both protocols were also used. In groups, A was the control and group B was applied just UV. In group C was applied UV and freezing. In addition to UV and freezing, in group D was applied hypertonic solution while group E was applied SDS (0.03 %). In group F was applied UV, freezing, hypertonic solution and SDS (0.03 %). In group G was applied UV, hypertonic solution, SDS (0.03 %) and freezing, respectively. Based on the histological and quantitative analyses, F and G groups were found as the most efficient decellularization protocols in rat amniotic membranes. Then, group F and G decellularized amniotic membranes were used to form scaffolds and thus-formed matrices were further characterized in vitro cell culture studies and mechanical tests. Cytotoxicity analyses performed using MTT showed a good cell viability in F and G groups scaffolds. The percentage viability rate was higher in G group (81.3 %) compared to F (75.33 %) and also cell viability in G group was found more meaningful according to p value which was obtained 0.007. Cellular adhesions after in vitro cell culture and morphology of scaffolds were evaluated by scanning electron microscopy (SEM). It was observed that the cells cultivated in equal amounts of tissue scaffolds were higher in the F compared to that observed in group G. The mechanical testing with 40 N force revealed 0.77 mm displacement in group F while it was 0.75 mm in group G. Moreover, according to force-controlled test, 2.9 mm displacement of F group and 1.2 mm displacement of G group was measured. As a result, this study shows that the multilayered decellularized amniotic membrane scaffolds support cell survival and adhesion and can form a flexible biomaterial with desired handling properties.

In vitro selection of DNA aptamers against human osteosarcoma.

BACKGROUND: Osteosarcoma is a highly malignant bone tumor, most frequently occurring in the rapid bone growth phase. Effective treatment of this disease is hindered by the lack of specific probes for early diagnosis and the fast cancer widespread. METHODS: To find such probes, the cell-Systematic Evolution of Ligands by EXponential enrichment (cell-SELEX) methodology was implemented against the human osteosarcoma MG-63 cell line towards the selection of new specific aptamers. After 10 rounds of selection, the aptamer DNA pool was Sanger sequenced and the sequences were subjected to a bioinformatic analysis that included sequence alignment, phylogenetic relationship, and secondary structure prediction. RESULTS: A DNA aptamer (OS-7.9), with a dissociation constant (Kd) value in the nanomolar range (12.8 ± 0.9 nM), revealed high affinity against the target cells at the physiological temperature. Furthermore, the selected aptamer also recognized lung carcinoma and colon colorectal adenocarcinoma cell lines, which are reported as common metastasis sites of osteosarcoma. CONCLUSIONS: These results suggest that OS-7.9 could recognize a common protein expressed in these cancer cells, possibly becoming a potential molecular probe for early diagnosis and targeted therapies for metastatic disease. Moreover, to the best of our knowledge, this was the first attempt to generate a DNA aptamer (OS-7.9 aptamer) against the MG-63-cell line by cell-SELEX.

Preparation of decellularized optic nerve grafts.

BACKGROUND: Decellularized tissues based on well-conserved extracellular matrices (ECMs) are a common area of research in tissue engineering. Although several decellularization protocols have been suggested for several types of tissues, studies on the optic nerve have been limited. METHODS: We report decellularization protocol with different detergent for the preparation of acellular optic nerve and tissues were examined. DNA, glycosaminoglycan (GAG), and collagen content of the groups were evaluated with biochemical analyses and examined with histological staining. Mechanical properties, chemical components as well as cytotoxic properties of tissues were compared. RESULTS: According to the results, it was determined that TX-100 (Triton X-100) was insufficient in decellularization when used alone. In addition, it was noticed that 85% of GAG content was preserved by using TX-100 and TX-100-SD (sodium deoxycholate), while this ratio was calculated as 30% for SDS. In contrast, the effect of the decellularization protocols on ECM structure of the tissues was evaluated by scanning and transmission electron microscopy (SEM and TEM) and determined their mechanical properties. Cytotoxicity analyses were exhibited minimum 95% cell viability for all groups, suggesting that there are no cytotoxic properties of the methods on L929 mouse fibroblast cells. CONCLUSIONS: The combination of TX-100-SD and TX-100-SDS (sodium dodecyl sulfate) were was determined as the most effective methods to the literature for optic nerve decellularization.

Biofabrication of Poly(glycerol sebacate) Scaffolds Functionalized with a Decellularized Bone Extracellular Matrix for Bone Tissue Engineering.

The microarchitecture of bone tissue engineering (BTE) scaffolds has been shown to have a direct effect on the osteogenesis of mesenchymal stem cells (MSCs) and bone tissue regeneration. Poly(glycerol sebacate) (PGS) is a promising polymer that can be tailored to have specific mechanical properties, as well as be used to create microenvironments that are relevant in the context of BTE applications. In this study, we utilized PGS elastomer for the fabrication of a biocompatible and bioactive scaffold for BTE, with tissue-specific cues and a suitable microstructure for the osteogenic lineage commitment of MSCs. In order to achieve this, the PGS was functionalized with a decellularized bone (deB) extracellular matrix (ECM) (14% and 28% by weight) to enhance its osteoinductive potential. Two different pore sizes were fabricated (small: 100-150 µm and large: 250-355 µm) to determine a preferred pore size for in vitro osteogenesis. The decellularized bone ECM functionalization of the PGS not only improved initial cell attachment and osteogenesis but also enhanced the mechanical strength of the scaffold by up to 165 kPa. Furthermore, the constructs were also successfully tailored with an enhanced degradation rate/pH change and wettability. The highest bone-inserted small-pore scaffold had a 12% endpoint weight loss, and the pH was measured at around 7.14. The in vitro osteogenic differentiation of the MSCs in the PGS-deB blends revealed a better lineage commitment of the small-pore-sized and 28% (w/w) bone-inserted scaffolds, as evidenced by calcium quantification, ALP expression, and alizarin red staining. This study demonstrates a suitable pore size and amount of decellularized bone ECM for osteoinduction via precisely tailored PGS elastomer BTE scaffolds.

Fabrication and characterization of carbon aerogel/poly(glycerol-sebacate) patches for cardiac tissue engineering.

Cardiovascular diseases (CVDs) are responsible for the major number of deaths around the world. Among these is heart failure after myocardial infarction whose latest therapeutic methods are limited to slowing the end-state progression. Numerous strategies have been developed to meet the increased demand for therapies regarding CVDs. This study aimed to establish a novel electrically conductive elastomer-based composite and assess its potential as a cardiac patch for myocardial tissue engineering. The electrically conductive carbon aerogels (CAs) used in this study were derived from waste paper as a cost-effective carbon source and they were combined with the biodegradable poly(glycerol-sebacate) (PGS) elastomer to obtain an electrically conductive cardiac patch material. To the best of our knowledge, this is the first report about the conductive composites obtained by the incorporation of CAs into PGS (CA-PGS). In this context, the incorporation of the CAs into the polymeric matrix significantly improved the elastic modulus (from 0.912 MPa for the pure PGS elastomer to 0.366 MPa for the CA-PGS) and the deformability (from 0.792 MPa for the pure PGS to 0.566 MPa for CA-PGS). Overall, the mechanical properties of the obtained structures were observed similar to the native myocardium. Furthermore, the addition of CAs made the obtained structures electrically conductive with a conductivity value of 65 × 10-3S m-1which falls within the range previously recorded for human myocardium. Thein vitrocytotoxicity assay with L929 murine fibroblast cells revealed that the CA-PGS composite did not have cytotoxic characteristics. On the other hand, the studies conducted with H9C2 rat cardiac myoblasts revealed that final structures were suitable for MTE applications according to the successes in cell adhesion, cell proliferation, and cell behavior.