CAR-T cell-derived exosomes: a new perspective for cancer therapy.

Chimeric antigen receptor (CAR)-T cell adoptive immunotherapy is a promising cancer treatment that uses genetically engineered T cells to attack tumors. However, this therapy can have some adverse effects. CAR-T cell-derived exosomes are a potential alternative to CAR-T cells that may overcome some limitations. Exosomes are small vesicles released by cells and can carry a variety of molecules, including proteins, RNA, and DNA. They play an important role in intercellular communication and can be used to deliver therapeutic agents to cancer cells. The application of CAR-T cell-derived exosomes could make CAR-T cell therapy more clinically controllable and effective. Exosomes are cell-free, which means that they are less likely to cause adverse reactions than CAR-T cells. The combination of CAR-T cells and exosomes may be a more effective way to treat cancer than either therapy alone. Exosomes can deliver therapeutic agents to cancer cells where CAR-T cells cannot reach. The appropriate application of both cellular and exosomal platforms could make CAR-T cell therapy a more practicable treatment for cancer. This combination therapy could offer a safe and effective way to treat a variety of cancers.

Engineered artificial articular cartilage made of decellularized extracellular matrix by mechanical and IGF-1 stimulation.

Cartilage engineering has the potential to overcome clinical deficiency in joint disorders. Decellularized extracellular matrix (dECM) has great biocompatibility and bioactivity and can be considered an appropriate natural scaffold for tissue engineering applications. Both insulin-like growth factor-1 (IGF-1) and mechanical compression stimulate the production of cartilage ECM, modulate mechanical properties, and gene expression. The current investigation aimed to fabricate a high-quality moldable artificial cartilage by exposing the chondrocytes in biomimicry conditions using cartilage dECM, IGF-1, and mechanical stimulations. In this study, an ad hoc bioreactor was designed to apply dynamic mechanical stimuli (10 % strain, 1 Hz) on chondrocyte-laden cartilage dECM-constructs with/without IGF-1 supplementation for 2 weeks, 3 h/day. Our data revealed that mechanical stimulation had no adverse effect on cell viability and proliferation. However, it elevated the expression of chondrogenic markers such as collagen type II (COL2A1), aggrecan (ACAN), and proteoglycan-4 (PRG-4), and reduced the expression of matrix metalloproteinase-3 (MMP-3). Mechanical stimulation also promoted higher newly formed glycosaminoglycan (GAG) and produced more aligned fibers that can be responsible for higher Young's modulus of the engineered construct. Even though IGF-1 demonstrated some extent of improvement in developing neocartilage, it was not as effective as mechanical stimulation. Neither IGF-1 nor compression elevated the collagen type I expression. Compression and IGF-1 showed a synergistic impact on boosting the level of COL2A1 but not the other factors. In conclusion, mechanical stimulation on moldable cartilage dECM can be considered a good technique to fabricate artificial cartilage with higher functionality.

Synergistic impact of platelet rich plasma-heparin sulfate with hydroxyapatite/zirconia on the osteoblast differentiation potential of adipose-derived mesenchymal stem cells.

3D porous hydroxyapatite (HA) has been reinforced by zirconia (ZrO2) coating and impregnation with a combination of platelet rich plasma (PRP) as a source of growth factors (GFs) and Heparin sulfate (HS) to sustain the release of GFs. Adipose mesenchymal stem cells (ADMSCs) were characterized by flow cytometry for CD (cluster of differentiation) 44, CD105, CD106, CD34 and CD144, along with checking the multipotency by differentiation into the adipocytes and osteoblasts. Then, they were cultured on the scaffold treated with and without osteogenic media on days 7, 14 and 21. Electron micrograph and PKH staining show that the ADMSCs have a fusiform phenotype in the absence of osteogenic induction. Cell viability assay shows a higher number of the viable cells on the PRP-containing scaffolds than PRP-free scaffolds on day 7. Colorimetric evaluation, quantitative RT-PCR and immunocytochemistry demonstrate that PRP and HS significantly elevate the alkaline phosphatase enzyme activity and also accelerate the production of both early and mid-osteogenic markers, including collagen I and osteopontin expression with and without osteogenic conditions. The PRP-HS also accelerates the expression of the late osteogenic marker, osteocalcin, in both mRNA and protein level expression with a peak on day 21. In conclusion, supplementation of HA/ZrO2 with PRP/HS has a synergistic impact on the ADMSCs, even in the absence of chemical induction. It seems that HA/ZrO2/PRP/HS scaffold provides a higher osteoconductive microenvironment for stem cell differentiation to osteoblasts.

Evaluation of the Possible Synergic Regenerative Effects of Platelet-Rich Plasma and Hydroxyapatite/Zirconia in the Rabbit Mandible Defect Model.

BACKGROUND: Platelet-rich plasma (PRP) and bioceramics such as hydroxyapatite (HA) and zirconium oxide (ZrO2) are used to reconstruct mandibular defects. We sought to determine the synergistic effects of HA/ZrO2 and PRP and compare their osteogenic activity. METHODS: ZrO2 scaffolds were constructed by the slurry method and were then coated with HA and impregnated by PRP/heparan sulfate (HS). Bilateral mandibular defects were created in 26 male rabbits. In 20 rabbits, the left defects were treated with HA/ZrO2/PRP (Group 1) and the corresponding right defects were filled with HA/ZrO2 (Group 2). The 6 remaining models were treated with PRP gels at both sides (Group 3). The osteoconductivity of HA/ZrO2/PRP was compared with that of HA/ZrO2 or PRP by radiological and histological methods after the follow-up period, at weeks 2, 6 and 8. The statistical analyses were performed by ANOVA and LSD using SPSS, version 16.0, for Windows (P<0.05). RESULTS: After 2 weeks, the percentage of the surface occupied by bone was significantly higher in the HA/ZrO2/PRP-treated defects than in the PRP-treated defects (P=0.007). Osteoblast and osteocyte counts were higher significantly in the PRP-treated group (P=0.032); however, the cells had not started matrix formation on a large scale and just small islands of osteoid with trapped osteocytes were observed. In the long term, the regenerative potential of all the scaffolds was the same. CONCLUSION: HA/ZrO2 showed a superior osteoconductive capacity over PRP in the short term; however, they showed no long-term synergic effects.

Fabrication and characterization of platelet-rich plasma scaffolds for tissue engineering applications.

Platelet-Rich Plasma (PRP), as a rich source of growth factor, can form a fibrin gel that recapitulates the extracellular matrix of the tissues. The aim of this study was to evaluate the effects of different concentrations of CaCl2 on the PRP scaffold structure which in turn could change the cell's behavior. PRP was mixed with 2.5, 5 and 10% (w/v) CaCl2. Then, the tensile strength, biodegradability and water content of the scaffolds were evaluated. We also performed immunostaining for assessment of the actin stress fiber orientation and SEM for detecting the cell phenotype and physical properties of the fibers. Cell viability, attachment and migration were also evaluated. The highest cell attachment and short term proliferation rate was observed on the scaffolds with 2.5% CaCl2. The cells cultured on the scaffold with higher CaCl2 concentration had fusiform phenotype with few cell processes and parallel arrangement of stress fibers while those cultured on the other scaffolds were fibroblast-like with more processes and net-like stress fibers. The scaffolds with 10% CaCl2 demonstrated the highest osmolarity (358.75±4.99mOsmole), fiber thickness (302.1±54.3nm), pore size (332.1±118.9nm2) and the longest clotting time (12.2±0.776min) compared with the other scaffolds. Water content, branching angle, porosity, orientation and tensile strength did not change by gelation with different CaCl2 concentrations. In conclusion, the cell shape, viability and proliferation were modified by culturing on the PRP scaffolds prepared with various concentrations of CaCl2, and as a result, the scaffolds showed different physical and biological properties.

Haplotypes of NOS3 gene polymorphisms in dilated cardiomyopathy.

Dilated Cardiomyopathy (DCM) is characterized by systolic dysfunction, followed by heart failure necessitating cardiac transplantation. The genetic basis is well established by the identification of mutations in sarcomere and cytoskeleton gene/s. Modifier genes and environmental factors are also considered to play a significant role in the variable expression of the disease, hence various mechanisms are implicated and one such mechanism is oxidative stress. Nitric Oxide (NO), a primary physiological transmitter derived from endothelium seems to play a composite role with diverse anti-atherogenic effects as vasodilator. Three functional polymorphisms of endothelial nitric oxide synthase (NOS3) gene viz., T-786C of the 5' flanking region, 27bp VNTR in intron4 and G894T of exon 7 were genotyped to identify their role in DCM. A total of 115 DCM samples and 454 controls were included. Genotyping was carried out by PCR -RFLP method. Allelic and genotypic frequencies were computed in both control & patient groups and appropriate statistical tests were employed. A significant association of TC genotype (T-786C) with an odds ratio of 1.74, (95% CI 1.14 - 2.67, p = 0.01) was observed in DCM. Likewise the GT genotypic frequency of G894T polymorphism was found to be statistically significant (OR 2.10, 95% CI 1.34-3.27, p = 0.0011), with the recessive allele T being significantly associated with DCM (OR 1.64, 95% CI 1.18 - 2.30, p = 0.003). The haplotype carrying the recessive alleles of G894T and T-786C, C4bT was found to exhibit 7 folds increased risk for DCM compared to the controls. Hence C4bT haplotype could be the risk haplotype for DCM. Our findings suggest the possible implication of NOS3 gene in the disease phenotype, wherein NOS3 may be synergistically functioning in DCM associated heart failure via the excessive production of NO in cardiomyocytes resulting in decreased myocardial contractility and systolic dysfunction, a common feature of DCM phenotype.