Chondrocytes Proliferation of Patients with Cartilage Lesions in Their Own Body for Use in Cartilage Tissue Engineering: Hypotheses on a New Approach for the Proliferation of Autologous Chondrocytes

Osteoarthritis is one of the most common chronic diseases, which have involved 250 million people around the world. One of the challenges in the field of cartilage tissue engineering is to provide an adequate source of chondrocytes to prevent changes in gene expression profile as a result of multiple passages. We hypothesized that by creating a low invasive lesion by scalpel or shear laser in the outer ear cartilage and stimulation of wound healing process, hyperplasia occurs and will provide an appropriate number of autologous chondrocytes for extraction and use in articular cartilage tissue engineering. Also, due to the effect of platelet-rich plasma and biomechanical forces in stimulating and accelerating of the repair process, these two factors can be used to achieve more desirable results. We describe a new approach to proliferate chondrocytes in the body. To evaluate this idea, various techniques of gene expression at the level of RNA or protein and animal experiments for histological studies can be used. Also, flowcytometry technique can be used to determine the cell viability and counting them. The use of autologous cell sources with minimal changes in gene expression profile can be promising in tissue engineering products. Keywords Author Keywords:Chondrocyte; Cell Proliferation; Cartilage; Tissue Regeneration; Osteoarthritis KeyWords Plus:HUMAN ARTICULAR CHONDROCYTES; GENE-EXPRESSION; STRATEGIES; REPAI

Antiproliferative effects of fresh water crab hemolymph and meat extract on breast cancer cell line

<strong>Background and aims:</strong> Despite the advances in drugs, side effects of chemotherapy drugs continue to exist. Therefore, more attention has been paid to the compounds derived from medicinal herbs and aquatic organisms. This study aimed to investigate the effect of freshwater crab hemolymph and meat extract on breast cancer (BC) cell line (4T1). <strong>Methods:</strong> After isolation of freshwater crab hemolymph and meat extract, protein concentration and total antioxidant capacity were analyzed by bicinchoninic acid (BCA) and cupric reducing antioxidant capacity (CUPRAC) methods. The 4T1 cells and bone marrow mesenchymal stem cells (BMSCs) were treated with crab hemolymph (1, 2, 10 mg/mL) and meat extract (0.1, 0.2 and 1 mg/mL), and cell survival was analyzed using 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (MTT) at 48 and 72 hours. Nitric oxide (NO) secretion was measured by Griess method. Data were analyzed using one-way analysis of variance (ANOVA). <strong>Results:</strong> Protein concentration of 23.25 mg/mL was shown in crab hemolymph, and 2.3 mg/mL in meat extract. Total antioxidant capacity was reported as 1.036 µM/mL and 1.104 µM/mL in crab hemolymph and meat extract, respectively. Cell survival in the 4T1 cells was decreased in a dose- and time-dependent manner (<em>P</em>�0.001). NO secretion of 4T1 cells was decreased after treatment with different concentrations of crab hemolymph and meat extract at 48 and 72 hours. Cellular growth was observed in BMSCs after treatment with different concentrations of crab hemolymph and meat extract at 48 and 72 hours. <strong>Conclusion:</strong> Since crab hemolymph and meat extract have protein and antioxidant activities, they can have anti-cancer effects on 4T1 cells

Evaluation of vacuum washing in the removal of SDS from decellularized bovine pericardium: method and device description.

AIMS: The aim of this study was to present a new method for removing Sodium dodecyl sulfate (SDS) detergent from decellularized bovine pericardium using vacuum. MATERIALS AND METHODS: The cows' pericardia were collected and decellularized. The samples were incubated with SDS1% for 48 h at 40 °C. To perform vacuum washing (VW: negative pressure was used to wash and remove detergents), every decellularized tissue was cut in 75mm diameter and fixed via a stainless-steel ring with 60mm diameter in the center of filtration Buchner Funnel which was connected to glass filtration flask The system was connected to a vacuum pump by a hose, and a negative pressure of -100 mmHg was applied for 15 min. Then, the samples were shaken and washed at 40-rpm in 100 ml of distilled water for 45 min. This process was repeated for samples of each group (6 times for sample VW6h, 12 times for sample VW12h, and 24 times for sample VW24h). At the end of every cycle, the effluent was collected to take a sample for SDS measurement. The normal washing (NW) group containing distilled water (NWd) and PBS (Phosphate buffered saline) (NWp) were used to wash and remove detergents. SDS measurements, MTT Assay, histological and tensile test, to compare two methods were used. RESULTS: The highest SDS in the effluent was in groups VW12h and VW24h (P ≤ 0.001) and the lowest residual SDS in scaffold was in two groups of VW12h and VW24h (P ≤ 0.001). MTT assay showed that cell survival in the VW12h and VW24h groups was higher than other groups and there' was no significant difference between cell survival in the VW12h and VW24h groups. Histological study showed destruction of tissue in the VW24h group. The results of the tensile test were shown that the native group had the highest module and the lowest amount was the VW24h sample which was reported with P ≤ 0.001 significance for all groups. CONCLUSION: VW12h can be used as an effective method for SDS removal from decellularized pericardium which morphologically demonstrated a good structure in ECM. KEYWORDS: Acellular; Cell biology; Cell culture; Cytotoxicity; Extracellular matrix; Pericardium; Regenerative medicine; Sodium dodecyl sulfate; Stem cells research; Toxicit

Evaluating the effects of vacuum on the microstructure and biocompatibility of bovine decellularized pericardium

The aim of this study was evaluating the effects of vacuum on microstructure and biocompatibility of bovine decellularized pericardium. So the bovine pericardia were decellularized and then the vacuum was applied for two periods of time; 90 and 180 min. DNA, glucose amino glycan, collagen and elastin content assay, scanning electron microscopy (SEM) examination, hematoxylin and eosin (H&E) and Masson's trichrome stainings performed to evaluate microstructure of tissues. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test, subcutaneous implantation, and tensile test were used to assay biocompatibility and mechanical properties of decellularized tissues. The results showed that applying vacuum reduced residual DNA significantly. Vacuum after 180 min reduced more residual DNA. There were no significant differences in the content of glucose amino glycan (GAG), collagen, and elastin between the vacuumed and control groups. SEM examination was revealed that vacuum for 180 min increased pore size and porosity more than 90 min and control groups. H&E and Masson's trichrome stainings revealed extracellular matrix preservation after decellularization in all groups. Cell viability was increased in vacuumed samples significantly after 72 h in vaccumed samples. H&E staining and tensile test after implantation of tissues were showed less inflammation in the vacuum applied tissues and increased durability. The vacuum increased DNA removal, pore size, porosity, and biocompatibility in vitro and in vivo and durability of bovine decellularized pericardium in vivo. Considering the important role of time, more studies should be performed to optimize time, intensity, and method of application of vacuum in decellularization of different tissues as well as bovine pericardium

Evaluation of the Effect of Cardiac Rehabilitation on Left Ventricular Diastolic and Systolic Function and Cardiac Chamber Size in Patients Undergoing Percutaneous Coronary Intervention

Background: Exercise and rehabilitation are important methods for decreasing the risk factors of coronary artery disease (CAD). We aimed to evaluate the effect of the cardiac rehabilitation (CR) exercise program on the cardiac structure and physiology in patients undergoing percutaneous coronary intervention (PCI). Methods: In this randomized controlled study, 146 patients with CAD were divided equally into two groups: case group (undertaking CR after PCI) and control group (without rehabilitation after PCI). All the patients in the case group underwent echocardiography (before and after CR), and echocardiography was performed for the control group simultaneously. The CR exercise program encompassed 24 sessions, twice or three times a week, with each session lasting between 15 and 45 minutes, depending on the individual patient’s tolerance. Left ventricular (LV) ejection fraction, LV diastolic function, LV end-systolic and diastolic diameter, and right ventricular (RV) end-diastolic diameter were measured in the CR group before and after rehabilitation and compared to those in the control group at the same times. Results: In this study, 146 patients (46 female and 100 male) were evaluated: 73 in the rehabilitation group and 73 in the control group. The mean age of the patients in the CR and control groups was 58.05 ± 10.27 and 56.76 ± 10.07 years, respectively. The CR exercise program had useful effects on LV diastolic function after PCI. The distribution of LV diastolic dysfunction after the CR exercise program was changed significantly only in the CR group (p value = 0.043). In the CR group, normal, grade I, grade II, and grade III LV diastolic dysfunction were observed in 20.5%, 69.8%, 6.8%, and 2.7%, respectively. This distribution was changed respectively to 30.1%, 61.6%, 5.4%, and 2.7% following CR, which showed a significant improvement due to CR in LV diastolic function, most prominently in the patients with grade I diastolic dysfunction (p value = 0.390). There was no significant change in LV and RV diameter before and after rehabilitation, while the ejection fraction increased significantly (p value < 0.05) in both groups. Conclusion: The RC exercise program can be effective in the augmentation of LV diastolic dysfunction after PCI, withoutsignificant changes in LV diameters

Isolation and Characterization of Crab Haemolymph Exosomes and Its Effects on Breast Cancer Cells (4T1)

Objective: The use of animal or plant exosomes in cancer treatment is promising because of their easy access and low cost. Freshwater crabs are used in traditional Iranian medicine to treat cancer. This study aims to determine the anti-cancer properties of exosomes removed from freshwater crabs on a breast cancer cell line (4T1) compared to bone marrow mesenchymal stem cells (BMSCs). Materials and Methods: In this experimental study, crab haemolymph exosomes were isolated via the precipitation method and characterised by electron microscopy, dynamic light scattering (DLS), and Western blot analysis. The protein concentration and total antioxidant capacity of these exosomes were determined by bicinchoninic acid (BCA) and cupric reducing antioxidant capacity (CUPRAC). The 4T1 cells and BMSCs were treated with exosomes and we assessed the cell survival by the resazurin and MTT assays. The level of nitric oxide (NO) secretion from the 4T1 cells was determined after treatment with the exosomes. Results: Electron microscopy, DLS and Western blot for CD63 confirmed that the isolated exosomes were <100 nm in size and expressed CD63. The total antioxidant capacity in these exosomes was 1.003 mu M/ml and the protein concentration was 650 mg/ml. Resazurin and MTT assay results showed a decrease in survival of the 4T1 cells (P <= 0.001) after treatment with the exosomes compared to cell growth in the exosome-treated BMSCs. Conclusion: Crab haemolymph contains protein-rich exosomes with antioxidant activities that can have anti-cancer effects on 4T1 cells. These exosomes may be proposed for breast cancer therapeutics

Retinoic acid and taurine enhance differentiation of the human bone marrow stem cells into cone photoreceptor cells and retinal ganglion cells

Degeneration and apoptotic death of the photoreceptor cell-layer of retina are a major cause of irreversible blindness in the development era. The stem cell replacement therapy is one of the strategies for the retinal repairing. In addition, exogenous signals critically contribute to the direction of lineage decisions that causes the fate-restricted photoreceptor progenitors from stem cell progeny in culture. It has been found that epidermal growth factor (EGF), taurine, and retinoic acid (RA) initially act in the instructive as well as lineage-restricted way in the progenitor lineage for producing neuroretinal cells or photoreceptor like cells from stem cell. The study aims to investigate the effect of RA and taurine in differentiation of the human bone marrow stem cell into cone photoreceptors cells and retinal ganglion cells. Mesenchymal stem cell was derived from human bone marrow of the term delivery. Therefore, the cultured cells have been treated with Dulbecco's modified Eagle's medium (DMEM)/high glucose (H+). After the four-cell passage, basal medium was replaced with DMEM/F12 complemented with 50 mu mol/L taurine, RA (1 mu M) and EGF (1 mu g/ml). Subsequently cellular change morphology was detected following 7 and 14 days. Then, gene expression of neuroretinal and photoreceptor cell biomarkers (CRX, OTX2, PKC-alpha, recoverin, and Rho) were examined by quantitative polymerase chain reaction (Q-PCR). Also, cells were cultured, fixed, and then immunocytochemical analyzed. Primary antibodies included CRX and Rho. Cellular morphology demonstrated spindle elongated morphology. Taurine alone and combination of RA upregulate neuroretinal and photoreceptor cell biomarkers in messenger RNA and protein levels but along with EGF have not significant effect. Our data showed that taurine combination with RA can differentiate bone marrow mesenchymal stem cells into neuroretinal or photoreceptor like cells in vitro that can offer an attractive treatment ground for transplantation in the cell-replacement therapy for some forms of the retinal degeneration

Preparation and in vitro characterization of electrospun scaffolds composed of chitosan, gelatin and 58S bioactive glass nanoparticles for skin tissue engineering

Background and aims: The presence of an appropriate scaffold at the wound site could significantly improve the healing process. In this study, we aimed to prepare a biomimetic nanocomposite scaffold composed of chitosan, gelatin, and 58S bioglass nanoparticles for skin tissue engineering. Methods: The nanocomposite scaffolds composed of chitosan, gelatin, and 58S bioglass nanoparticles were fabricated through electrospinning process. Then the cell viability assay was performed in order to evaluate the biological properties of the membranes. The optimum concentration of bioglass nanoparticles was determined for further studies. In vitro characterization was also performed to evaluate physicochemical properties of the scaffolds. Results: The chitosan/gelatin scaffold containing 2% of 58S bioglass nanoparticles showed no cell toxicity, and the dermal fibroblasts were found capable of proliferation on the membrane. The in vitro results obtained from the scanning electron microscopy (SEM), attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR), and porosity tests demonstrated the appropriate properties of the membrane as a scaffold for skin regeneration. Conclusions: It was concluded that a chitosan-gelatin membrane containing 2% of 58S bioglass nanoparticles had the potential to function as a scaffold to accelerate wound healing due to its suitable properties, such as high porosity, high surface/volume ratio, and excellent biocompatibility. Keywords: Chitosan, Gelatin, Bioactive glass, Wound healing, Tissue engineerin

The Protective Effects of Lavender officinalis Extract Against Impairment of Antioxidant–Detoxification System Induced by Glucose Deprivation Through Nrf2 Expression

peer reviewedDespite tremendous progress in the prevention and treatment of cerebral ischemia, stroke remains one of the major causes of mortality and paralysis among the elderly. The design and study of neuroprotective drugs in vitro, such as in the Serum-Glucose Deprivation (SGD) condition, are proved to be efficient to understand how these drugs defend neuronal cells from ischemia-induced damage. Neurological defects and stroke volume are reduced by lavender plant’s potent and abundant compounds reduces. In this study, we intend to study the mechanism of a protective effect of the lavender extract on insult caused by glucose deprivation in the PC12 cell line, focusing on the Nrf2 pathway. In order to investigate the protective effects of the extract, the cells were subjected to serum/glucose deprivation for a period of 12 h at non-toxic concentrations and viability was investigated. Expression levels of Nrf2, HO-1, and NQO1 genes were evaluated. Also, the activity of CAT, SOD, and GPx enzymes was measured. The results illustrate an increase in cell viability, expression of HO-1 and NQO1 genes, and enzymatic activity of CAT, SOD, GPx. In conclusion, pretreatment of PC12 cells with hydroalcoholic extract of lavender reinforces antioxidant system and detoxification enzymes under oxidative stress

Fabrication of functional and nano-biocomposite scaffolds using strontium-doped bredigite nanoparticles/polycaprolactone/poly lactic acid via 3D printing for bone regeneration

Bone tissue engineering is a field to manufacture scaffolds for bone defects that cannot repair without medical interventions. Ceramic nanoparticles such as bredigite have importance roles in bone regeneration. We synthesized a novel strontium (Sr) doped bredigite (Bre) nanoparticles (Bre-Sr) and then developed new nanocomposite scaffolds using polycaprolactone (PCL), poly lactic acid (PLA) by the 3D-printing technique. Novel functional nanoparticles were synthesized and characterized using field emission scanning electron microscopy (FESEM), X-ray diffraction (XRD), and energy dispersive spectroscopy (EDS: map). The nanoparticles were uniformly distributed in the polymer matrix composites. The 3D- printed scaffolds were investigated using scanning electron microscopy (SEM), X-ray diffraction (XRD), attenuated total reflection-fourier transform infrared (ATR-FTIR), degradation rate porosity, mechanical tests, apatite formation and cell culture. Degradation rate and mechanical strength were increased in the PLA/PCL/Bre-5%Sr nanocopmposite scaffolds. Hydroxyapatite crystals were also created on the scaffold surface in the bioactivity test. The scaffolds supported viability and proliferation of human osteoblasts. Gene expression and calcium deposition in the samples containing nanoparticles indicated statistical different than the scaffolds without nanoparticles. The nanocomposite scaffolds were implanted into the critical-sized calvarial defects in rat for 3 months. The scaffolds containing Bre-Sr ceramic nanoparticles exhibited the best potential to regenerate bone tissue