Multiple Sclerosis Gene Therapy Using Recombinant Viral Vectors: Overexpression of IL-4, IL-10 and Leukemia Inhibitory Factor in Wharton's Jelly Stem Cells in The EAE Mice Model

Objective: Immunotherapy and gene therapy play important roles in modern medicine. The aim of this study is to evaluate the overexpression of interleukin-4 (IL-4), IL-10 and leukemia inhibitory factor (LIF) in Wharton's jelly stem cells (WJSCs) in the experimental autoimmune encephalomyelitis (EAE) mice model. Materials and Methods: In this experimental study, a DNA construction containing IL-4, IL-10 and LIF was assembled to make a polycistronic vector (as the transfer vector). Transfer and control vectors were co-transfected into Human Embryonic Kidney 293 (HEK-293T) cells with helper plasmids which produced recombinant lentiviral viruses (rLV). WJSCs were transduced with rLV to make recombinant WJSC (rWJSC). In vitro protein and mRNA overexpression of IL-4, LIF, and IL-10 were evaluated using quantitative polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA) and western blot (WB) analysis. EAE was induced in mice by MOG-CFA and pertussis toxin. EAE mice were injected twice with 2x10(5) rWJSCs. The in vivo level of IL-4, LIF, IL-10 cytokines and IL-17 were measured by ELISA. Brain tissues were analyzed histologically for evaluation of EAE lesions. Results: Isolated WJSCs were performed to characterize by in vitro differentiation and surface markers were analyzed by flow cytometry method. Cloning of a single lentiviral vector with five genes was done successfully. Transfection of transfer and control vectors were processed based on CaPO4 method with > 90% efficiency. Recombinant viruses were produced and results of titration showed 2-3x10(7) infection-unit/ml. WJSCs were transduced using recombinant viruses. IL-4, IL-10 and LIF overexpression were confirmed by ELISA, WB and qPCR. The EAE mice treated with rWJSC showed reduction of Il-17, and brain lesions as well as brain cellular infiltration, in vivo. Weights and physical activity were improved in gene-treated group. Conclusion: These results showed that gene therapy using anti-inflammatory cytokines can be a promising approach against multiple sclerosis (MS). In addition, considering the immunomodulatory potential of WJSCs, an approach using a combination of WJSCs and gene therapy will enhance the treatment efficacy

β-mangostin suppresses LA-7 cells proliferation in vitro and in vivo: involvement of antioxidant enzyme modulation; suppression of matrix metalloproteinase and α6β4 integrin signalling pathways

β-mangostin (βM) was isolated from Cratoxylum arborescens to investigate anti-breast cancer effect in vitro and in vivo. βM exhibited an inhibitory effect on the growth of LA-7 cells in vitro with apoptosis formation. In the animal model, βM treatment was found to be effective in improving the tissue antioxidant enzymes such as superoxide dismutase and catalase activity (P < 0.05). βM treatment clearly exhibited apoptosis in mammary tumour tissues, and it was associated with regulation of PCNA and p53. The cDNA microarray gene expression followed by qRT-PCR based validation demonstrated that βM could mediate tumour reduction and prevent metastasis by reduction of MMP-9, MMP-13, and MMP-27. Moreover, the reduction of both 14-3-3β and ITGB4 genes indicated the involvement of α6β4 integrin signalling pathway. These findings showed that β-mangostin is a promising compound candidate as an anti-tumour agent against breast cancer

Multiple Sclerosis Gene Therapy with Recombinant Viral Vectors: Overexpression of IL-4, Leukemia Inhibitory Factor, and IL-10 in Wharton's Jelly Stem Cells Used in EAE Mice Model.

OBJECTIVES: Immunotherapy and gene therapy play important roles in modern medicine. The aim of this study is to evaluate the overexpression of interleukin-4 (IL-4), IL-10 and leukemia inhibitory factor (LIF) in Wharton's jelly stem cells (WJSCs) in the experimental autoimmune encephalomyelitis (EAE) mice model. MATERIALS AND METHODS: In this experimental study, a DNA construction containing IL- 4, IL-10 and LIF was assembled to make a polycistronic vector (as the transfer vector). Transfer and control vectors were co-transfected into Human Embryonic Kidney 293 (HEK-293T) cells with helper plasmids which produced recombinant lentiviral viruses (rLV). WJSCs were transduced with rLV to make recombinant WJSC (rWJSC). In vitro protein and mRNA overexpression of IL-4, LIF, and IL-10 were evaluated using quantitative polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA) and western blot (WB) analysis. EAE was induced in mice by MOG-CFA and pertussis toxin. EAE mice were injected twice with 2×105 rWJSCs. The in vivo level of IL-4, LIF, IL-10 cytokines and IL-17 were measured by ELISA. Brain tissues were analyzed histologically for evaluation of EAE lesions. RESULTS: Isolated WJSCs were performed to characterize by in vitro differentiation and surface markers were analyzed by flow cytometry method. Cloning of a single lentiviral vector with five genes was done successfully. Transfection of transfer and control vectors were processed based on CaPO4 method with >90% efficiency. Recombinant viruses were produced and results of titration showed 2-3×107 infection-unit/ml. WJSCs were transduced using recombinant viruses. IL-4, IL-10 and LIF overexpression were confirmed by ELISA, WB and qPCR. The EAE mice treated with rWJSC showed reduction of Il-17, and brain lesions as well as brain cellular infiltration, in vivo. Weights and physical activity were improved in gene-treated group. CONCLUSIONS: These results showed that gene therapy using anti-inflammatory cytokines can be a promising approach against multiple sclerosis (MS). In addition, considering the immunomodulatory potential of WJSCs, an approach using a combination of WJSCs and gene therapy will enhance the treatment efficacy

β-Mangostin induces p53-dependent G2/M cell cycle arrest and apoptosis through ROS mediated mitochondrial pathway and NfkB suppression in MCF-7 cells

β-Mangostin (βM) was isolated from Cratoxylum arborescens to investigate its anti-cancer effect in MCF-7 cells. βM induced apoptosis by down-regulation of Bcl2 and up-regulation of Bax, triggering the cytochrome c release from mitochondria to cytosol. The release of caspase-9 and -7 and consequently cleaved PARP leading to apoptotic was observed upon treatment. Reduction of both bid and caspase 8 and the up regulation of Fas showed the involvement of the extrinsic pathway. Significantly up regulated GADD45A and HRK genes were observed upon treatment, with concomitant inhibition of NF-kB to nucleus. The protein array had demonstrated the expression of HSP 70, HSP 60, XIAP, Survivin, p53 and Bax. Moreover, βM had showed p53-dependent G2/M cell cycle arrest by down regulation of cdc2 and PCNA. Together, the results demonstrated that the βM induced anti-proliferative effect, leading to G2/M phase cell cycle arrest and apoptosis through both the extrinsic and mitochondrial pathways with the involvement of the multiple pro and anti-apoptosis and NF-kB signalling pathways

The association between amount and distribution of protein intake with body composition, resting energy expenditure, substrate oxidation, and muscle function in bariatric surgery candidates: a preliminary study

Background: Obesity is associated with disrupted energy metabolism and skeletal muscle dysfunction. Dietary protein is important for the preservation of skeletal muscle mass and strength. Objectives: The objectives of this preliminary study were: 1) to assess the association of dietary protein intake with body composition, and muscle function; 2) to examine weather protein distribution pattern throughout the day are associated with body composition and substrate oxidation; and 3) to explore the association between body composition with substrate oxidation and energy expenditure in males and females with severe obesity. Methods: This cross-sectional study included 17 male and female bariatric surgery candidates. Dietary data, including protein intake was collected using a 3-day food journal and 24-hour food recall. Body composition was assessed using dual-energy X-ray absorptiometry. Substrate oxidation and resting energy expenditure were measured by indirect calorimetry. Muscle function was assessed by handgrip strength and 6-minute walk test. Results: Mean age and BMI were 43.9 ± 7.8 y and 46.9 ± 4.9 kg/m2, respectively. Daily protein distribution was uneven and had a skewness toward dinner. Males who ate ≥ 20 g protein at each meal had greater ALM in comparison to those who did not (38.0 ± 0.9 vs 35.2 ± 0.7, P = 0.03). ALM was an independent predictor of REE and fat oxidation rate. Females had altered substrate metabolism and decreased rates of fat oxidation. Conclusion: This study provided preliminary results that will help improve study design and considerations in the continued recruitment of participants

SIMULTANEOUS SENSING OF PRESSURE AND TEMPERATURE USING A SELF-TEMPERATURE-COMPENSATED FABRY–PÉROT MEMS MECHANISM

This thesis presents the design and development of a self-temperature-compensated sensor for measuring temperature and pressure in harsh environments using a combination of Fabry–Pérot interferometry and microelectromechanical systems (MEMS). A silicon-on-insulator (SOI) wafer is etched to form a dual mechanism consisting of a membrane and a solid block that is then coupled with two optical fibers contained in a unique and simple protective stainless-steel housing. The solid block uses the thermo-optical properties of silicon for temperature measurements, while the deflection of the membrane is used for pressure sensing. An empirically based model combines solid mechanics and optical theory and is in good agreement with experimental measurements. As part of this work, the thermo-optic coefficient (TOC) of the silicon was also investigated theoretically and experimentally. The results show a good agreement between the TOC extracted from the experimental data and such a coefficient in published literature. Furthermore, a novel optical model for the demodulation of the intensity-based pressure-sensing mechanism was developed. This model relates the whole sensor-response profile to the measured parameters and eliminates linear range limitations. By using this model, one can also obtain the initial cavity lengths of an FFPI sensor, which can be very challenging at the microscale. A series of experiments conducted to test the performance of this multi-functional sensor showed that it can easily withstand pressures up to 1,000 psi and temperatures of up to 120°C, where the range of the temperature measurements are restricted only by the fiber optic materials. The developed self-temperature-compensated multi-functional sensor therefore serves as a promising tool in the precise characterization of pressure and temperature in harsh and/or complex environments

Determination of Dioctyl phthalate (DEHP) concentration in polyvinyl chloride (PVC) plastic parts of toothbrushes

Phthalates are chemicals that are widely used in daily products and polyvinyl chloride (PVC) plastics, mainly because of their ability to enhance flexibility and durability. The presence of phthalates in various products contributes to the substantial exposure that all consumers may experience. Continuity in phthalate exposure has several side effects, including endocrine disruption, neurological damage, asthma, hormonal imbalances, obesity, infertility, genital defects, and testicular cancer. The purpose of this research experiment was to determine the Dioctyl phthalate (DEHP) concentration of PVC plastic parts of toothbrushes. The method used to conduct the experiment was Gas Chromatography/Mass Spectrometry. The result showed that the phthalate concentration of a Reach adult’s toothbrush and a Equate kid’s toothbrush were 1.97 ppm and 0.56 ppm respectively. Therefore, this study found that considerable concentrations of phthalates were also present in toothbrushes, which, due to consumers’ daily exposure, would certainly have negative effects on users’ health in the long term

Determination of Dioctyl phthalate (DEHP) concentration in polyvinyl chloride (PVC) plastic parts of toothbrushes

Phthalates are chemicals that are widely used in daily products and polyvinyl chloride (PVC) plastics, mainly because of their ability to enhance flexibility and durability. The presence of phthalates in various products contributes to the substantial exposure that all consumers may experience. Continuity in phthalate exposure has several side effects, including endocrine disruption, neurological damage, asthma, hormonal imbalances, obesity, infertility, genital defects, and testicular cancer. The purpose of this research experiment was to determine the Dioctyl phthalate (DEHP) concentration of PVC plastic parts of toothbrushes. The method used to conduct the experiment was Gas Chromatography/Mass Spectrometry. The result showed that the phthalate concentration of a Reach adult’s toothbrush and a Equate kid’s toothbrush were 1.97 ppm and 0.56 ppm respectively. Therefore, this study found that considerable concentrations of phthalates were also present in toothbrushes, which, due to consumers’ daily exposure, would certainly have negative effects on users’ health in the long term

Mid-term follow-up of the transcatheter closure of perimembranous ventricular septal defects in children using the Amplatzer

Background: The ventricular septal defect (VSD) is the most common form of congenital heart defects. The purpose of this study was to evaluate the results of the early complications and mid-term follow-up of the transcatheter closure of the VSD using the Amplatzer VSD Occluder. Methods: Between April 2012 and October 2013, 110 patients underwent the percutaneous closure of the perimembranous VSD. During the procedure, the size and type of the VSD were obtained via ventriculography. A device at least 2 mm larger than the VSD diameter measured via ventriculography was deployed. The size of the VSD, size of the Amplatzer, and devicesize to VSD-size ratio were calculated. After the confirmation of the suitable position of the device via echocardiography and left ventriculography, the device was released. Follow-up evaluations were done at discharge as well as at 1, 6, and 12 months and yearly thereafter for the VSD occlusion and complete heart block. Results: The study population comprised 62 females and 48 males. The mean age and weight of the patients at procedure were 4.3 ± 5.6 years (range: 2 to 14) and 14.9 ± 10.8 kg (range: 10 to 43). The average device size was 7.0 ± 2.5 mm (range: 4 to 14). The VSD occlusion rate was 72.8 at the completion of the procedure and rose up to 99.0 during the follow-up. The most serious significant complication was complete atrioventricular block, which was seen in 2 patients. The mean follow-up duration was 10.9 ± 3.6 months. Conclusion: The transcatheter closure of the perimembranous VSD was a safe and effective treatment with excellent closure rates in our study population. This procedure had neither mortality nor serious complications. © 2015, Tehran Heart Center. All right recived