فهرست مطالب aram mokarizadeh

Recently, prevalence of hepatitis B virus (HBV), and hepatitis C virus (HCV) co-infection with Human immunodeficiency virus (HIV), has dramatically increased worldwide due to their shared routes of transmission. Compared to sporadic infection with HIV, HBV, and HCV, concurrent infection with these agents increases the effects and complications of these viruses. Furthermore, co-infection may also alter therapeutic strategies against HIV. Accordingly, choosing appropriate biomarkers to detect these co -infections is one of the main concerns in the field of diagnostic pathology. Up to now, several markers have been introduced for simultaneous diagnosis of HIV, HBV, and HCV. In this regard, serum adenosine deaminase activity (ADA), Fibro Tests, AST-to-Platelet Ratio Index (APRI), Fibrosis-4, Hyaluronic acid, and micro ribonucleic acids have been investigated as potential biomarkers for diagnosis of HIV-HCV/HBV co-infections. This work summarizes the diagnostic value of current and emerging biomarkers in HIV patients concurrently infected with HBV and HCV.

Drastic pH drop is a common consequence of scaling up a mammalian cell culture process, where it may affect the final performance of cell culture. Although CO2 sparging and base addition are used as common approaches for pH control, these strategies are not necessarily successful in large scale bioreactors due to their effect on osmolality and cell viability. Accordingly, a series of experiments were conducted using an IgG1 producing Chinese Hamster Ovary (CHO-S) cell culture in 30 L bioreactor to assess the efficiency of an alternative strategy in controlling culture pH.

Factors inducing partial pressure of CO2 and lactate accumulation (as the main factors altering culture pH) were assessed by Plackett-Burman design to identify the significant ones. As culture pH directly influences process productivity, protein titer was measured as the response variable. Subsequently, Central Composite Design (CCD) was employed to obtain a model for product titer prediction as a function of individual and interaction effects of significant variables.

The results indicated that the major factor affecting pH is non-efficient CO2 removal. CO2 accumulation was found to be affected by an interaction between agitation speed and overlay air flow rate. Accordingly, after increasing the agitation speed and headspace aeration, the culture pH was successfully maintained in the range of 6.95-7.1, resulting in 51% increase in final product titer. Similar results were obtained during 250 L scale bioreactor culture, indicating the scalability of the approach.

The obtained results showed that pH fluctuations could be effectively controlled by optimizing CO2 stripping

The purification of Schwann cells has proven to be a difficult process, with most methods requiring the use of special equipment. However, obtaining a sufficient number and high purity of Schwann cells is an integral aspect in their use for clinical application. Therefore, the aim of this study was to establish a simple and effective protocol for the isolation and purification of Schwann cells from the sciatic nerve of C57BL/6 mice. Furthermore, we aimed to provide a protocol for the isolation of exosomes from these cells.
To purify Schwann cells, we used a combination of in situ nerve pre-degeneration and fetal bovine serum. To determine the most effective method of cell purification, we treated the culture with varying concentrations of fetal bovine serum and examined which concentration provided the highest Schwann cell purity. Exosomes were then isolated from Schwann cells through a process of repeated centrifugation and filtration steps.
We were able to increase the purified population of Schwann cells from C57BL/6 mice by reducing the concentration of FBS. The purity of Schwann cells at FBS concentrations of 10%, 5%, and 2% were 93.42%, 91.25%, and 97.83%, respectively.
When using a concentration of 2% FBS, we obtained the highest purification yield of Schwann cells. Our protocol does not require special equipment or materials. We have created a protocol that is simple, fast, and safe while providing a high yield of purified Schwann cells. The exosome isolation method described in this paper is an appropriate approach with a high quality and yield.

This study aimed to investigate the maturation and fertilization rates of immature mouse oocytes using Embryonic Stem Cell Conditioned Medium (ESCM).
Germinal Vesicle (GV) stage oocytes were observed in 120 NMRI mice, aged 4-6 weeks. GV oocytes with or without cumulus cells were subjected to IVM in either ESCM, Embryonic Stem Cell Growth Medium (ESGM), or α-minimum essential medium (α-MEM). After recording the Metaphase II (MII) oocyte maturation rate, the oocytes were fertilized in vitro. The fertilization success rate was recorded after 24 hr. The embryos were maintained in potassium Simplex Optimization Medium (KSOM) for 96 hr and allowed to grow until the blastocyst stage. After recording developmental competence, they were transferred into the uteri of pseudopregnant mice and their birth rates were recorded.
No significant difference existed between the maturation rates in α-MEM (68.18%) and ESCM (64.67%; p>0.05), whereas this rate was significantly higher for both α-MEM and ESCM compared to ESGM (32.22%; p0.05), similar birth rate between α-MEM and ESCM (47 vs. 40%).
ESCM is an effective medium for preantral follicle growth, oocyte maturation, and subsequent embryo development.

Brucellosis is one of the most prevalent diseases common between humans and animals. It is also called Malta fever, Undulant fever and Mediterranean fever. This disease is spread by consuming milk and its unpasteurized derivatives. Clinical symptoms of brucellosis in humans are fever, chills, headache, muscular pain, tiredness, loss of appetite, joint pain, weight loss, constipation, sore throat, and dry cough. The present study aimed at surveying the seroprevalence of brucellosis in pregnant women and those women who suffered from spontaneous abortion.

This case- control study was conducted in Sanandaj (Iran) in 2016 and included 2 groups of pregnant women: one group included 160 pregnant women and the other included 160 women who suffered from spontaneous abortion. Then, the participants were asked to fill out the questionnaire. After receiving permission from an obstetrician, a 10-cc blood sample was taken from each person to be used in the Rose Bengal, Wright, 2ME, and Coombs tests. Independent samples t test and Chi-square test were used to analyze the data and compare the groups.

Mean±SD age of women in the case group was 30.9±7.3 years, while it was 27.74±5.41 years in control women. The Rose Bengal, Wright, and 2ME prevalence for both groups was negative, but the Coombs and Wright tests score was 33 (20.6%) in pregnant women and it was 27 (16.9%) in women who experienced spontaneous abortion. No meaningful relationship was observed between spontaneous abortion and brucellosis (p= 0.39).

Even though the present study did not find a meaningful relationship between spontaneous abortion and brucellosis (p=0.39), high brucella seroprevalence rates between both groups of women indicated that screening tests should be considered before gestation as an appropriate therapeutic strategy.

Saccharomyces cerevisiae, a common probiotic, can induce in vitro apoptosis in human cancer cells, which could explain its antitumor activity..
The present study was conducted to investigate the in vitro effects of a cytoplasmic extract from S. cerevisiae on the proliferation and viability of a K562 (chronic myeloid leukemia) cell line..
S. cerevisiae was cultured and then disrupted by sonication. After centrifugation, the harvested supernatant was considered to be a cytoplasmic extract. The protein concentration was determined by the Biuret method and the extract was diluted to concentrations of 500, 1000, and 2000 µg protein/ml. The frequencies of apoptosis and necrosis were assessed in extract-treated K562 cells by electrophoresis to show DNA segmentation and by flow cytometry..
The cytoplasmic extract exhibited a time-dependent antitumor activity. DNA electrophoresis did not reveal apoptosis and necrosis in the treated cells, but the DNA bands were weak. The flow cytometry results indicated the induction of apoptosis as well as necrosis in the K562 cell line and the intensity of apoptosis increased with time..
The cytoplasmic extract of S. cerevisiae investigated here may inhibit cell growth and induce apoptosis of chronic myeloid leukemia cells..

Despite great advances in clarifying the pathogenesis of multiple sclerosis (MS), the exact underlying mechanism has not been definitely established. However, the responsibility of cross-reactive antibodies as the initiating factor in MS pathogenesis is a novel idea. Recently, an antibody against-α-gliadin 33-mer peptide which is found in most patients with gluten sensitivity have shown to cross-react significantly with various neural antigens including asialoganglioside, synapsin, and myelin basic protein (MBP). Furthermore, evidence indicates that IL-17, circulating immune complexes and even antibodies produced during gluten sensitivity can contribute to blood–brain barrier (BBB) permeability. Accordingly, extravasation of these anti-α-gliadin antibodies (AGA; especially IgG isotype) through the impaired BBB thought to target asialoganglioside, synapsin, and MBP in neurons. This opsonization may trigger a series of cascade pathways including complement activation, antibody-dependent microglial cytotoxicity against neurons, secretion of inflammatory mediators, myelin sheath damage, chemokine expression, CNS inflammation, BBB disruption and then leukocyte infiltration. The present hypothesis introduces a new antibody-dependent alternative pathway which may lead to multiple sclerosis (MS) during gluten sensitivity.

Acute treatment with metformin has a cardio-protective effects by suppression of inflammatory responses during myocardial infarction (MI) through activation of AMP-activated protein kinase (AMPK). Neutrophils have a pivotal role during MI-induced inflammatory responses. Some anti-inflammatory treatments have decreased cardiac injury and infarct size in MI. Here we evaluated the effects of chronic pretreatment with metformin on myocardial remodeling and neutrophil recruitment after isoproterenol-induced MI. Male wistar rats were randomly assigned into 6 groups (n=6) of untreated control, sham, isoproterenol (Iso), and pre-treated orally with 25, 50, and 100 mg/kg of metformin, twice daily, for 14 days. Isoproterenol was injected subcutaneously (sc) at 13th and 14th days for induction of acute MI. Histopathological examinations were done on the harvested hearts. Number of neutrophils in peripheral blood and their infiltration to myocardium were evaluated by Gimsa staining and myeloperoxidase (MPO) assay, respectively. Histopathological analysis showed a significant attenuation of isoproterenol induced cardiomyocyte necrosis and fibrosis by all three doses of metformin. The heart to body weight ratio was also decreased with all doses of metformin. Pre-treatment with metformin in comparison to Iso (MI) group reduced peripheral neutrophils (p<0.05, p<0.01, and p 0.001 at 25, 50, and 100 mg/kg; respectively) as well as MPO activity (p<0.05 and p<0.01 at 50 and 100 mg/ kg, respectively). Pre-treatment with metformin decreased post-MI myocardial injuries by reducing cardiac remodeling and myocardial neutrophil activity. The results could be explained as a new mechanism for cardio-protective effect of metformin.

Recently، bone-marrow-derived cells have introduced new therapeutic approaches to the management of wound healing in severe skin injuries. Bone marrow-derived stromal cells are described as a heterogeneous population، including mesenchymal stem cells، hematopoietic stem cells، and fibro-blast cells. Results derived from several studies indicate that these cells may contribute to tissue regeneration whether through producing variety of bioactive growth factors and/or by differentiation into mesoderm lineage. The aim of the present study was to investigate the effect of subcutaneous administration of bone marrow-derived stromal cells in repairing or regeneration of skin wounds induced by third-degree burn in a mouse model. In an experimental study that was performed in Urmia University research center from December 2011 to June 2012، The third-degree skin burn was induced on the shaved backs of healthy 7-8 week old male mice (N=18) using a metal rods heated in boiling water. After 1 hour، based on the equal physical condition mice were randomly divided into two separate groups and then subcutaneously administered with phosphate buffered saline (PBS; 400 µl) or bone marrow-derived stromal cells (106 cell in 400µl PBS) at the burn site. 7، 14 and 21 days after induction of burn injury، biopsies were taken from burn wounds and then the sections were prepared. Subsequently the prepared sections were stained with hematoxylin/eosin and Masson''s trichrome to explore histopathological changes evoke by administration of bone marrow derived stromal cells in comparison with control subjects. Considering investigated parameters including formation of granulation tissue (respectively on days 7، 14 and 21 P≤ 0/007، P≤ 0/0013 and P≤ 0/001)، angiogenesis (on day 21 P≤ 0/002) and collagen deposition، in mice treated with bone marrow-derived stromal cells the rate of healing of third-degree thermal burns was significantly accelerated when compared to the PBS-treated mice. This experimental modulation of wound healing suggests that bone marrow-derived stromal cells can significantly enhance the rate of wound healing possibly through stimulation of granulation tissue، angiogenesis، fibroblast proliferation and collagen deposition.

To investigate the IgG content and its variations in uterine fluid (UF) during the estrous cycle of the cow and to compare them with those of the blood serum (S), six pairs of serum and UF samples for each phase of the cycle selected out of 240 bovine genital tracts and blood samples were collected from Urmia abattoir. The UF samples were collected by gentle scraping of the endometrium using a curette after uterine incision and their IgG content and those of the serum were measured by single radial immuno-diffusion (SRID) assay. Serum IgG values (Mean ± SEM) were generally higher than the UF values throughout the cycle except for di-estrus (S: 38.50 ± 0.90, UF: 51.60 ± 2.10 mg mL-1), in which the highest values were observed in UF samples. In met-estrus the difference was not significant (S: 34.80 ± 1.80mg mL-1, UF: 30.80 ± 5.20 mg mL-1), however, in estrus the mean UF IgG value (12.50 ± 1.10 mg mL-1) was lower than that of the serum (31.30 ± 1.20 mg mL-1). In pro-estrus, the lowest values (S: 27.80 ± 1.30 mg mL-1, UF: 9.10 ± 1.50 mg mL-1) were obtained. The results showed a lower IgG values in the bovine UF than those of the serum in the follicular phase of the cycle, while in di-estrus the UF IgG content was the highest, suggesting some IgG production in the uterus at this phase.

Mesenchymal stem cells (MSCs) are appropriate target for gene and cell-based therapy of hemophilia B patients. MSCs possess several unique properties such as capability of differentiating into multiple lineages and lower immunogenecity in transplant procedure that make them attractive candidates for cell and gene therapy. One of the challenges in the gene therapy is the low expression level of transgene. To improve expression، strong regulatory elements in the context of vectors could contribute to improve efficacy of gene therapy strategies. In this study four human factor IX (hFIX) -expressing plasmids equipped with various combination of human -globin (hBG) introns and Kozak sequence were transfected into the MSCs and expression of the hFIX was evaluated in vitro. MSCs were obtained from tibias and the femora of rats and phenotypic characterization of the MSCs was determined by flow cytometry. Four hFIX-expressing plasmids were introduced into the culture-expanded MSCs using transfection agent. 48 hours after transfection، ability of the MSCs for expression of the hFIX and efficacies of the plasmids were evaluated by performing sandwich ELISA on cultured media as well as semi-quantitative RT-PCR. All analyses were performed with One-way ANOVA using SPSS software. The highest expression level of the hFIX was obtained from intron-less and hBG intron-I containing construct. The highest biological activity was obtained from hBG intron-I،II containing construct. Successful expression of the hFIX was obtained from recombinant MSCs. MSCs were able to splice heterologous hBG intron-I from the hFIX-cDNA. Application of thehBG introns reduced the hFIX expression levels، probably due to improper splicing of the hBG introns.

The initiating event in multiple sclerosis (MS) pathogenesis is not known yet. However, in general, breakdown of the blood-brain barrier (BBB) and subsequent infiltration of immune cells into the central nervous system (CNS) has been thought to be the main initiating event. Nonetheless, the mechanism by which the BBB gets disrupted and allows immune cells to infiltrate into the CNS is not fully understood. Evidence indicates that prior to cellular infiltration, over passing peripherally generated cross-reactive immunoglobulin G (IgG) through the transiently permeable BBB during systemic inflammation, hypoxia, hyperthermia, transient hypertension or acute stresses may cause CNS inflammation, BBB breakdown and then initiation of MS disease. Here, we discuss the possible detailed mechanisms that may be involved in cross-reactive IgG-mediated MS autoimmunity.

Leptin, as a 16 kDa adipokine, is a pleiotropic cytokine-like hormone that primarily secreted from adipose tissue. It also involves in the regulation of energy homeostasis, neuroendocrine function, immunity, lipid and glucose homeostasis, fatty acid oxidation, angiogenesis, puberty and reproduction. The aim of this study was to investigate the effects of in vitro addition of leptin to in vitro maturation (IVM) medium on buffalo oocyte maturation and apoptosis. In this experimental study, Ovaries from apparently normal reproductive organs of slaughtered adult buffaloes (Bubalus bubalis) with unknown breeding history were collected from Urmia Abattoir, Urmia, Iran, and were transported immediately to the laboratory in a thermos flask containing sterile normal saline with added antibiotics. Oocytes were aspirated from 2-8 mm visible follicles of the ovaries using an 18-G needle attached to a 10 ml syringe. IVM medium included tissue culture medium-199 (TCM-199), 10% fetal bovine serum (FBS), 22 μg/ml sodium pyruvate, 0.5 IU/ml ovine follicle-stimulating hormone (oFSH), 0.5 IU/ml ovine luteinizing hormone (oLH), 1 μg/ml oestradiol, 50 μg/ml gentamycin, and leptin [0 (control), 10, 50, and 100 ng/ml]. The good quality buffalo oocytes (batches of 10 oocytes) were placed in a culture plate containing six 50 μl droplets of maturation medium, covered with sterilized mineral oil, and then incubated at 38.5˚C with 5% CO2 in air for 24 hours. The maturation of oocytes was evaluated under a stereomicroscope by detecting the first polar body extrusion of oocytes. FITC-Annexin V - propidium iodide (PI) staining method was used to detect oocyte apoptosis. From a total of 115 collected ovaries, 1100 oocytes were recovered among which 283 oocyte were suitable for IVM. In the groups of leptin treated with 0 (control), 10,and 100 ng/ml, the percentage of oocytes maturation was 74.65, 83.81, 77.85, and 75.40%, while the percentage of oocytes apoptosis was 9.83, 9.54, 9.93, and 10.42%, respectively. Our results showed that addition of 10 ng/ml leptin to buffalo IVM medium increased oocyte maturation, significantly, as compared with that in control group. However, addition of leptin to IVM medium had no significant influence on buffalo oocyte apoptosis. Our findings suggested that addition of 10 ng/ml leptin to IVM medium of buffalo oocyte can improve oocyte nuclear maturation. Furthermore, we showed that there is no relation between in vitro addition of leptin to buffalo oocyte IVM medium and oocyte apoptosis.

Auto-reactive cells-mediated immune responses are responsible for the current tissue damages during autoimmunity. Accordingly، functional modulation of auto-reactive cells has been a pivotal aim in many of recent studies. In the current study، we investigated the possibility for insertion of regulatory molecules onto auto-reactive cells through exosomal nano-shuttles as a novel approach for phenotype modification of auto-reactive cells. The exosomes were isolated from supernatant of mesenchymal stem cells culture. Resultant exosomes co-cultured with lymphocytes were harvested from established EAE mice in the presence of antigenic MOG35-55 peptide. After 24 hr، insertion of exosomal tolerogenic molecules (PD-L1، TGF-β، galectin-1) onto auto-reactive cells were explored through flow cytometry. The potency of exosomal inserted membrane molecules to modulate phenotype of auto-reactive lymphocytes was assessed upon ELISA test for their-derived cytokines IFN-γ and IL-17. Incorporation of exosomal molecules into lymohocytes’ membrane was confirmed by flow cytometric analyses for surface levels of mentioned molecules. Additionally، the decreased secretion of IFN-γ and IL-17 were detected in exosome pre-treated lymphocytes upon stimulation with MOG peptide. Mesenchymal stem cells -derived exosomes showed to be efficient organelles for insertion of bioactive tolerogenic molecules onto auto-reactive cells and modulation of their phenotypes

The aim of the present study was to investigate the effect of in vitro supplementation of selenium on fresh and frozen spermatozoa quality of buffalo (Bubalus bubalis) bulls. Five healthy buffalo bulls (5 ejaculates from each bull) were used. Each ejaculate was diluted at 37 ˚C with tris-based extender containing 0 (control)، 0. 5، 1، 2، 4 and 8 μg mL-1 sodium selenite and the sperm motility and viability were evaluated at 0 (T0) (immediately after dilution)، 60 (T1) and 120 (T2) min after diluting semen. In the second step، semen samples were diluted with tris-egg yolk-glycerol extender containing the same amounts of sodium selenite، cooled to 4 ˚C، equilibrated and semen parameters (motility، viability، membrane integrity and DNA damage) were estimated. Then، the semen was packed in 0. 5 mL French straws and frozen in liquid nitrogen. Later، the semen was thawed and analyzed for the same parameters، as well as total antioxidant capacity. Results showed that addition of 1 and 2 μgmL-1 selenium to the semen extender significantly increased the sperm motility of fresh and equilibrated semen compared to the control without affecting other parameters. However، in frozen-thawed semen، extenders containing 1 and 2 μg mL-1 selenium significantly improved sperm motility، viability، membrane integrity and semen total antioxidant capacity and also resulted in lower DNA damaged sperms. In this study selenium supplementation of semen extender of 4 and 8 μg mL-1 had deleterious effects on sperm parameters as early as the samples were prepared for freezing