Royan Institute for Reproductive Medicine

Tehrān, Iran

Royan Institute for Reproductive Medicine

Tehrān, Iran

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Abouzaripour M.,Tehran University of Medical Sciences | Pasbakhsh P.,Tehran University of Medical Sciences | Atlasi N.,Tehran University of Medical Sciences | Shahverdi A.H.,Royan Institute for Reproductive Medicine | And 2 more authors.
Cell Journal | Year: 2016

Objective: Bone marrow has recently been recognized as a novel source of stem cells for the treatment of wide range of diseases. A number of studies on murine bone marrow have shown a homogenous population of rare stage-specific embryonic antigen 1 (SSEA-1) positive cells that express markers of pluripotent stem cells. This study focuses on SSEA-1 positive cells isolated from murine bone marrow in an attempt to differentiate them into insulin-secreting cells (ISCs) in order to investigate their differentiation potential for future use in cell therapy. Materials and Methods: This study is an experimental research. Mouse SSEA-1 positive cells were isolated by Magnetic-Activated cell sorting (MACS) followed by characterization with flow cytometry. Induced SSEA-1 positive cells were differentiated into ISCs with specific differentiation media. In order to evaluate differentiation quality and analysis, dithizone (DTZ) staining was use, followed by reverse transcription polymerase chain reaction (RT-PCR), immunocytochemistry and insulin secretion assay. Statistical results were analyzed by one-way ANOVA. Results: The results achieved in this study reveal that mouse bone marrow contains a population of SSEA-1 positive cells that expresses pluripotent stem cells markers such as SSEA-1, octamer-binding transcription factor 4 (OCT-4) detected by immunocytochemistry and C-X-C chemokine receptor type 4 (CXCR4) and stem cell antigen-1 (SCA-1) detected by flow cytometric analysis. SSEA-1 positive cells can differentiate into ISCs cell clusters as evidenced by their DTZ positive staining and expression of genes such as Pdx1 (pancreatic transcription factors), Ngn3 (endocrine progenitor marker), Insulin1 and Insulin2 (pancreaticβ-cell markers). Additionally, our results demonstrate expression of PDX1 and GLUT2 protein and insulin secretion in response to a glucose challenge in the differentiated cells. Conclusion: Our study clearly demonstrates the potential of SSEA-1 positive cells to differentiate into insulin secreting cells in defined culture conditions for clinical applications.


PubMed | Royan Institute for Reproductive Medicine, Tehran University of Medical Sciences and Yasuj University of Medical Sciences
Type: Journal Article | Journal: Cell journal | Year: 2016

Bone marrow has recently been recognized as a novel source of stem cells for the treatment of wide range of diseases. A number of studies on murine bone mar- row have shown a homogenous population of rare stage-specific embryonic antigen 1 (SSEA-1) positive cells that express markers of pluripotent stem cells. This study focuses on SSEA-1 positive cells isolated from murine bone marrow in an attempt to differentiate them into insulin-secreting cells (ISCs) in order to investigate their differentiation potential for future use in cell therapy.This study is an experimental research. Mouse SSEA-1 positive cells were isolated by Magnetic-activated cell sorting (MACS) followed by characteriza- tion with flow cytometry. Induced SSEA-1 positive cells were differentiated into ISCs with specific differentiation media. In order to evaluate differentiation quality and analysis, dithizone (DTZ) staining was use, followed by reverse transcription polymerase chain reaction (RT-PCR), immunocytochemistry and insulin secretion assay. Statistical results were analyzed by one-way ANOVA.The results achieved in this study reveal that mouse bone marrow contains a population of SSEA-1 positive cells that expresses pluripotent stem cells markers such as SSEA-1, octamer-binding transcription factor 4 (OCT-4) detected by immunocytochem- istry and C-X-C chemokine receptor type 4 (CXCR4) and stem cell antigen-1 (SCA-1) detected by flow cytometric analysis. SSEA-1 positive cells can differentiate into ISCs cell clusters as evidenced by their DTZ positive staining and expression of genes such as Pdx1 (pancreatic transcription factors), Ngn3 (endocrine progenitor marker), Insulin1 and Insulin2 (pancreatic-cell markers). Additionally, our results demonstrate expression of Pdx1 and Glut2 protein and insulin secretion in response to a glucose challenge in the differentiated cells.Our study clearly demonstrates the potential of SSEA-1 positive cells to differentiate into insulin secreting cells in defined culture conditions for clinical ap- plications.


Hosseini S.M.,Royan Institute for Biotechnology | Hajian M.,Royan Institute for Biotechnology | Forouzanfar M.,Royan Institute for Biotechnology | Ostadhosseini S.,Royan Institute for Biotechnology | And 8 more authors.
Animal Reproduction Science | Year: 2015

The present study aimed to facilitate widespread application of a previously described manual method of somatic cell nuclear transfer (SCNT) by investigating the effects of demecolcine (a microtubule-depolymerizing chemical), cytochalasin-B (a microfilament-depolymerizing chemical: 2.5. μg/ml for 15. min) and MG-132 (a proteasome inhibitor chemical) on the (i) incidence of cytoplasmic protrusion of MII chromosomes, (ii) improvement of manual oocyte enucleation, and (iii) in vitro and in vivo developmental competence of SCNT embryos in the goat. Following in vitro maturation, around 65% of goat oocytes contained a characteristic cytoplasmic protrusion of MII-chromosomes. Treatment with demecolcine (0.4. μg/ml for 30. min) significantly increased this rate to 92.2 ± 4.5%. Treatment with MG-132 (2. μM for 30. min) could not improve this rate when used alone (61.4 ± 11.5%), but when combined with demecolcine (86.4 ± 8.1%). Treatment with cytochalasin-B completely suppressed this rate whenever used, either alone (7.7 ± 5.1%) or in combination with demecolcine (3.9 ± 1.3%). In a direct comparison, there was no significant difference in quantity and quality of embryos propagated by the manual vs. micromanipulation-based methods of SCNT (cleavage: 85.3 ± 4.5 vs. 89.5 ± 8.9%, blastocyst: 19.5 ± 4.3 vs. 24.3 ± 4.4%, grade 1 and 2 blastocyst: 33.8 ± 7.1 vs. 29.5 ± 6.3%, total cell count: 125 ± 11.1 vs. 122 ± 10.5, respectively). Furthermore, development to live kids at term was not significant between the two SCNT methods. From both technical and economical points of view, the overall in vitro and in vivo efficiency of this manual method of SCNT proved it a simple, fast and efficient alternative for large scale production of cloned goats. © 2015 Elsevier B.V.


Hosseini S.M.,Royan Institute for Biotechnology | Hajian M.,Royan Institute for Biotechnology | Ostadhosseini S.,Royan Institute for Biotechnology | Forouzanfar M.,Royan Institute for Biotechnology | And 7 more authors.
Reproductive BioMedicine Online | Year: 2015

This study compared the efficiency of two embryo culture media (SOF1/SOF2 and G1.2/G2.2) for pre- and post-implantation development of somatic cell nuclear transfer goat embryos derived from non-transgenic and transgenic (for htPA and hrcfIX genes) fibroblasts. Despite similar cleavage rates, G1.2/G2.2 supported significantly higher blastocyst development than SOF1/SOF2 (30-35% versus 21%; P < 0.05), irrespective of cell transgenesis. However, following embryo transfer, pregnancy outcomes (establishment, full-term development and live birth) were all significantly higher (P < 0.05) for embryos developed in SOF1/SOF2 versus G1.2/G2.2. Gene expression profiling of 17 developmentally important genes revealed that: (i) SOX2, FOXD3, IFNT, FZD, FGFR4, ERK1, GCN5, PCAF, BMPR1, SMAD5, ALK4, CDC25 and LIFR were significantly induced in blastocysts developed in SOF1/SOF2 but not G1.2/G2.2; (ii) OCT4, CTNNB and CDX2 were similarly expressed in both groups; and (iii) AKT was significantly higher in G1.2/G2.2 than SOF1/SOF2 (P < 0.05). Following IVF, although blastocyst development in G1.2/G2.2 was significantly higher than SOF1/SOF2 counterparts, the majority of assessed genes were similarly expressed in blastocysts developed in both groups. It was concluded that the long-term programming effects of embryo culture medium and/or embryo production method may irreversibly affect post-implantation development of cloned embryos through defined molecular pathways. © 2015 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.


Bazrgar M.,Royan Institute for Reproductive Medicine | Bazrgar M.,Human Genetic Research Group | Peiravian F.,Islamic Azad University at Kazeroon | Abedpour F.,Islamic Azad University at Kazeroon | Karimi M.,Shiraz University of Medical Sciences
Pediatric Hematology and Oncology | Year: 2011

There are limited studies that have focused on the causes for hospitalization as an indicator of morbidity in patients with β-thalassemia major (BTM). A cross-sectional study was conducted to determine the main causes for hospitalization and death in hospitalized BTM patients in a referral hospital in Shiraz, southern Iran. During a 5-year period, 555 BTM patients were admitted to the hospital, of which 390 (67.7%) were 10 to 20 years of age. The most frequent causes for hospitalization were splenectomy (23%), heart failure (22.6%), liver biopsy (22.2%), uncontrolled diabetes (10.9%), arrhythmia (7.2%), cholecystectomy (3.8%), hypoparathyroidism (2.1%), and sepsis (2%). Of the hospitalized patients, 65 (11%), with a mean age of 16.1 ±Â± 4.2 years, died. The most common causes of death were cardiomyopathy (72.3%), infections (17%), malignancies (3.1%), and cerebrovascular accidents (3.1%). Survival of our patients was less than in developed countries and cardiac complications were the most common cause of mortality and morbidity in these patients. Regarding the key role of iron chelation in prevention of different complications in BTM, correction of iron chelation regimen should be well considered. © 2011 Informa Healthcare USA, Inc.


PubMed | Royan Institute for Reproductive Medicine
Type: Journal Article | Journal: Pediatric hematology and oncology | Year: 2011

There are limited studies that have focused on the causes for hospitalization as an indicator of morbidity in patients with -thalassemia major (BTM). A cross-sectional study was conducted to determine the main causes for hospitalization and death in hospitalized BTM patients in a referral hospital in Shiraz, southern Iran. During a 5-year period, 555 BTM patients were admitted to the hospital, of which 390 (67.7%) were 10 to 20 years of age. The most frequent causes for hospitalization were splenectomy (23%), heart failure (22.6%), liver biopsy (22.2%), uncontrolled diabetes (10.9%), arrhythmia (7.2%), cholecystectomy (3.8%), hypoparathyroidism (2.1%), and sepsis (2%). Of the hospitalized patients, 65 (11%), with a mean age of 16.1 4.2 years, died. The most common causes of death were cardiomyopathy (72.3%), infections (17%), malignancies (3.1%), and cerebrovascular accidents (3.1%). Survival of our patients was less than in developed countries and cardiac complications were the most common cause of mortality and morbidity in these patients. Regarding the key role of iron chelation in prevention of different complications in BTM, correction of iron chelation regimen should be well considered.


Jafarpour F.,Royan Institute for Animal Biotechnology | Hosseini S.M.,Royan Institute for Animal Biotechnology | Hajian M.,Royan Institute for Animal Biotechnology | Forouzanfar M.,Islamic Azad University at Marvdasht | And 8 more authors.
Cellular Reprogramming | Year: 2011

5-Aza-2′-deoxycytidine (AzC), trichostatin A (TSA), and its natural mimetic, sodium butyrate (NaB), are antineoplastic drugs that can modify the epigenetic status of donor cells prior to somatic cell nuclear transfer (SCNT). In this study, we used fibroblast cells treated with these drugs to investigate the direct and indirect effects of induced changes in DNA methylation and acetylation of the lysine 9 residue of histone H3 (H3K9). Additionally, we assayed cellular characteristics (cell growth, cell proliferation, cell cycle progression, and apoptosis) and SCNT efficiency in response to these drugs as well as monitoring these effects 24h after removing the drugs. We observed the following: (1) AzC, TSA, and NaB all showed dose-dependent effects on different cellular characteristics; (2) TSA and NaB induced H3K9 hyperacetylation accompanied by DNA hypermethylation, whereas AzC induced DNA hypomethylation with no effect on H3K9 hyperacetylation; (3) TSA and NaB improved cloning efficiency, whereas AzC reduced it; and (4) unlike AzC, the effects of TSA and NaB on cellular characteristics and SCNT efficiency were reversed following drug removal. Our results indicate that somatic cells treated with TSA and NaB show better survival and recovery rates following the removal of these drugs. Moreover, H3K9 hyperacetylation (induced with TSA and NaB), but not DNA hypomethylation (induced with AzC), favors cloning efficiency. © 2011 Mary Ann Liebert, Inc.


PubMed | Royan Institute for Reproductive Medicine
Type: Journal Article | Journal: Theriogenology | Year: 2015

Frozen-thawed rooster semen is not reliable for use in artificial insemination in commercial stocks. Low-density lipoprotein (LDL) has been assessed for effectiveness as a cryoprotectant in the extender to improve the quality of frozen-thawed rooster semen. Although LDL has been evaluated in a few studies in other species for semen cryopreservation, so far no study has been conducted to examine this cryoprotectant for cryopreservation of fowl semen. Thus, this study aims to analyze the effects of different concentrations of LDL (0%, 2%, 4%, 6%, and 8%) in a Beltsville extender for cryopreservation of rooster spermatozoa. In experiment 1, motion parameters, membrane integrity, acrosome integrity, apoptosis status, and mitochondria activity were assessed after freeze-thawing. The highest quality frozen-thawed semen was selected to be used for evaluation of the fertility rate in experiment 2. Semen was collected from six roosters, twice weekly, then extended in a Beltsville extender that contained different concentrations of LDL as follows: 0% (control), 1% (Beltsville plus 1% LDL [BLDL1]), 2% (BLDL2), 4% (BLDL4), 6% (BLDL6), and 8% (BLDL8). Supplementation of the Beltsville extender with 4% LDL produced the most significant percentage of motility (43.1 1.3), membrane integrity (59.4 2.1),mitochondria activity (49.1 1.19), and viable spermatozoa (45 2.28) compared with the control treatment with the results of 22.7 1.3 (motility), 38.4 2.1 (membrane integrity), 40.25 1.19 (mitochondrial activity), and 37.8 2.28 (viability). In experiment 2, a significantly higher percentage of fertility rate was observed for frozen-thawed semen in the extender supplemented with 4% LDL (49.5 1.6) compared with the control (29.2 2.9). Progressive motility and acrosome integrity were not affected by LDL levels in the extenders. The results revealed that supplementation of the Beltsville extender with 4% LDL resulted in higher quality of frozen-thawed rooster sperm.

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