Habib Rezanejad | Harvard Medical School (original) (raw)

Papers by Habib Rezanejad

Nature Chemical Biology

Regenerating pancreatic β-cells is a potential curative approach for diabetes. We previously iden... more Regenerating pancreatic β-cells is a potential curative approach for diabetes. We previously identified the small molecule CID661578 as a potent inducer of β-cell regeneration, but its target and mechanism of action have remained unknown. We now screened 257 million yeast clones and determined that CID661578 targets MAP kinase-interacting serine/threonine kinase 2 (MNK2), an interaction we genetically validated in vivo. CID661578 increased β-cell neogenesis from ductal cells in zebrafish, neonatal pig islet aggregates and human pancreatic ductal organoids. Mechanistically, we found that CID661578 boosts protein synthesis and regeneration by blocking MNK2 from binding eIF4G in the translation initiation complex at the mRNA cap. Unexpectedly, this blocking activity augmented eIF4E phosphorylation depending on MNK1 and bolstered the interaction between eIF4E and eIF4G, which is necessary for both hypertranslation and β-cell regeneration. Taken together, our findings demonstrate a targe...

Glaucoma remains one of the major causes of blindness in today's world. The progressive fiel... more Glaucoma remains one of the major causes of blindness in today's world. The progressive field of stem cells proposes an exciting potential for discovering novel therapies. Here, we report the development of an easy and high throughput method for differentiation of retinal ganglion cells (RGCs) and bipolar cells from human adipose tissue-derived mesenchymal stem cells (hADSCs) using PAX6 (+5a) gene expression, a master gene in development of the vertebrate visual system. HADSCs were isolated from fat tissues and confirmed by their cell surface markers and differentiation potential into adipocytes and osteocytes lineages. Then, the coding region of human PAX6 (+5a) gene was cloned and lentiviral particles were produced. HADSCs differentiation was characterized by morphological characteristics, qRT-PCR and immunocytochemistry (ICC). The hADSCs were isolated successfully with high yield and purity (99%). After 30 hours post transduction by pLEX-PAX6- pur lentiviral vectors in fibro...

Glaucoma remains one of the major causes of blindness in today's world. The progressive field... more Glaucoma remains one of the major causes of blindness in today's world. The progressive field of stem cells proposes an exciting potential for discovering novel therapies. Here, we report the development of an easy and high throughput method for differentiation of retinal ganglion cells (RGCs) and bipolar cells from human adipose tissue-derived mesenchymal stem cells (hADSCs) using PAX6 (+5a) gene expression, a master gene in development of the vertebrate visual system. HADSCs were isolated from fat tissues and confirmed by their cell surface markers and differentiation potential into adipocytes and osteocytes lineages. Then, the coding region of human PAX6 (+5a) gene was cloned and lentiviral particles were produced. HADSCs differentiation was characterized by morphological characteristics, qRT-PCR and immunocytochemistry (ICC). The hADSCs were isolated successfully with high yield and purity (99%). After 30 hours post transduction by pLEX-PAX6 pur lentiviral vectors in fibrone...

Current Protocols in Cell Biology, 2018

Traditionally, studies on cells and tissues have been performed on isolated primary cells or immo... more Traditionally, studies on cells and tissues have been performed on isolated primary cells or immortalized cell lines by culturing them in 2D culture dishes or flasks. However, a caveat of 2D culture is that the cells poorly recapitulate their in vivo counterparts mainly due to the lack of 3D cell-cell and cell-extracellular matrix interactions. In recent years, the development of in vitro organoids as a 3D culture has gained substantial attention as a model to study different tissues. In adults, pancreatic ductal cells have been considered as a source of stem or progenitor cells, thus developing new methods to study ductal cells would be useful. Here, we explain a protocol to isolate mouse pancreatic ductal cells and a cost-effective protocol to generate 3D organoid structures from such ductal cells. Finally, we have devised a protocol for immunostaining of intact whole-organoids without sectioning.

Diabetes, Jul 6, 2018

Previously we showed thyroid hormone (T) enhanced β-cell functional maturation through induction ... more Previously we showed thyroid hormone (T) enhanced β-cell functional maturation through induction of High levels of T have been linked to decreased lifespan in mammals, and low levels to lengthened lifespan, suggesting a relationship between thyroid hormone and aging. Here we show T increased (a β-cell senescence marker and effector) mRNA in rodent and human β-cells. The kinetics of and induction suggested both genes as targets of thyroid hormone via TH receptor (THR) binding to specific response elements (TRE). Using specific agonists CO23 and GC1, we show expression was controlled by THRA and by THRB. Using chromatin immunoprecipitation and a transient transfection yielding biotinylated THRB1 or THRA isoforms to achieve specificity, we determined that THRA isoform bound to whereas THRB1 bound to but not to On a cellular level, T treatment accelerated cell senescence as shown by increased number of β-cells with acidic β-galactosidase activity. Our data show T can simultaneously indu...

Cell Therapy and Regenerative Medicine Journal, 2016

Stem cells are considered as the promising candidates to regenerate damaged tissues. A number of ... more Stem cells are considered as the promising candidates to regenerate damaged tissues. A number of clinical trials have encouraged the researchers and clinicians to use stem cells especially mesenchymal stem cells (MSCs) due to its several unique properties. These cells, in addition to multilinage differentiation potential and high proliferation capacity, exert paracrine effects via the release of exosomes and microvesicles which are known as the main mediators of cell to cell communication (Figure 1). The concept that lncRNAs might be paracrine or even endocrine effectors, is based on the evidence that they can transport via extracellular vesicles (EVs), i.e. exosomes and microvesicles (MVs) between cells, either within a surrounding tissue or remotely by entering into blood (1, 2). It has been shown that the EVs may contain mRNA, noncoding RNA (microRNA, LncRNA, and etc) and sometimes even DNA, in addition to lipids and proteins; secreting from the stem cells and having the capacity to alter the phenotype and cell fate of neighboring cells to restore injury tissues/organs (2, 3). Recently, Wang et al. have investigated the expression profiles of lncRNAs

Cellular reprogramming, 2012

Pluripotent stem cells (PSCs) have the potential to differentiate into many cell types and theref... more Pluripotent stem cells (PSCs) have the potential to differentiate into many cell types and therefore can be a valuable source for cell therapy. Embryonic stem cells (ESCs), which are derived from the inner cell mass (ICM) of the blastocyst, are representative of PSCs. However, use of these cells has some limitations, especially ethical restrictions and immune response. As a result, researchers have been looking for other cell sources or strategies to overcome these limitations. One kind of cellular reprogramming is the process of guiding mature cells into a state of gene expression similar to PSCs. It has been demonstrated that somatic cells can be reprogrammed by various methods, including somatic cell nuclear transfer (SCNT) and cell fusion with ESCs or treatment with their extracts. This implies that some factors in oocytes and ESCs are able to initiate the reprogramming process. Accordingly, induced pluripotent stem cells (iPSCs) have been derived from somatic cells by ectopic e...

Journal of Inorganic Biochemistry, 2012

In the present study, we report a facile and eco-friendly method for the preparation of a novel s... more In the present study, we report a facile and eco-friendly method for the preparation of a novel silver nanocomposite hydrogel (SNH) based on poly(acrylic acid) (PAA) grafted onto salep as a water soluble polysaccharide backbone. The presence of inorganic silver nanoparticles (nano-Ag) in the hydrogel was confirmed by thermo-gravimetric (TG) analysis. The TEM images illustrated the presence of embedded nano-Ag throughout the hydrogel matrix. In addition, the transmission electron microscopy (TEM) images showed that the formed nano-Ag had an average particle size of 5-10 nm. The potential of obtained SNH was examined for Tetracycline hydrochloride (TH) release in simulated colon conditions. Lastly, the in vitro antibacterial properties of the obtained optimum sample were successfully evaluated against gram-negative and gram-positive bacteria.

Cell and Tissue Research, 2014

The neural retina is subjected to various degenerative conditions. Regenerative stem-cell-based t... more The neural retina is subjected to various degenerative conditions. Regenerative stem-cell-based therapy holds great promise for treating severe retinal degeneration diseases, although many drawbacks remain to be overcome. One important problem is to gain authentically differentiated cells for replacement. Paired box 6 protein (5a) (PAX6 (5a)) is a highly conserved master control gene that has an essential role in the development of the vertebrate visual system. Human adiposetissue-derived stem cell (hADSC) isolation was performed by using fat tissues and was confirmed by the differentiation potential of the cells into adipocytes and osteocytes and by their surface marker profile. The coding region of the human PAX6 (5a) gene isoform was cloned and lentiviral particles were propagated in HEK293T. The differentiation of hADSCs into retinal cells was characterized by morphological characteristics, quantitative real-time reverse transcription plus the polymerase chain reaction (qPCR) and immunocytochemistry (ICC) for some retinal cell-specific and retinal pigmented epithelial (RPE) cell-specific markers. hADSCs were successfully isolated. Flow cytometric analysis of surface markers indicated the high purity (∼97 %) of isolated hADSCs. After 30 h of post-transduction, cells gradually showed the characteristic morphology of neuronal cells and small axon-like processes emerged. qPCR and ICC confirmed the differentiation of some neural retinal cells and RPE cells. Thus, PAX6 (5a) transcription factor expression, together with medium supplemented with fibronectin, is able to induce the differentiation of hADSCs into retinal progenitors, RPE cells and photoreceptors.

ACS Applied Materials & Interfaces, 2012

Poly ((2-dimethylaminoethyl) methacrylate) grafted onto κ-carrageenan (κC-g-PDMA) as a biopolymer... more Poly ((2-dimethylaminoethyl) methacrylate) grafted onto κ-carrageenan (κC-g-PDMA) as a biopolymer was synthesized and applied for surface modification of water-soluble CdTe quantum dots (QDs). The effects of DMA concentration, molar ratio of κC-g-PDMA/CdTe, reaction temperature and time on optical properties of CdTe QDs were investigated via fluorescent (FL) and UV-visible spectra. The results showed that the κC-g-PDMA significantly affects the optical properties of CdTe QDs. The obtained samples were characterized by Fourier transform infrared spectrum (FT-IR), thermogravimetric (TG) analysis, and transmission electron microscopy (TEM). The antibacterial activity, antifungal assays, and cytotoxicity of modified QDs were examined, and a good biocompatibility was observed.

Glaucoma remains one of the major causes of blindness in today's world. The progressive f... more Glaucoma remains one of the major causes of blindness in today's world. The progressive field of stem cell proposes an exciting potential for discovering novel therapies. Here, we report the development of an easy and high throughput method for differentiation of retinal ganglion cells (RGC) and bipolar cells from human adipose tissue-derived mesenchymal stem cells (hADSCs) using PAX6 (+5a) gene expression, a master gene in development of the vertebrate visual system. HADSCs was isolated from fat tissues and confirmed by their surface markers and differentiation potential into adipocytes and osteocytes lineages. Then, the coding region of human PAX6 (+5a) gene was cloned and lentiviral particles were produced. HADSCs differentiation was characterized by morphological characteristics, qRT-PCR and immunocytochemistry (ICC). The hADSCs were isolated successfully with high yield and purity (99%). After 30 hours post transduction by pLEX-pax6- pur lentiviral vectors in fibronectin supplemented medium, cells gradually showed the characteristic morphology of neuronal cells. QRT- PCR and ICC confirmed deriving of mainly RGC and marginally bipolar cells. The current investigation demonstrates the feasibility of differentiation of RGCs and bipolar cells from hADSCs using expression of PAX6 (+5a) in the medium supplemented by fibronectin.

Nature Communications

Diabetes can be caused by an insufficiency in β-cell mass. Here, we performed a genetic screen in... more Diabetes can be caused by an insufficiency in β-cell mass. Here, we performed a genetic screen in a zebrafish model of β-cell loss to identify pathways promoting β-cell regeneration. We found that both folate receptor 1 (folr1) overexpression and treatment with folinic acid, stimulated β-cell differentiation in zebrafish. Treatment with folinic acid also stimulated β-cell differentiation in cultures of neonatal pig islets, showing that the effect could be translated to a mammalian system. In both zebrafish and neonatal pig islets, the increased β-cell differentiation originated from ductal cells. Mechanistically, comparative metabolomic analysis of zebrafish with/without β-cell ablation and with/without folinic acid treatment indicated β-cell regeneration could be attributed to changes in the pyrimidine, carnitine, and serine pathways. Overall, our results suggest evolutionarily conserved and previously unknown roles for folic acid and one-carbon metabolism in the generation of β-ce...

Stem cell reports, Jan 15, 2018

Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic gr... more Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic growth and regeneration. However, genetic lineage-tracing experiments with pancreatic duct-specific Cre expression have given conflicting results. Using immunofluorescence and flow cytometry, we show heterogeneous expression of both HNF1β and SOX9 in adult human and murine ductal epithelium. Their expression was dynamic and diminished significantly after induced replication. Purified pancreatic duct cells formed organoid structures in 3D culture, and heterogeneity of expression of Hnf1β and Sox9 was maintained even after passaging. Using antibodies against a second cell surface molecule CD51 (human) or CD24 (mouse), we could isolate living subpopulations of duct cells enriched for high or low expression of HNF1β and SOX9. Only the CD24(Hnfβ/Sox9) subpopulation was able to form organoids.

Nature Communications, 2021

Diabetes can be caused by an insufficiency in β-cell mass. Here, we performed a genetic screen in... more Diabetes can be caused by an insufficiency in β-cell mass. Here, we performed a genetic screen in a zebrafish model of β-cell loss to identify pathways promoting β-cell regeneration. We found that both folate receptor 1 (folr1) overexpression and treatment with folinic acid, stimulated β-cell differentiation in zebrafish. Treatment with folinic acid also stimulated β-cell differentiation in cultures of neonatal pig islets, showing that the effect could be translated to a mammalian system. In both zebrafish and neonatal pig islets, the increased β-cell differentiation originated from ductal cells. Mechanistically, comparative metabolomic analysis of zebrafish with/without β-cell ablation and with/without folinic acid treatment indicated β-cell regeneration could be attributed to changes in the pyrimidine, carnitine, and serine pathways. Overall, our results suggest evolutionarily conserved and previously unknown roles for folic acid and one-carbon metabolism in the generation of β-cells.

Curr Protoc Cell Biol, 2018

Traditionally, studies of cells and tissues have been performed on isolated primary cells or immo... more Traditionally, studies of cells and tissues have been performed on isolated primary cells or immortalized cell lines by culturing them in 2D culture dishes or flasks. However, a caveat regarding 2D culture is that the cells poorly recapitulate their in vivo counterparts, mainly due to a lack of 3D cell-cell and cell-extracellular matrix interactions. In recent years, the development of in vitro organoids as 3D culture has gained substantial attention as a model to study different tissues. In adults, pancreatic ductal cells are considered as a source of stem or progenitor cells, so developing new methods to study ductal cells would be useful. Here, we provide a protocol to isolate mouse pancreatic ductal cells and a cost-effective protocol to generate 3D organoid structures from such ductal cells. Additionally, we have devised a protocol for immunostaining of intact whole organoids without sectioning.

Diabetes, 2018

Previously, we showed that thyroid hormone (TH) triiodothyronine (T3) enhanced β-cell functional ... more Previously, we showed that thyroid hormone (TH) triiodothyronine (T3) enhanced β-cell functional maturation through induction of Mafa High levels of T3 have been linked to decreased life span in mammals and low levels to lengthened life span, suggesting a relationship between TH and aging. Here, we show that T3 increased p16Ink4a (a β-cell senescence marker and effector) mRNA in rodent and human β-cells. The kinetics of Mafa and p16Ink4a induction suggested both genes as targets of TH via TH receptors (THRs) binding to specific response elements. Using specific agonists CO23 and GC1, we showed that p16Ink4a expression was controlled by THRA and Mafa by THRB. Using chromatin immunoprecipitation and a transient transfection yielding biotinylated THRB1 or THRA isoforms to achieve specificity, we determined that THRA isoform bound to p16Ink4a , whereas THRB1 bound to Mafa but not to p16Ink4a On a cellular level, T3 treatment accelerated cell senescence as shown by increased number of β-cells with acidic β-galactosidase activity. Our data show that T3 can simultaneously induce both maturation (Mafa) and aging (p16Ink4a ) effectors and that these dichotomous effects are mediated through different THR isoforms. These findings may be important for further improving stem cell differentiation protocols to produce functional β-cells for replacement therapies in diabetes.

Stem Cell Reports, 2018

Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic gr... more Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic growth and regeneration. However, genetic
lineage-tracing experiments with pancreatic duct-specific Cre expression have given conflicting results. Using immunofluorescence and
flow cytometry, we show heterogeneous expression of both HNF1b and SOX9 in adult human and murine ductal epithelium. Their
expression was dynamic and diminished significantly after induced replication. Purified pancreatic duct cells formed organoid structures
in 3D culture, and heterogeneity of expression of Hnf1b and Sox9 was maintained even after passaging. Using antibodies against a second
cell surface molecule CD51 (human) or CD24 (mouse), we could isolate living subpopulations of duct cells enriched for high or low
expression of HNF1b and SOX9. Only the CD24high (Hnfb high/Sox9 high) subpopulation was able to form organoids.

Previously we showed thyroid hormone (T 3 ) enhanced β-cell functional maturation through inducti... more Previously we showed thyroid hormone (T 3 ) enhanced β-cell functional maturation through induction of Mafa. High levels of T 3 have been linked to decreased lifespan in mammals, and low levels to lengthened lifespan, suggesting a relationship between thyroid hormone and aging.

SUMMARY Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancr... more SUMMARY Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic growth and regeneration. However, genetic lineage-tracing experiments with pancreatic duct-specific Cre expression have given conflicting results. Using immunofluorescence and flow cytometry, we show heterogeneous expression of both HNF1b and SOX9 in adult human and murine ductal epithelium. Their expression was dynamic and diminished significantly after induced replication. Purified pancreatic duct cells formed organoid structures in 3D culture, and heterogeneity of expression of Hnf1b and Sox9 was maintained even after passaging. Using antibodies against a second cell surface molecule CD51 (human) or CD24 (mouse), we could isolate living subpopulations of duct cells enriched for high or low expression of HNF1b and SOX9. Only the CD24 high (Hnfb high /Sox9 high) subpopulation was able to form organoids.

Nature Chemical Biology

Regenerating pancreatic β-cells is a potential curative approach for diabetes. We previously iden... more Regenerating pancreatic β-cells is a potential curative approach for diabetes. We previously identified the small molecule CID661578 as a potent inducer of β-cell regeneration, but its target and mechanism of action have remained unknown. We now screened 257 million yeast clones and determined that CID661578 targets MAP kinase-interacting serine/threonine kinase 2 (MNK2), an interaction we genetically validated in vivo. CID661578 increased β-cell neogenesis from ductal cells in zebrafish, neonatal pig islet aggregates and human pancreatic ductal organoids. Mechanistically, we found that CID661578 boosts protein synthesis and regeneration by blocking MNK2 from binding eIF4G in the translation initiation complex at the mRNA cap. Unexpectedly, this blocking activity augmented eIF4E phosphorylation depending on MNK1 and bolstered the interaction between eIF4E and eIF4G, which is necessary for both hypertranslation and β-cell regeneration. Taken together, our findings demonstrate a targe...

Glaucoma remains one of the major causes of blindness in today's world. The progressive fiel... more Glaucoma remains one of the major causes of blindness in today's world. The progressive field of stem cells proposes an exciting potential for discovering novel therapies. Here, we report the development of an easy and high throughput method for differentiation of retinal ganglion cells (RGCs) and bipolar cells from human adipose tissue-derived mesenchymal stem cells (hADSCs) using PAX6 (+5a) gene expression, a master gene in development of the vertebrate visual system. HADSCs were isolated from fat tissues and confirmed by their cell surface markers and differentiation potential into adipocytes and osteocytes lineages. Then, the coding region of human PAX6 (+5a) gene was cloned and lentiviral particles were produced. HADSCs differentiation was characterized by morphological characteristics, qRT-PCR and immunocytochemistry (ICC). The hADSCs were isolated successfully with high yield and purity (99%). After 30 hours post transduction by pLEX-PAX6- pur lentiviral vectors in fibro...

Glaucoma remains one of the major causes of blindness in today's world. The progressive field... more Glaucoma remains one of the major causes of blindness in today's world. The progressive field of stem cells proposes an exciting potential for discovering novel therapies. Here, we report the development of an easy and high throughput method for differentiation of retinal ganglion cells (RGCs) and bipolar cells from human adipose tissue-derived mesenchymal stem cells (hADSCs) using PAX6 (+5a) gene expression, a master gene in development of the vertebrate visual system. HADSCs were isolated from fat tissues and confirmed by their cell surface markers and differentiation potential into adipocytes and osteocytes lineages. Then, the coding region of human PAX6 (+5a) gene was cloned and lentiviral particles were produced. HADSCs differentiation was characterized by morphological characteristics, qRT-PCR and immunocytochemistry (ICC). The hADSCs were isolated successfully with high yield and purity (99%). After 30 hours post transduction by pLEX-PAX6 pur lentiviral vectors in fibrone...

Current Protocols in Cell Biology, 2018

Traditionally, studies on cells and tissues have been performed on isolated primary cells or immo... more Traditionally, studies on cells and tissues have been performed on isolated primary cells or immortalized cell lines by culturing them in 2D culture dishes or flasks. However, a caveat of 2D culture is that the cells poorly recapitulate their in vivo counterparts mainly due to the lack of 3D cell-cell and cell-extracellular matrix interactions. In recent years, the development of in vitro organoids as a 3D culture has gained substantial attention as a model to study different tissues. In adults, pancreatic ductal cells have been considered as a source of stem or progenitor cells, thus developing new methods to study ductal cells would be useful. Here, we explain a protocol to isolate mouse pancreatic ductal cells and a cost-effective protocol to generate 3D organoid structures from such ductal cells. Finally, we have devised a protocol for immunostaining of intact whole-organoids without sectioning.

Diabetes, Jul 6, 2018

Previously we showed thyroid hormone (T) enhanced β-cell functional maturation through induction ... more Previously we showed thyroid hormone (T) enhanced β-cell functional maturation through induction of High levels of T have been linked to decreased lifespan in mammals, and low levels to lengthened lifespan, suggesting a relationship between thyroid hormone and aging. Here we show T increased (a β-cell senescence marker and effector) mRNA in rodent and human β-cells. The kinetics of and induction suggested both genes as targets of thyroid hormone via TH receptor (THR) binding to specific response elements (TRE). Using specific agonists CO23 and GC1, we show expression was controlled by THRA and by THRB. Using chromatin immunoprecipitation and a transient transfection yielding biotinylated THRB1 or THRA isoforms to achieve specificity, we determined that THRA isoform bound to whereas THRB1 bound to but not to On a cellular level, T treatment accelerated cell senescence as shown by increased number of β-cells with acidic β-galactosidase activity. Our data show T can simultaneously indu...

Cell Therapy and Regenerative Medicine Journal, 2016

Stem cells are considered as the promising candidates to regenerate damaged tissues. A number of ... more Stem cells are considered as the promising candidates to regenerate damaged tissues. A number of clinical trials have encouraged the researchers and clinicians to use stem cells especially mesenchymal stem cells (MSCs) due to its several unique properties. These cells, in addition to multilinage differentiation potential and high proliferation capacity, exert paracrine effects via the release of exosomes and microvesicles which are known as the main mediators of cell to cell communication (Figure 1). The concept that lncRNAs might be paracrine or even endocrine effectors, is based on the evidence that they can transport via extracellular vesicles (EVs), i.e. exosomes and microvesicles (MVs) between cells, either within a surrounding tissue or remotely by entering into blood (1, 2). It has been shown that the EVs may contain mRNA, noncoding RNA (microRNA, LncRNA, and etc) and sometimes even DNA, in addition to lipids and proteins; secreting from the stem cells and having the capacity to alter the phenotype and cell fate of neighboring cells to restore injury tissues/organs (2, 3). Recently, Wang et al. have investigated the expression profiles of lncRNAs

Cellular reprogramming, 2012

Pluripotent stem cells (PSCs) have the potential to differentiate into many cell types and theref... more Pluripotent stem cells (PSCs) have the potential to differentiate into many cell types and therefore can be a valuable source for cell therapy. Embryonic stem cells (ESCs), which are derived from the inner cell mass (ICM) of the blastocyst, are representative of PSCs. However, use of these cells has some limitations, especially ethical restrictions and immune response. As a result, researchers have been looking for other cell sources or strategies to overcome these limitations. One kind of cellular reprogramming is the process of guiding mature cells into a state of gene expression similar to PSCs. It has been demonstrated that somatic cells can be reprogrammed by various methods, including somatic cell nuclear transfer (SCNT) and cell fusion with ESCs or treatment with their extracts. This implies that some factors in oocytes and ESCs are able to initiate the reprogramming process. Accordingly, induced pluripotent stem cells (iPSCs) have been derived from somatic cells by ectopic e...

Journal of Inorganic Biochemistry, 2012

In the present study, we report a facile and eco-friendly method for the preparation of a novel s... more In the present study, we report a facile and eco-friendly method for the preparation of a novel silver nanocomposite hydrogel (SNH) based on poly(acrylic acid) (PAA) grafted onto salep as a water soluble polysaccharide backbone. The presence of inorganic silver nanoparticles (nano-Ag) in the hydrogel was confirmed by thermo-gravimetric (TG) analysis. The TEM images illustrated the presence of embedded nano-Ag throughout the hydrogel matrix. In addition, the transmission electron microscopy (TEM) images showed that the formed nano-Ag had an average particle size of 5-10 nm. The potential of obtained SNH was examined for Tetracycline hydrochloride (TH) release in simulated colon conditions. Lastly, the in vitro antibacterial properties of the obtained optimum sample were successfully evaluated against gram-negative and gram-positive bacteria.

Cell and Tissue Research, 2014

The neural retina is subjected to various degenerative conditions. Regenerative stem-cell-based t... more The neural retina is subjected to various degenerative conditions. Regenerative stem-cell-based therapy holds great promise for treating severe retinal degeneration diseases, although many drawbacks remain to be overcome. One important problem is to gain authentically differentiated cells for replacement. Paired box 6 protein (5a) (PAX6 (5a)) is a highly conserved master control gene that has an essential role in the development of the vertebrate visual system. Human adiposetissue-derived stem cell (hADSC) isolation was performed by using fat tissues and was confirmed by the differentiation potential of the cells into adipocytes and osteocytes and by their surface marker profile. The coding region of the human PAX6 (5a) gene isoform was cloned and lentiviral particles were propagated in HEK293T. The differentiation of hADSCs into retinal cells was characterized by morphological characteristics, quantitative real-time reverse transcription plus the polymerase chain reaction (qPCR) and immunocytochemistry (ICC) for some retinal cell-specific and retinal pigmented epithelial (RPE) cell-specific markers. hADSCs were successfully isolated. Flow cytometric analysis of surface markers indicated the high purity (∼97 %) of isolated hADSCs. After 30 h of post-transduction, cells gradually showed the characteristic morphology of neuronal cells and small axon-like processes emerged. qPCR and ICC confirmed the differentiation of some neural retinal cells and RPE cells. Thus, PAX6 (5a) transcription factor expression, together with medium supplemented with fibronectin, is able to induce the differentiation of hADSCs into retinal progenitors, RPE cells and photoreceptors.

ACS Applied Materials & Interfaces, 2012

Poly ((2-dimethylaminoethyl) methacrylate) grafted onto κ-carrageenan (κC-g-PDMA) as a biopolymer... more Poly ((2-dimethylaminoethyl) methacrylate) grafted onto κ-carrageenan (κC-g-PDMA) as a biopolymer was synthesized and applied for surface modification of water-soluble CdTe quantum dots (QDs). The effects of DMA concentration, molar ratio of κC-g-PDMA/CdTe, reaction temperature and time on optical properties of CdTe QDs were investigated via fluorescent (FL) and UV-visible spectra. The results showed that the κC-g-PDMA significantly affects the optical properties of CdTe QDs. The obtained samples were characterized by Fourier transform infrared spectrum (FT-IR), thermogravimetric (TG) analysis, and transmission electron microscopy (TEM). The antibacterial activity, antifungal assays, and cytotoxicity of modified QDs were examined, and a good biocompatibility was observed.

Glaucoma remains one of the major causes of blindness in today's world. The progressive f... more Glaucoma remains one of the major causes of blindness in today's world. The progressive field of stem cell proposes an exciting potential for discovering novel therapies. Here, we report the development of an easy and high throughput method for differentiation of retinal ganglion cells (RGC) and bipolar cells from human adipose tissue-derived mesenchymal stem cells (hADSCs) using PAX6 (+5a) gene expression, a master gene in development of the vertebrate visual system. HADSCs was isolated from fat tissues and confirmed by their surface markers and differentiation potential into adipocytes and osteocytes lineages. Then, the coding region of human PAX6 (+5a) gene was cloned and lentiviral particles were produced. HADSCs differentiation was characterized by morphological characteristics, qRT-PCR and immunocytochemistry (ICC). The hADSCs were isolated successfully with high yield and purity (99%). After 30 hours post transduction by pLEX-pax6- pur lentiviral vectors in fibronectin supplemented medium, cells gradually showed the characteristic morphology of neuronal cells. QRT- PCR and ICC confirmed deriving of mainly RGC and marginally bipolar cells. The current investigation demonstrates the feasibility of differentiation of RGCs and bipolar cells from hADSCs using expression of PAX6 (+5a) in the medium supplemented by fibronectin.

Nature Communications

Diabetes can be caused by an insufficiency in β-cell mass. Here, we performed a genetic screen in... more Diabetes can be caused by an insufficiency in β-cell mass. Here, we performed a genetic screen in a zebrafish model of β-cell loss to identify pathways promoting β-cell regeneration. We found that both folate receptor 1 (folr1) overexpression and treatment with folinic acid, stimulated β-cell differentiation in zebrafish. Treatment with folinic acid also stimulated β-cell differentiation in cultures of neonatal pig islets, showing that the effect could be translated to a mammalian system. In both zebrafish and neonatal pig islets, the increased β-cell differentiation originated from ductal cells. Mechanistically, comparative metabolomic analysis of zebrafish with/without β-cell ablation and with/without folinic acid treatment indicated β-cell regeneration could be attributed to changes in the pyrimidine, carnitine, and serine pathways. Overall, our results suggest evolutionarily conserved and previously unknown roles for folic acid and one-carbon metabolism in the generation of β-ce...

Stem cell reports, Jan 15, 2018

Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic gr... more Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic growth and regeneration. However, genetic lineage-tracing experiments with pancreatic duct-specific Cre expression have given conflicting results. Using immunofluorescence and flow cytometry, we show heterogeneous expression of both HNF1β and SOX9 in adult human and murine ductal epithelium. Their expression was dynamic and diminished significantly after induced replication. Purified pancreatic duct cells formed organoid structures in 3D culture, and heterogeneity of expression of Hnf1β and Sox9 was maintained even after passaging. Using antibodies against a second cell surface molecule CD51 (human) or CD24 (mouse), we could isolate living subpopulations of duct cells enriched for high or low expression of HNF1β and SOX9. Only the CD24(Hnfβ/Sox9) subpopulation was able to form organoids.

Nature Communications, 2021

Diabetes can be caused by an insufficiency in β-cell mass. Here, we performed a genetic screen in... more Diabetes can be caused by an insufficiency in β-cell mass. Here, we performed a genetic screen in a zebrafish model of β-cell loss to identify pathways promoting β-cell regeneration. We found that both folate receptor 1 (folr1) overexpression and treatment with folinic acid, stimulated β-cell differentiation in zebrafish. Treatment with folinic acid also stimulated β-cell differentiation in cultures of neonatal pig islets, showing that the effect could be translated to a mammalian system. In both zebrafish and neonatal pig islets, the increased β-cell differentiation originated from ductal cells. Mechanistically, comparative metabolomic analysis of zebrafish with/without β-cell ablation and with/without folinic acid treatment indicated β-cell regeneration could be attributed to changes in the pyrimidine, carnitine, and serine pathways. Overall, our results suggest evolutionarily conserved and previously unknown roles for folic acid and one-carbon metabolism in the generation of β-cells.

Curr Protoc Cell Biol, 2018

Traditionally, studies of cells and tissues have been performed on isolated primary cells or immo... more Traditionally, studies of cells and tissues have been performed on isolated primary cells or immortalized cell lines by culturing them in 2D culture dishes or flasks. However, a caveat regarding 2D culture is that the cells poorly recapitulate their in vivo counterparts, mainly due to a lack of 3D cell-cell and cell-extracellular matrix interactions. In recent years, the development of in vitro organoids as 3D culture has gained substantial attention as a model to study different tissues. In adults, pancreatic ductal cells are considered as a source of stem or progenitor cells, so developing new methods to study ductal cells would be useful. Here, we provide a protocol to isolate mouse pancreatic ductal cells and a cost-effective protocol to generate 3D organoid structures from such ductal cells. Additionally, we have devised a protocol for immunostaining of intact whole organoids without sectioning.

Diabetes, 2018

Previously, we showed that thyroid hormone (TH) triiodothyronine (T3) enhanced β-cell functional ... more Previously, we showed that thyroid hormone (TH) triiodothyronine (T3) enhanced β-cell functional maturation through induction of Mafa High levels of T3 have been linked to decreased life span in mammals and low levels to lengthened life span, suggesting a relationship between TH and aging. Here, we show that T3 increased p16Ink4a (a β-cell senescence marker and effector) mRNA in rodent and human β-cells. The kinetics of Mafa and p16Ink4a induction suggested both genes as targets of TH via TH receptors (THRs) binding to specific response elements. Using specific agonists CO23 and GC1, we showed that p16Ink4a expression was controlled by THRA and Mafa by THRB. Using chromatin immunoprecipitation and a transient transfection yielding biotinylated THRB1 or THRA isoforms to achieve specificity, we determined that THRA isoform bound to p16Ink4a , whereas THRB1 bound to Mafa but not to p16Ink4a On a cellular level, T3 treatment accelerated cell senescence as shown by increased number of β-cells with acidic β-galactosidase activity. Our data show that T3 can simultaneously induce both maturation (Mafa) and aging (p16Ink4a ) effectors and that these dichotomous effects are mediated through different THR isoforms. These findings may be important for further improving stem cell differentiation protocols to produce functional β-cells for replacement therapies in diabetes.

Stem Cell Reports, 2018

Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic gr... more Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic growth and regeneration. However, genetic
lineage-tracing experiments with pancreatic duct-specific Cre expression have given conflicting results. Using immunofluorescence and
flow cytometry, we show heterogeneous expression of both HNF1b and SOX9 in adult human and murine ductal epithelium. Their
expression was dynamic and diminished significantly after induced replication. Purified pancreatic duct cells formed organoid structures
in 3D culture, and heterogeneity of expression of Hnf1b and Sox9 was maintained even after passaging. Using antibodies against a second
cell surface molecule CD51 (human) or CD24 (mouse), we could isolate living subpopulations of duct cells enriched for high or low
expression of HNF1b and SOX9. Only the CD24high (Hnfb high/Sox9 high) subpopulation was able to form organoids.

Previously we showed thyroid hormone (T 3 ) enhanced β-cell functional maturation through inducti... more Previously we showed thyroid hormone (T 3 ) enhanced β-cell functional maturation through induction of Mafa. High levels of T 3 have been linked to decreased lifespan in mammals, and low levels to lengthened lifespan, suggesting a relationship between thyroid hormone and aging.

SUMMARY Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancr... more SUMMARY Pancreatic duct epithelial cells have been suggested as a source of progenitors for pancreatic growth and regeneration. However, genetic lineage-tracing experiments with pancreatic duct-specific Cre expression have given conflicting results. Using immunofluorescence and flow cytometry, we show heterogeneous expression of both HNF1b and SOX9 in adult human and murine ductal epithelium. Their expression was dynamic and diminished significantly after induced replication. Purified pancreatic duct cells formed organoid structures in 3D culture, and heterogeneity of expression of Hnf1b and Sox9 was maintained even after passaging. Using antibodies against a second cell surface molecule CD51 (human) or CD24 (mouse), we could isolate living subpopulations of duct cells enriched for high or low expression of HNF1b and SOX9. Only the CD24 high (Hnfb high /Sox9 high) subpopulation was able to form organoids.