An expression system of channelrhodopsin-2 driven by a minimal Arc/Arg3.1 promoter and Tet system was developed in human neuroblastoma cells.

Advances in neuroscience have relied on the development of techniques that examine neuronal cell activities. One major challenge involves the limitations in labeling and controlling neuronal activities relating to the cell's activation state. In this study, the modified human codon-optimized channelrhodopsin-2 photoreceptor hChR2(C128S) was integrated into function with inducible gene expression methods and materials: the Tet system and the highly efficient minimum promoter of Arc/Arg3.1. The system successfully expressed the target fusion gene exclusively in activated SH-SY5Y human neuroblastoma cells while maintaining the essential characteristics of ChR2. The expression of the channelrhodopsin construct was observed, while the expression duration was refined by treatment with doxycycline. The optogenetic construct here tested the application of the minimum Arc/Arg3.1 promoter, an advanced immediate-early gene promoter, for the expression of the channelrhodopsin gene. Along with its noninvasive nature, this expression system promises to serve dual functions as a cell activity indicator and cell actuator, creating the possibility for researchers to precisely label cells according to their activation state and control the activities of specific neuronal cell populations.

Expression of nano-ferritin in neuronal cells encompassed by minimal Arc promoter system.

Genetic engineering for neuronal cell activity labeling and neuronal cell activity modulation are invaluable for elucidating the underlying characteristics of the brain and neurons. In this study, ferritin fusion protein (FFP) was combined with Tet expression construct under a modified immediate-early gene (IEG) Arc/Arg3.1 promoter so-called SARE-ArcMin. This expression system is a neuronal activity-dependent expression module for nano-ferritin, a radio/magnetic wave-sensitive protein well-accepted as a potential recombinant neuronal actuator. The system was characterized in transcriptional and translational levels in human neuroblastoma SH-SY5Y cells. The mRNA and protein expression levels of nano-ferritin were significant in the activated neurons suggesting that the activity dependent expression patterns of the ferritin also acted as a neuronal cell activation indicator. The system sufficed the need for precise neuronal cell activity specific expression and demonstrated a platform that suggested the use of the nano-ferritin for the study of neuronal cells.

Modelling the neuropathology of lysosomal storage disorders through disease-specific human induced pluripotent stem cells.

Mucopolysaccharidosis II (MPS II) is a lysosomal storage disorder (LSD), caused by iduronate 2-sulphatase (IDS) enzyme dysfunction. The neuropathology of the disease is not well understood, although the neural symptoms are currently incurable. MPS II-patient derived iPSC lines were established and differentiated to neuronal lineage. The disease phenotype was confirmed by IDS enzyme and glycosaminoglycan assay. MPS II neuronal precursor cells (NPCs) showed significantly decreased self-renewal capacity, while their cortical neuronal differentiation potential was not affected. Major structural alterations in the ER and Golgi complex, accumulation of storage vacuoles, and increased apoptosis were observed both at protein expression and ultrastructural level in the MPS II neuronal cells, which was more pronounced in GFAP + astrocytes, with increased LAMP2 expression but unchanged in their RAB7 compartment. Based on these finding we hypothesize that lysosomal membrane protein (LMP) carrier vesicles have an initiating role in the formation of storage vacuoles leading to impaired lysosomal function. In conclusion, a novel human MPS II disease model was established for the first time which recapitulates the in vitro neuropathology of the disorder, providing novel information on the disease mechanism which allows better understanding of further lysosomal storage disorders and facilitates drug testing and gene therapy approaches.

Optimisation of electroporation and lipofection protocols to derive the black tiger shrimp cell line (Penaeus monodon).

To achieve in creating permanent shrimp cell lines, cellular arrest of primary cells in the culture is needed to be firstly solved. Considering the insertion of some markers affecting cellular proliferation into primary haemocytes in order to produce the black tiger shrimp cell line and the very low percent of transduced cells previously reported in penaeid shrimps, these paved us the way to set up suitable gene delivery protocols to increase percent of transduced cells in the shrimp as our primary aim. In this study, electroporation and lipofection were used to transfer construct plasmids (pLL3.7 plasmids containing CMV promoters and pGL-IE1-126(A)-EGFP plasmids carrying WSSV IE1 promoters) into primary haemocytes. As it was difficult to distinguish between cells expressing EGFP signal and auto-fluorescence of many dead cells occurred by electroporation during the first 72 h of experiment; so, only lipofection was managed to deliver plasmids into primary cells. Surprisingly, numbers of suspected proliferative cells were derived after electroporation with no insertion of immortalising markers. These cells survived in vitro for up to 45 days with high rate of cell viability, but the number of viable cells decreased throughout the experiment. In addition, these cells expressed genes and proteins closely related to hyaline cells determined using RT-PCR and western blot. For the lipofection experiment, no green fluorescence signal was detected in any primary cell introduced with these plasmids, suggesting that plasmids were not successfully inserted into cells. Also, a number of primary haemocytes had the apoptotic cell death characteristic within 5 days after lipofection. These possibly result from using inappropriate lipofection protocol and chemical substances. In summary, finding out suitable protocols to elevate the percent of transduced cells is still necessary. Additionally, continuous shrimp cell lines would be possibly established by transforming suspected proliferative cells derived from electroporation in this study.

A robust model of natural hepatitis C infection using hepatocyte-like cells derived from human induced pluripotent stem cells as a long-term host.

BACKGROUND: Hepatitis C virus (HCV) could induce chronic liver diseases and hepatocellular carcinoma in human. The use of primary human hepatocyte as a viral host is restrained with the scarcity of tissue supply. A culture model restricted to HCV genotype 2a (JFH-1) has been established using Huh7-derived hepatocyte. Other genotypes including the wild-type virus could not propagate in Huh7, Huh7.5 and Huh7.5.1 cells. METHODS: Functional hepatocyte-like cells (HLCs) were developed from normal human iPS cells as a host for HCV infection. Mature HLCs were identified for selective hepatocyte markers, CYP450s, HCV associated receptors and HCV essential host factors. HLCs were either transfected with JFH-1 HCV RNA or infected with HCV particles derived from patient serum. The enhancing effect of α-tocopherol and the inhibitory effects of INF-α, ribavirin and sofosbuvir to HCV infection were studied. The HCV viral load and HCV RNA were assayed for the infection efficiency. RESULTS: The fully-developed HLCs expressed phase I, II, and III drug-metabolizing enzymes, HCV associated receptors (claudin-1, occludin, CD81, ApoE, ApoB, LDL-R) and HCV essential host factors (miR-122 and SEC14L2) comparable to the primary human hepatocyte. SEC14L2, an α-tocopherol transfer protein, was expressed in HLCs, but not in Huh7 cell, had been implicated in effective HCVser infection. The HLCs permitted not only the replication of HCV RNA, but also the production of HCV particles (HCVcc) released to the culture media. HLCs drove higher propagation of HCVcc derived from JFH-1 than did the classical host Huh7 cells. HLCs infected with either JFH-1 or wild-type HCV expressed HCV core antigen, NS5A, NS5B, NS3 and HCV negative-stand RNA. HLCs allowed entire HCV life cycle derived from either JFH-1, HCVcc or wild-type HCV (genotype 1a, 1b, 3a, 3b, 6f and 6n). Further increasing the HCVser infection in HLCs was achieved by incubating cell with α-tocopherol. The supernatant from infected HLCs could infect both naïve HLC and Huh7 cell. Treating infected HLC with INF-α and ribavirin decreased HCV RNA in both the cellular fraction and the culture medium. The HLCs reacted to HCVcc or wild-type HCV infection by upregulating TNF-α, IL-28B and IL-29. CONCLUSIONS: This robust cell culture model for serum-derived HCV using HLCs as host cells provides a remarkable system for investigating HCV life cycle, HCV-associated hepatocellular carcinoma development and the screening for new anti HCV drugs.

Induced pluripotency enables differentiation of human nullipotent embryonal carcinoma cells N2102Ep.

Embryonal carcinoma (EC) cells, which are considered to be malignant counterparts of embryonic stem cells, comprise the pluripotent stem cell component of teratocarcinomas, a form of testicular germ cell tumors (GCTs). Nevertheless, many established human EC cell lines are nullipotent with limited or no capacity to differentiate under normal circumstances. In this study, we tested whether an over-expression of Yamanaka's reprogramming factors OCT4, SOX2, c-MYC and KLF4 might enable differentiation of the human nullipotent EC cells N2102Ep. Using OCT4 knockdown differentiated N2102Ep cells, we are able to derive reprogrammed N2102Ep cell lines. The induced pluripotency of N2102Ep allows the cells to differentiate toward neural lineage by retinoic acid; the expression of SSEA3 and SSEA4 is down-regulated, whereas that of neural surface markers is up-regulated. Consistent with the up-regulation of neural surface markers, the expression of the master neuroectodermal transcription factor PAX6 is also induced in reprogrammed N2102Ep. We next investigated whether PAX6 might induce spontaneous differentiation of nullipotent stem cells N2102Ep. However, while an ectopic expression of PAX6 promotes differentiation of NTERA2, it induces cell death in N2102Ep. We nevertheless find that upon induction of retinoic acid, the reprogrammed N2102Ep cells form mature neuronal morphology similar to differentiated pluripotent stem cells NTERA2 as determined by TUJ1 expression, which is absent in N2102Ep parental cells. Altogether, we conclude that the nullipotent state of human EC cells can be reprogrammed to acquire a more relaxed state of differentiation potential by Yamanaka's factors.

Generation of human induced pluripotent stem cell line (MUi033-A) from a male with homozygous for Hemoglobin E.

Hemoglobin E (HbE), a common variant in Southeast Asian populations, results from a G to A substitution at codon 26 of the HBB gene, causing abnormal Hb and mild ß-thalassemia-like symptoms. Here, we derived an induced pluripotent stem cell (iPSC) line, named MUi033-A, from a male homozygous for HbE. The iPSC line demonstrates a normal karyotype and embryonic stem cell-like properties including pluripotency gene expression, and tri-lineage differentiation potential. This iPSC resource holds the potential for investigating gene therapy targeting HbE mutation.

BDNF-, IGF-1- and GDNF-secreting human neural progenitor cells rescue amyloid ß-induced toxicity in cultured rat septal neurons.

Alzheimer's disease (AD) is characterized by the depositions of amyloid-ß (Aß) proteins, resulting in a reduction of choline acetyltransferase (ChAT) activity of AD brain in the early stages of the disease. Several growth factors, including brain-derived neurotrophic factor (BDNF), insulin-like growth factor (IGF)-1 and glial cell-derived neurotrophic factor (GDNF) are known to protect neuronal cell death in several neurodegenerative both in vitro and in vivo models. In this study, septal neurons were prepared from septal nucleus of embryonic (day 16-17) rat brain and treated with monomeric, oligomeric or fibrillar Aß(1-42) peptide. Oligomeric Aß(1-42), (10 µM) was the most potent at sublethal dose. Septal neuron cultures treated with BDNF, IGF-1 or GDNF or co-cultured with genetically modified human neural progenitor cells (hNPCs) secreting these neurotrophic factors (but not allowing contact between the two cell types), were protected from oligomeric Aß(1-42) peptide-induced cell death, and these trophic factors enhanced cholinergic functions by increasing ChAT expression level. These results indicate the potential of employing transplanted hNPCs for treatment of AD.

Establishment and Characterization of MUi027-A: A Novel Patient-Derived Cell Line of Polycystic Kidney Disease with PKD1 Mutation.

Autosomal dominant polycystic kidney disease (ADPKD) is one of the most prevalent genetic diseases affecting the kidneys. A genetically specific mutation model is required to comprehend its pathophysiology and to develop a drug treatment. In this study, we successfully developed human induced pluripotent stem cells (hiPSCs) named MUi027-A from skin fibroblasts of a patient diagnosed with ADPKD and carrying the PKD1 frameshift mutation (c.7946_7947delCT). MUi027-A cells showed the same genetic fingerprints as the parental cells, including the presence of the PKD1 mutation. MUi027-A hiPSCs displayed embryonic stem cell-like characteristics with the capability of differentiating into the three germ layers. Upon directed differentiation, MUi027-A hiPSCs could be differentiated into tubular organoids with the expression of renal cell markers. Furthermore, we compared the efficiency of cyst formation in two human iPSC lines with different PKD1 mutations. When cyst formation was induced by either forskolin or blebbistatin, MUi027-A hiPSC-derived kidney organoids displayed higher frequencies of cyst formation when compared to organoids generated from an iPSC cell line with non-truncating PKD1 mutation genotype (c.5878C > T), suggesting the presence of physiological differences in the mechanism of cyst formation between different PKD1 mutants. Overall, we generated and characterized a novel human iPSC line with a specific PKD mutation and demonstrated its potential as a disease model to study the pathophysiology of genetic determinants in the development of ADPKD disease.

Generation of a human induced pluripotent stem cells (MUi015-A) from skin fibroblast of retinoblastoma patient with 13q deletion syndrome.

The 13q deletion syndrome is a rare chromosomal disorder caused by loss of the long arm of chromosome 13, and usually entails developmental delay, intellectual disability, behavioral problems and distinctive facial features. In this study, we successfully generated a human iPSC line (MUi015-A) from skin fibroblasts of a patient who had large deletion of chromosome 13, del(13)(q14q22). The MUi015-A line exhibited embryonic stem cell characteristics with consistent pluripotency marker expression and the capability of differentiating into three germ layers. The cell line provides a good tool in studying pathophysiology of the tumors, drug testing and gene therapy.

Amelioration of osteogenesis in iPSC-derived mesenchymal stem cells from osteogenesis imperfecta patients by endoplasmic reticulum stress inhibitor.

AIM: Osteogenesis imperfecta (OI) is a hereditary connective tissue disorder primarily caused by mutations in COL1A1 or COL1A2, which encode type I collagen. These mutations affect the quantity and/or quality of collagen composition in bones, leading to bone fragility. Currently, there is still a lack of treatment that addresses disease-causing factors due to an insufficient understanding of the pathological mechanisms involved. MAIN METHODS: Induced pluripotent stem cells (iPSCs) were generated from OI patients with glycine substitution mutations in COL1A1 and COL1A2 and developed into mesenchymal stem cells (iPS-MSCs). OI-derived iPS-MSCs underwent in vitro osteogenic induction to study cell growth, osteogenic differentiation capacity, mRNA expression of osteogenic and unfolded protein response (UPR) markers and apoptosis. The effects of 4-phenylbutyric acid (4-PBA) were examined after treatment of OI iPS-MSCs during osteogenesis. KEY FINDINGS: OI-derived iPS-MSCs exhibited decreased cell growth and impaired osteogenic differentiation and collagen expression. Expression of UPR genes was increased, which led to an increase in apoptotic cell death. 4-PBA treatment decreased apoptotic cells and reduced expression of UPR genes, including HSPA5, XBP1, ATF4, DDIT3, and ATF6. Osteogenic phenotypes, including RUNX2, SPP1, BGLAP, and IBPS expression, as well as calcium mineralization, were also improved. SIGNIFICANCE: MSCs differentiated from disease-specific iPSCs have utility as a disease model for identifying disease-specific treatments. In addition, the ER stress-associated UPR could be a pathogenic mechanism associated with OI. Treatment with 4-PBA alleviated OI pathogenesis by attenuating UPR markers and apoptotic cell death.

Brain-derived neurotrophic factor increases cell number of neural progenitor cells derived from human induced pluripotent stem cells.

BACKGROUND: Several pieces of evidence from in vitro studies showed that brain-derived neurotrophic factor (BDNF) promotes proliferation and differentiation of neural stem/progenitor cells (NSCs) into neurons. Moreover, the JAK2 pathway was proposed to be associated with mouse NSC proliferation. BDNF could activate the STAT-3 pathway and induce proliferation in mouse NSCs. However, its effects on proliferation are not fully understood and JAK/STAT pathway was proposed to play a role in this activity. METHODS: In the present study, the effects of BDNF on cell proliferation and neurite outgrowth of Alzheimer's disease (AD) induced pluripotent stem cells (iPSCs)-derived human neural progenitor cells (hNPCs) were examined. Moreover, a specific signal transduction pathway important in cell proliferation was investigated using a JAK2 inhibitor (AG490) to clarify the role of that pathway. RESULTS: The proliferative effect of BDNF was remarkably observed as an increase in Ki-67 positive cells. The cell number of hNPCs was significantly increased after BDNF treatment represented by cellular metabolic activity of the cells measured by MTT assay. This noticeable effect was statistically shown at 20 ng/ml of BDNF treatment. BDNF, however, did not promote neurite outgrowth but increased neuronal cell number. It was found that AG490 suppressed hNPCs proliferation. However, this inhibitor partially decreased BDNF-induced hNPCs proliferation. These results demonstrated the potential role of BDNF for the amelioration of AD through the increase of AD-derived hNPCs number.

Generation of human induced pluripotent stem cell line (MUi026-A) from a patient with autosomal dominant polycystic kidney disease carrying PKD1 point mutation.

Autosomal dominant polycystic kidney disease (ADPKD) is one of the common genetic kidney disorders that are caused by mutations in PKD1 or PKD2 gene. In this report, the MUi026-A human induced pluripotent stem cell (hiPSC) line was established from the skin fibroblasts of a female ADPKD patient who had the PKD1 mutation with c.5878C > T. The iPSC line retained normal karyotype. The cells displayed embryonic stem cell-like characteristics with pluripotency marker expression and were able to differentiate into three germ layers.

Overexpression of anti-fibrotic factors ameliorates anti-fibrotic properties of Wharton's jelly derived mesenchymal stem cells under oxidative damage.

Transplantation with Wharton's jelly derived mesenchymal stem cells (WJ-MSCs) showed great benefits for restoring myocardial function. However, the outcome of WJ-MSCs transplantation was unsuccessful due to multiple factors including oxidative damage. The presence of oxidative stress due to myocardium injury influences fibrous tissue formation, which causes disability of cardiac muscle. Hepatocyte growth factor (HGF), insulin-like growth factor (IGF1), and sonic hedgehog (SHH) are well-known master regulators in anti-fibrosis when secreted by WJ-MSCs. They showed a beneficial role in the recovery of cardiac fibrosis after WJ-MSCs transplantation. This study hypothesizes whether the reduction of the anti-fibrosis property in WJ-MSCs from oxidative damage can be recovered by overexpression of the HGF, IGF1, or SHH gene. Overexpression was attained by transfection of WJ-MSCs with pCMV3-HGF, pCMV3-IGF1, or pCMV3-SHH followed by H2O2 exposure and co-culturing with cardiac fibroblasts. Myofibroblast specific markers comprised of alpha-smooth muscle actin (α-SMA) and collagen type 1 (COL1) were evaluated. The WJ-MSCs treated with H2O2 influenced the expression of myofibroblastic markers, whereas the overexpression of HGF, IGF1 or SHH reduced myofibroblastic formation. These results indicate that the oxidative stress impaired anti-fibrotic property of WJ-MSCs, leads to an increase of myofibroblasts. Overexpression of anti-fibrotic genes restored the endogenous HGF, IGF1, and SHH alleviating improvement of cardiac function.

Generation of a human induced pluripotent stem cell line (MUi010-A) from skin fibroblast of patient carrying a c.2104C>T mutation in MYH9 gene.

Mutations in MYH9 gene is one of the major causes of inherited thrombocytopenia resulted from nonfunctional myosin-9 protein. We have generated a human induced pluripotent stem cell line MUi010-A from skin fibroblasts of a patient who had a point mutation c.2104C>T (p.R702C) in the exon 16 of MYH9 gene using a non-integrative reprogramming method. The MUi010-A exhibited embryonic stem cell-like characteristics with consistent pluripotent markers expression, was capable of all three embryonic germ layers differentiation, and had a normal karyotype.

Potential effects and molecular mechanisms of melatonin on the dopaminergic neuronal differentiation of human amniotic fluid mesenchymal stem cells.

Melatonin, a highly lipophilic molecule secreted by the pineal gland in the brain, plays a role in various biological functions. Previous studies reported that melatonin exerts its effect on mesenchymal stem cell (MSC) survival and differentiation into osteogenic- and adipogenic-lineage. However, the effect of melatonin in neurogenic differentiation in amniotic fluid (AF)-MSCs remains to be explored, thus we investigated the potential role of melatonin on dopaminergic neuron differentiation in AF-MSCs. The results showed that various concentrations of melatonin did not affect cell viability and proliferative effects of AF-MSCs. Increases in the levels of neuronal protein marker (ßIII-tubulin) and dopaminergic neuronal markers (tyrosine hydroxylase, TH and NURR1), but decrease in the level of glial fibrillary acidic protein (GFAP), were observed in melatonin-treated AF-MSCs. Melatonin induced alteration in differential expression patterns of mesenchymal stem cell antigens by reducing CD29, CD45, CD73, CD90 and CD105, but no changing CD34 expressing cells. AF-MSCs were sequentially induced in neurobasal medium containing standard inducing cocktails (ST: bFGF, SHH, FGF8, BDNF), 1 µM melatonin, or a combination of ST and melatonin. The levels of TUJ1, TH, MAP2, NURR1 and dopamine transporter (DAT) were significantly increased in all treated groups when compared with control-untreated cells. Pretreated AF-MSCs with non-selective MT1/MT2 receptors antagonist, luzindole and selective MT2 receptor antagonist, 4-P-PDOT diminished melatonin-induced increase in dopaminergic neuronal markers and phosphorylated ERK but did not diminish increase in phosphorylated CaMKII by melatonin. Pretreatment with mitogen-activated protein kinase (MEK) inhibitor, PD98059 and CaMKII inhibitor, KN-93 were able to abolish increase in the levels of dopaminergic markers in melatonin-treated AF-MSCs. These findings suggest that melatonin promotes dopaminergic neuronal differentiation of AF-MSCs possibly via the induction in ERK and CaMKII pathways through melatonin receptor-dependent and -independent mechanisms, respectively.

Generation of induced pluripotent stem cells from peripheral blood CD34+ hematopoietic progenitors of a 31year old healthy woman.

The MUi019-A human induced pluripotent stem cell line was generated from peripheral blood CD34+ hematopoietic progenitors of a healthy woman using a non-integrative reprogramming method. Episomal vectors carrying reprogramming factors OCT4, SOX2, KLF4, L-MYC, LIN28, and shRNA of TP53 and EBNA-1 were delivered using electroporation. The iPSC line can be used as a control in studying disease mechanisms. Furthermore, gene editing approaches can be used to introduce specific mutations into the MUi019-A to model disease while the cell type affected by the disease is inaccessible.

Establishment of MUi009 - A human induced pluripotent stem cells from a 32year old male with homozygous ß°-thalassemia coinherited with heterozygous α-thalassemia 2.

The thalassemias are a group of genetic disorders characterized by a deficiency in the synthesis of globin chains. In this study the MUi009-A human induced pluripotent stem cell line was successfully generated from peripheral blood CD34+ haematopoietic progenitors of a 32year old male who had coinherited a homozygous ß°-thalassemia mutation at codon 41/42 (-TCTT) and a heterozygous α-thalassemia 4.2 deletion. The MUi009-A cell line exhibited embryonic stem cell characteristics with consistent pluripotency marker expression and the capability of differentiating into the three germ layers. The cell line may provide a tool for drug testing and gene therapy studies.

Derivation of the human induced pluripotent stem cell line MUi017-A from a patient with homozygous Hemoglobin Constant Spring.

Hemoglobin Constant Spring (HbCS, HBA2: c.427T>C) is a common nondeletional α-thalassemia resulting from a nucleotide substitution at the termination codon of the HBA2 gene. Homozygosity for HbCS is characterized with mild anemia, jaundice, and splenomegaly. In this study, the human induced pluripotent stem cell line MUi017-A was successfully generated from peripheral blood CD34+ hematopoietic progenitors of a 52year old female with homozygous HbCS. The MUi017-A cell line exhibited embryonic stem cell characteristics with consistent expression of specific pluripotency markers and the capability of differentiating into the three germ layers. The cell line may be used for the disease modeling.

Enhancement of ß-Globin Gene Expression in Thalassemic IVS2-654 Induced Pluripotent Stem Cell-Derived Erythroid Cells by Modified U7 snRNA.

The therapeutic use of patient-specific induced pluripotent stem cells (iPSCs) is emerging as a potential treatment of ß-thalassemia. Ideally, patient-specific iPSCs would be genetically corrected by various approaches to treat ß-thalassemia including lentiviral gene transfer, lentivirus-delivered shRNA, and gene editing. These corrected iPSCs would be subsequently differentiated into hematopoietic stem cells and transplanted back into the same patient. In this article, we present a proof of principle study for disease modeling and screening using iPSCs to test the potential use of the modified U7 small nuclear (sn) RNA to correct a splice defect in IVS2-654 ß-thalassemia. In this case, the aberration results from a mutation in the human ß-globin intron 2 causing an aberrant splicing of ß-globin pre-mRNA and preventing synthesis of functional ß-globin protein. The iPSCs (derived from mesenchymal stromal cells from a patient with IVS2-654 ß-thalassemia/hemoglobin (Hb) E) were transduced with a lentivirus carrying a modified U7 snRNA targeting an IVS2-654 ß-globin pre-mRNA in order to restore the correct splicing. Erythroblasts differentiated from the transduced iPSCs expressed high level of correctly spliced ß-globin mRNA suggesting that the modified U7 snRNA was expressed and mediated splicing correction of IVS2-654 ß-globin pre-mRNA in these cells. Moreover, a less active apoptosis cascade process was observed in the corrected cells at transcription level. This study demonstrated the potential use of a genetically modified U7 snRNA with patient-specific iPSCs for the partial restoration of the aberrant splicing process of ß-thalassemia. Stem Cells Translational Medicine 2017;6:1059-1069.