Publications by authors named "Hongguang Huo"

13 Publications

  • Page 1 of 1

Function of human pluripotent stem cell-derived photoreceptor progenitors in blind mice.

Sci Rep 2016 07 13;6:29784. Epub 2016 Jul 13.

Astellas Institute for Regenerative Medicine, 33 Locke Dr, Marlborough, MA 01752, USA.

Photoreceptor degeneration due to retinitis pigmentosa (RP) is a primary cause of inherited retinal blindness. Photoreceptor cell-replacement may hold the potential for repair in a completely degenerate retina by reinstating light sensitive cells to form connections that relay information to downstream retinal layers. This study assessed the therapeutic potential of photoreceptor progenitors derived from human embryonic and induced pluripotent stem cells (ESCs and iPSCs) using a protocol that is suitable for future clinical trials. ESCs and iPSCs were cultured in four specific stages under defined conditions, resulting in generation of a near-homogeneous population of photoreceptor-like progenitors. Following transplantation into mice with end-stage retinal degeneration, these cells differentiated into photoreceptors and formed a cell layer connected with host retinal neurons. Visual function was partially restored in treated animals, as evidenced by two visual behavioral tests. Furthermore, the magnitude of functional improvement was positively correlated with the number of engrafted cells. Similar efficacy was observed using either ESCs or iPSCs as source material. These data validate the potential of human pluripotent stem cells for photoreceptor replacement therapies aimed at photoreceptor regeneration in retinal disease.
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http://dx.doi.org/10.1038/srep29784DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4942817PMC
July 2016

Scalable generation of universal platelets from human induced pluripotent stem cells.

Stem Cell Reports 2014 Nov 16;3(5):817-31. Epub 2014 Oct 16.

Advanced Cell Technology, Marlborough, MA 01752, USA. Electronic address:

Human induced pluripotent stem cells (iPSCs) provide a potentially replenishable source for the production of transfusable platelets. Here, we describe a method to generate megakaryocytes (MKs) and functional platelets from iPSCs in a scalable manner under serum/feeder-free conditions. The method also permits the cryopreservation of MK progenitors, enabling a rapid "surge" capacity when large numbers of platelets are needed. Ultrastructural/morphological analyses show no major differences between iPSC platelets and human blood platelets. iPSC platelets form aggregates, lamellipodia, and filopodia after activation and circulate in macrophage-depleted animals and incorporate into developing mouse thrombi in a manner identical to human platelets. By knocking out the β2-microglobulin gene, we have generated platelets that are negative for the major histocompatibility antigens. The scalable generation of HLA-ABC-negative platelets from a renewable cell source represents an important step toward generating universal platelets for transfusion as well as a potential strategy for the management of platelet refractoriness.
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http://dx.doi.org/10.1016/j.stemcr.2014.09.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4235139PMC
November 2014

Derivation of human embryonic stem cells with NEMO deficiency.

Stem Cell Res 2012 May 4;8(3):410-5. Epub 2012 Jan 4.

Department of Newborn Medicine and Department of Medicine, Division of Genetics, Brigham & Women's Hospital, Boston, MA, USA.

Deficiency of the nuclear factor-kappa-B essential modulator (NEMO) is a rare X-linked disorder that presents in boys as hypohydrotic ectodermal dysplasia with immunodeficiency due to defective nuclear factor-κB activation. Here we report on the generation of 2 human embryonic stem cell lines from discarded in vitro fertilization (IVF) embryos ascertained via preimplantation genetic diagnosis. We have derived two human embryonic stem cell lines that carry a T458G hypomorphic mutation in exon 4 of the NEMO (or IKBKG) gene. One of the lines is diploid male; the other is diploid female but has clonally inactivated the X-chromosome that harbors the wild-type IKBKG gene. We show that both lines are pluripotent, have the capacity to differentiate into hematopoietic progenitors, and have defective inhibitor of nuclear factor kappa-B kinase activity. These NEMO deficiency hES cell lines provide an unlimited source for differentiated cell types and may serve as a unique tool to study NEMO deficiency and potentially lead to the development of new therapies for this disease.
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http://dx.doi.org/10.1016/j.scr.2011.12.007DOI Listing
May 2012

Donor cell type can influence the epigenome and differentiation potential of human induced pluripotent stem cells.

Nat Biotechnol 2011 Nov 27;29(12):1117-9. Epub 2011 Nov 27.

Stem Cell Transplantation Program, Division of Pediatric Hematology/Oncology, Manton Center for Orphan Disease Research, Howard Hughes Medical Institute, Children's Hospital Boston and Dana-Farber Cancer Institute, Boston, Massachusetts, USA.

We compared bona fide human induced pluripotent stem cells (iPSCs) derived from umbilical cord blood (CB) cells and neonatal keratinocytes (K). As a consequence of both incomplete erasure of tissue-specific methylation and aberrant de novo methylation, CB-iPSCs and K-iPSCs were distinct in genome-wide DNA methylation profiles and differentiation potential. Extended passage of some iPSC clones in culture did not improve their epigenetic resemblance to embryonic stem cells, implying that some human iPSCs retain a residual 'epigenetic memory' of their tissue of origin.
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http://dx.doi.org/10.1038/nbt.2052DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3357310PMC
November 2011

Clump passaging and expansion of human embryonic and induced pluripotent stem cells on mouse embryonic fibroblast feeder cells.

Curr Protoc Stem Cell Biol 2010 Aug;Chapter 1:Unit 1C.10

Stem Cell Program, Children's Hospital Boston, Boston, Massachusetts, USA.

The ability of human embryonic stem cells (hESCs) to differentiate into essentially all somatic cell types has made them a valuable tool for studying human development and has positioned them for broad applications in toxicology, regenerative medicine, and drug discovery. This unit describes a protocol for the large-scale expansion and maintenance of hESCs in vitro. hESC cultures must maintain a balance between the cellular states of pluripotency and differentiation; thus, researchers must use care when growing these technically demanding cells. The culture system is based largely on the use of a proprietary serum-replacement product and basic fibroblast growth factor (bFGF), with mouse embryonic fibroblasts as a feeder layer. These conditions provide the basis for relatively inexpensive maintenance and expansion of hESCs, as well as their engineered counterparts, human induced pluripotent stem cells (hiPSCs).
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http://dx.doi.org/10.1002/9780470151808.sc01c10s14DOI Listing
August 2010

Telomere elongation in induced pluripotent stem cells from dyskeratosis congenita patients.

Nature 2010 Mar 17;464(7286):292-6. Epub 2010 Feb 17.

Division of Hematology/Oncology, Children's Hospital Boston, Massachusetts 02115, USA.

Patients with dyskeratosis congenita (DC), a disorder of telomere maintenance, suffer degeneration of multiple tissues. Patient-specific induced pluripotent stem (iPS) cells represent invaluable in vitro models for human degenerative disorders like DC. A cardinal feature of iPS cells is acquisition of indefinite self-renewal capacity, which is accompanied by induction of the telomerase reverse transcriptase gene (TERT). We investigated whether defects in telomerase function would limit derivation and maintenance of iPS cells from patients with DC. Here we show that reprogrammed DC cells overcome a critical limitation in telomerase RNA component (TERC) levels to restore telomere maintenance and self-renewal. We discovered that TERC upregulation is a feature of the pluripotent state, that several telomerase components are targeted by pluripotency-associated transcription factors, and that in autosomal dominant DC, transcriptional silencing accompanies a 3' deletion at the TERC locus. Our results demonstrate that reprogramming restores telomere elongation in DC cells despite genetic lesions affecting telomerase, and show that strategies to increase TERC expression may be therapeutically beneficial in DC patients.
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http://dx.doi.org/10.1038/nature08792DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3058620PMC
March 2010

Live cell imaging distinguishes bona fide human iPS cells from partially reprogrammed cells.

Nat Biotechnol 2009 Nov 11;27(11):1033-7. Epub 2009 Oct 11.

Division of Pediatric Hematology/Oncology, Children's Hospital Boston and Dana-Farber Cancer Institute, Boston, Massachusetts, USA.

Somatic cells can be reprogrammed into induced pluripotent stem (iPS) cells by enforced expression of transcription factors. Using serial live imaging of human fibroblasts undergoing reprogramming, we identified distinct colony types that morphologically resemble embryonic stem (ES) cells yet differ in molecular phenotype and differentiation potential. By analyzing expression of pluripotency markers, methylation at the OCT4 and NANOG promoters and differentiation into teratomas, we determined that only one colony type represents true iPS cells, whereas the others represent reprogramming intermediates. Proviral silencing and expression of TRA-1-60, DNMT3B and REX1 can be used to distinguish the fully reprogrammed state, whereas alkaline phosphatase, SSEA-4, GDF3, hTERT and NANOG are insufficient as markers. We also show that reprogramming using chemically defined medium favors formation of fully reprogrammed over partially reprogrammed colonies. Our data define molecular markers of the fully reprogrammed state and highlight the need for rigorous characterization and standardization of putative iPS cells.
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http://dx.doi.org/10.1038/nbt.1580DOI Listing
November 2009

Generation of induced pluripotent stem cells from human blood.

Blood 2009 May 18;113(22):5476-9. Epub 2009 Mar 18.

Department of Medicine, Division of Pediatric Hematology Oncology, Children's Hospital Boston, Dana-Farber Cancer Institute, Boston, MA 02115, USA.

Human dermal fibroblasts obtained by skin biopsy can be reprogrammed directly to pluripotency by the ectopic expression of defined transcription factors. Here, we describe the derivation of induced pluripotent stem cells from CD34+ mobilized human peripheral blood cells using retroviral transduction of OCT4/SOX2/KLF4/MYC. Blood-derived human induced pluripotent stem cells are indistinguishable from human embryonic stem cells with respect to morphology, expression of surface antigens, and pluripotency-associated transcription factors, DNA methylation status at pluripotent cell-specific genes, and the capacity to differentiate in vitro and in teratomas. The ability to reprogram cells from human blood will allow the generation of patient-specific stem cells for diseases in which the disease-causing somatic mutations are restricted to cells of the hematopoietic lineage.
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http://dx.doi.org/10.1182/blood-2009-02-204800DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2689048PMC
May 2009

Disease-specific induced pluripotent stem cells.

Cell 2008 Sep 7;134(5):877-86. Epub 2008 Aug 7.

Department of Medicine, Division of Pediatric Hematology Oncology, Children's Hospital Boston, and Dana-Farber Cancer Institute, Boston, MA 02115, USA.

Tissue culture of immortal cell strains from diseased patients is an invaluable resource for medical research but is largely limited to tumor cell lines or transformed derivatives of native tissues. Here we describe the generation of induced pluripotent stem (iPS) cells from patients with a variety of genetic diseases with either Mendelian or complex inheritance; these diseases include adenosine deaminase deficiency-related severe combined immunodeficiency (ADA-SCID), Shwachman-Bodian-Diamond syndrome (SBDS), Gaucher disease (GD) type III, Duchenne (DMD) and Becker muscular dystrophy (BMD), Parkinson disease (PD), Huntington disease (HD), juvenile-onset, type 1 diabetes mellitus (JDM), Down syndrome (DS)/trisomy 21, and the carrier state of Lesch-Nyhan syndrome. Such disease-specific stem cells offer an unprecedented opportunity to recapitulate both normal and pathologic human tissue formation in vitro, thereby enabling disease investigation and drug development.
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http://dx.doi.org/10.1016/j.cell.2008.07.041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2633781PMC
September 2008

Generation of human-induced pluripotent stem cells.

Nat Protoc 2008 ;3(7):1180-6

Division of Pediatric Hematology Oncology, Children's Hospital Boston, 300 Longwood Avenue, Boston, Massachusetts 02115, USA.

Pluripotent cells, such as embryonic stem cells, are invaluable tools for research and can potentially serve as a source of cell- and tissue-replacement therapy. Rejection after transplantation of cells and tissue derived from embryonic stem cells is a significant obstacle to their clinical use. Recently, human somatic cells have been reprogrammed directly to pluripotency by ectopic expression of four transcription factors (Oct4, Sox2, Klf4 and Myc) to yield induced pluripotent stem (iPS) cells. Human iPS cells are a potential source of patient-specific pluripotent stem cells that would bypass immune rejection. iPS cells can also be used to study diseases for which there are no adequate human in vitro or animal models. In this protocol, we describe how to establish primary human fibroblasts lines and how to derive iPS cells by retroviral transduction of reprogramming factors. Overall, it takes 2 months to complete reprogramming human primary fibroblasts starting from biopsy.
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http://dx.doi.org/10.1038/nprot.2008.92DOI Listing
September 2008

Derivation and maintenance of human embryonic stem cells from poor-quality in vitro fertilization embryos.

Nat Protoc 2008 ;3(5):923-33

Division of Newborn Medicine, Brigham & Women's Hospital and Children's Hospital Boston, Karp Family Research Building 7214, 300 Longwood Avenue, Boston, Massachusetts 02115, USA.

Human embryonic stem (hES) cells are self-renewing, pluripotent cells that are valuable research tools and hold promise for use in regenerative medicine. Most hES cell lines are derived from cryopreserved human embryos that were created during in vitro fertilization (IVF) and are in excess of clinical need. Embryos that are discarded during the IVF procedure because of poor morphology and a low likelihood for generating viable pregnancies or surviving the cryopreservation process are also a viable source of hES cells. In this protocol, we describe how to derive novel hES cells from discarded poor-quality embryos and how to maintain the hES cell lines.
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http://dx.doi.org/10.1038/nprot.2008.60DOI Listing
August 2008

Human embryonic stem cell derivation from poor-quality embryos.

Nat Biotechnol 2008 Feb 27;26(2):212-4. Epub 2008 Jan 27.

Division of Newborn Medicine, Brigham & Women's Hospital, Boston, Massachusetts 02115, USA.

During in vitro fertilization, embryos deemed clinically useless based on poor morphology are typically discarded. Here we demonstrate a statistical correlation between the developmental stage of such poor-quality embryos and the yield of human embryonic stem (hES) cell lines. Early-arrested or highly fragmented embryos only rarely yield cell lines, whereas those that have achieved blastocyst stage are a robust source of normal hES cells.
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http://dx.doi.org/10.1038/nbt1378DOI Listing
February 2008

Reprogramming of human somatic cells to pluripotency with defined factors.

Nature 2008 Jan 23;451(7175):141-6. Epub 2007 Dec 23.

Division of Pediatric Hematology/Oncology, Children's Hospital Boston and Dana Farber Cancer Institute, Boston, Massachusetts 02115, USA.

Pluripotency pertains to the cells of early embryos that can generate all of the tissues in the organism. Embryonic stem cells are embryo-derived cell lines that retain pluripotency and represent invaluable tools for research into the mechanisms of tissue formation. Recently, murine fibroblasts have been reprogrammed directly to pluripotency by ectopic expression of four transcription factors (Oct4, Sox2, Klf4 and Myc) to yield induced pluripotent stem (iPS) cells. Using these same factors, we have derived iPS cells from fetal, neonatal and adult human primary cells, including dermal fibroblasts isolated from a skin biopsy of a healthy research subject. Human iPS cells resemble embryonic stem cells in morphology and gene expression and in the capacity to form teratomas in immune-deficient mice. These data demonstrate that defined factors can reprogramme human cells to pluripotency, and establish a method whereby patient-specific cells might be established in culture.
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http://dx.doi.org/10.1038/nature06534DOI Listing
January 2008