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Compositions and methods of generating reprogrammed adipocyte cells and methods of use therefore

a technology of adipocyte cells and adipocytes, which is applied in the direction of optical radiation measurement, chemical vapor deposition coating, fungi, etc., can solve the problems of adverse health effects and mortality, and insufficient methods for preventing or treating obesity and associated metabolic disorders, so as to reduce the probability of developing a disorder

Inactive Publication Date: 2012-08-30
THE GENERAL HOSPITAL CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0087]By “positioned for expression” is meant that the polynucleotide of the invention (e.g., a DNA molecule) is positioned adjacent to a DNA sequence that directs transcription and translation of the sequence (i.e., facilitates the production of, for example, a recombinant polypeptide of the invention, or an RNA molecule).
[0096]As used herein, the terms “prevent,”“preventing,”“prevention,”“prophylactic treatment” and the like refer to reducing the probability of developing a disorder or condition in a subject, who does not have, but is at risk of or susceptible to developing a disorder or condition.

Problems solved by technology

Obesity is a disease in which the natural energy reserve, stored in the adipose tissue of humans and other mammals, is increased to a point where it is associated with adverse health effects and mortality.
Despite recognition of the risks associated with obesity, methods for preventing or treating obesity and associated metabolic disorders are inadequate.
Obesity continues to pose a significant public health problem.
While mouse models for obesity exist, they represent rare single gene deletions that do not adequately recapitulate human disease processes, in part due to differences in mouse and human metabolism.
Research into adipocyte biology has also been limited by the fact that a renewable source of adipocytes is unavailable.
Although human adipose tissue is easily obtained, primary adipocytes are difficult to maintain in culture and are not amenable to expansion.

Method used

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  • Compositions and methods of generating reprogrammed adipocyte cells and methods of use therefore
  • Compositions and methods of generating reprogrammed adipocyte cells and methods of use therefore
  • Compositions and methods of generating reprogrammed adipocyte cells and methods of use therefore

Examples

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example 1

Reprogrammed Adipocyte Cells were Obtained by the Directed Differentiation of Human Embryonic Stem Cells

[0191]Known regulators of adipogenesis were cloned into a doxycycline-inducible lentiviral backbone (e.g. PPARγ2, C / EBPα, C / EBPβ, C / EBPδ, SREBP1c, CREB1, and KROX20) (FIG. 1). To test viral titers human adipocyte-derived mesenchymal stem cells (ADMSCs) and human embryonic stem cells were transduced with enhanced green fluorescent protein (eGFP) and reverse tetracycline-controlled transcriptional activator M2 (rtTA-M2) expressing lentiviruses. Fluoresence microscopy of ADMSCs followed by fluorescence-activated cell sorting (FACS) revealed a transfection efficiency of 93% (FIGS. 2A-2C) with a similar efficiency in human embryonic stem cells.

[0192]The adipogenic potential of lentiviral PPARγ2 overexpression in ADMSCs and human embyronic stem cells was tested. In both cases there was differentiation into adipocytes at high efficiency. Of note, discernible differences in cellular morph...

example 2

Reprogrammed Adipocyte Cells are Useful for Identifying Genetic Alterations Associated with Cardiac Pathology

[0197]The methods of the invention provide for the generation of subject-specific tissues for genetic profiling and other studies through the use of reprogrammed induced pluripotent stem (iPS) cells. iPS cell lines can be generated from patient-derived fibrobasts using methods described by (Park et al., Cell. 2008; 134:877-886; Maherali et al., Cell Stem Cell 3, 340-345). Fibroblasts suitable for reprogramming have been obtained from three different sources: skin punch biopsies, hair follicles, and peripheral blood mononuclear cells (PBMC) fractions from blood draws (FIG. 6B). The cells are transduced with a single doxycycline-inducible lentivirus that transiently expresses all of the four genes needed for fibroblast programming into iPS cells (FIGS. 6A-6D). Vectors and methods for generating induced pluripotent stem cells are known in the art and described, for example, in M...

example 3

PPARγ2 Expression is Sufficient for Adipocyte Differentiation In Vitro from both hEs Cells and ADMSCs

[0202]As described above, human somatic cells can be directly reprogrammed in vitro into a pluripotent embryonic stem cell-like state by introducing a combination of four transcription factors (OCT4, SOX2, and either cMYC and KLF4 or NANOG and LIN28) (Takahashi et al., Nat Protoc, 2007. 2(12):3081-9, Yu et al., Science, 2007. 318(5858): 1917-20, Lowry et al., Proc Natl Acad Sci USA, 2008. 105(8): p. 2883-8). These induced pluripotent stem (iPS) cells are nearly identical to embryonic stem (ES) cells in gene expression, DNA methylation and chromatin modifications (Takahashi et al., Nat Protoc, 2007. 2(12):3081-9, Yu et al., Science, 2007. 318(5858): 1917-20, Maherali supra). Using a retroviral system, human iPS cells have successfully been generated from patients with a variety of genetic diseases, including Parkinson disease, Huntington disease, and type 1 diabetes mellitus. A tetrac...

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Abstract

The invention provides therapeutic compositions comprising reprogrammed adipocyte cells for use as disease models, therapeutic compositions comprising reprogrammed adipocyte cells for the treatment of conditions characterized by a reduction in cell number or tissue mass, and methods of generating such cells.

Description

CROSS-REFERENCE TO RELATED APPLICATION[0001]This application claims the benefit of the following U.S. Provisional Application No. 61 / 100,946, filed Sep. 29, 2008, the entire contents of which are incorporated herein by reference.BACKGROUND OF THE INVENTION[0002]Obesity is a disease in which the natural energy reserve, stored in the adipose tissue of humans and other mammals, is increased to a point where it is associated with adverse health effects and mortality. Obesity is a complex, multi-factorial disease involving environmental, genetic, and behavioral components. It is also the second leading cause of preventable death in the U.S. Adipose tissue secretes exocrine mediators that lead to endothelial dysfunction and atherosclerosis. Increased triglycerides, decreased HDL levels and abnormal LDL composition characterize the primary dyslipidemia related to obesity and no doubt play a major role in the development of atherosclerosis and cardiovascular disease in obese individuals. Ob...

Claims

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Application Information

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IPC IPC(8): A61K35/12C40B30/04C12N1/19C12Q1/54C12N15/63C12N1/21C12N5/077G01N33/566B81C1/00H01L21/306G01J1/02G01P5/12H01L35/00
CPCB81C1/00103B81B2203/0384B81B2203/0353
Inventor LUM, DAVID H.AHFELDT, TIM D.COWAN, CHAD
Owner THE GENERAL HOSPITAL CORP
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