Pluripotent stem cell for treating diabetic skin ulcer

a stem cell and diabetic technology, applied in the field of regenerative medicine, can solve the problems of increasing the number of patients, increasing the risk of chronic skin ulcers, and affecting the survival of patients with diabetes mellitus, and achieve the effect of inhibiting the progression of skin ulcers

Inactive Publication Date: 2017-09-14
THE UNIV OF TOKYO +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0037]The present invention is able to inhibit the progression of skin ulcer and repair skin tissue by a skin tissue regeneration mechanism by which Muse cells diffe...

Problems solved by technology

In Japan, diabetes has become a modern-day national affliction and more than 95% of diabetes patients (which is estimated to exceed 20 million people when including persons at risk to the onset of diabetes) have been diagnosed with non-insulin-dependent diabetes mellitus, and increases in the number of patients is becoming a problem accompanying lifestyle changes.
Many serious complications of diabetes such as heart disease, kidney failure or blindness have an effect on individuals together with diabetes, and complications involving the lower extremities result in the greatest damage.
Patients afflicted with diabetes mellitus are at greater risk to the onset of chronic skin ulcers such as ulcers of the leg and foot accompanying long-term complications.
However, in cases in which microvascular disease (or ischemia) is present, this response is significantly impaired, and this is considered to most likely be important in the etiology of ulcers together with a tendency towards the occurrence of thrombosis in the microcirculation during reduced blood flow.
On the other hand, nerve disorders lack an adequately established treatment method with respect to both the symptomatic treatment thereof and the prevention of the progressive decline of nerve function, and is one of the major complications of diabetes mellitus.
The effects of peripher...

Method used

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  • Pluripotent stem cell for treating diabetic skin ulcer
  • Pluripotent stem cell for treating diabetic skin ulcer
  • Pluripotent stem cell for treating diabetic skin ulcer

Examples

Experimental program
Comparison scheme
Effect test

example 1

Preparation of Human Muse Cells

[0068](1) Sampling of Human Tissue and Cell Preparation

[0069]Lipoaspirates were acquired by liposuction surgery from the abdomens and / or thighs of five non-obese women (age: 26.6±8.7 years, BMI: 21.5±2.0) from whom consent was obtained with the approval of the ethics committee of the Graduate School of Medicine and Faculty of Medicine of the University of Tokyo. Stromal vascular fractions (SVF) containing adipose-derived stromal / stem cells (ASC) were isolated from the aspirated adipose tissue as previously described (see Yoshimura, K. et al., J. Cell Physiol., Vol. 208, p. 64-76 (2006)). Simply put, aspirated adipose tissue was washed with PBS and digested in PBS containing 0.075% collagenase for 30 minutes at 37° C. on a shaker. Mature adipocytes and connective tissue were separated from the pellet by centrifugal separation. The cell pellet was then re-suspended and lysed by passing through 100 μm, 70 μm and 40 μm screens. The cell pellet containing a...

example 2

Production of Immunosuppressed Diabetic Mice Model

[0072]Five-week-old severe combined immunodeficient (SCID) mice (C. B17 / Icr-scid scid / scid) were purchased from Clea Japan, Inc. (Tokyo, Japan). All animal experiments were carried out with the approval of the Institutional Animal Care and Use Committee of the University of Tokyo. After allowing the SCID mice to fast for 24 hours, the animals were intraperitoneally injected with freshly prepared citrate-buffered saline (pH 4.5) containing streptozotocin (STZ, 150 mg / kg, Sigma-Aldrich, St. Louis Mo.). Blood glucose levels were measured using a glucometer and test strips (Glucose Pilot, Aventir Biotech LLC, Carlsbad, Calif.) on the third day after injection of STZ. Mice were considered to have diabetes mellitus (DM) if blood glucose level exceeded 300 mg / dl. Those mice that did not exhibit hyperglycemia (blood glucose level in excess of 300 mg / dl) were subjected to a second round of STZ injection (150 mg / kg) followed by monitoring bloo...

example 3

Cytokine Production Assay (ELISA)

[0075]MSC are known to secrete growth factors (such as PDGF, bFGF, TGF-β or EGF) required in the inflammatory phase and cell growth phase of wound healing (Maxson, S., et al., Stem Cells Transl. Med., Vol. 1, p. 142-149 (2012)). Therefore, the Muse cell population and MSC were cultured in vitro under hypoxic and normoxic conditions to examine cytokines secreted into the culture broth. The experiment was carried out in the manner indicated below. 4.0×105 cells of the Muse cell population and MSC were disseminated in a 60 mm culture dish followed by culturing in serum-free DMEM under hypoxic (1% O2) and normoxic (6% O2) conditions. The culture medium was recovered 48 hours later and filtered using a 0.22 μm filter (Millex-GV Filter, Millipore, Billerica, Mass.). The amounts of cytokines secreted into the culture broth were compared and examined using an ELISA kit for hepatocyte growth factor (HGF) and stromal cell-derived factor 1 (SDF-1) (both availab...

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Abstract

The purpose of the present invention is to provide a novel medicinal use in regeneration medicine, said medicinal use comprising using pluripotent stem cells (Muse cells). Provided is a cell preparation for treating skin ulcer, said cell preparation comprising SSEA-3 positive pluripotent stem cells isolated from a mesenchymal tissue of a living organism or cultured mesenchymal cells. The cell preparation according to the present invention is based on such mechanism of skin tissue regeneration that, when the Muse cells are administered to a skin ulcer site of a subject suffering from the aforesaid disease, the Muse cells differentiate into skin-constituting cells.

Description

TECHNICAL FIELD[0001]The present invention relates to a cell preparation for use in regenerative medicine. More particularly, the present invention relates to a cell preparation comprising pluripotent stem cells effective for repair and regeneration of skin tissue in skin ulcers including skin ulcers caused by diabetes, and to a method for treating skin ulcers that uses these pluripotent stem cells.BACKGROUND ART[0002]Diabetes is a disease that is associated with persistent hyperglycemic state and is said to occur as a result of the actions of a diverse range of environmental and genetic factors. The main regulatory factor of blood sugar is insulin, and hyperglycemia is known to occur due to insulin deficiency or an excess of factors that inhibit the action thereof (such as genetic predisposition, lack of exercise, obesity or stress). Diabetes is classified as type 1 diabetes, which occurs primarily due to a decrease in pancreatic insulin secretory function attributable to such fact...

Claims

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Application Information

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IPC IPC(8): A61K35/28
CPCA61K35/28A61K35/12A61K35/545A61P17/02A61P3/10
Inventor YOSHIMURA, KOTAROKINOSHITA, KAHORIDEZAWA, MARI
Owner THE UNIV OF TOKYO
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