KR20100097574A - Cell composition for promoting hair growth - Google Patents

Abstract

PURPOSE: A composition for inducing hair growth containing adipose-derived cells and a therapeutic agent for hair loss are provided to transplant autologous cells to scalp and to enhance hair growth. CONSTITUTION: A composition for inducing hair growth contains autologous blood-derived monocytes and adipose-derived cells including adipose stem cells and mesenchymal stromal cells as active ingredients. The blood-derived monocytes are epithelial progenitor cells or hematopoetic stem cells. A therapeutic agent for hair loss contains the adipose-derived cells as active ingredients. A therapeutic agent for atrichia contains the adipose-derived cells as active ingredients.

Description

Cell composition for Promoting Hair Growth

The present invention relates to a composition for inducing hair growth, alopecia treatment agent and alopecia treatment agent containing adipose tissue-derived cells including autologous blood-derived monocytes and adipose stem cells and mesenchymal stromal cells as an active ingredient.

The cause of hair loss is a testosterone effect by DHT (Dihydrotestosterone), a type of male hormone. The male hormone binds to 5a-reductase in the cell and activates the enzyme.DHT, a reaction product produced in this process, inhibits protein synthesis in hair follicles during DNA transcription, which damages the hair follicles. Hair loss occurs. Other causes include genetic hair loss and hair loss caused by stress. In addition, hair loss caused by cranial development, which causes thinning of the scalp and blood flow disorders due to age, because the scalp stops developing faster than the skull. , Hair loss due to aging has been reported (Sheen, Fighting Hair Loss , USA Library Publishing, p. cm. 97-090157, 1966).

Various methods have been introduced as a treatment for hair loss, and the most commonly used method is self hair transplantation (self hair transplantation). This method does not have a problem such as rejection because the hair tissue is transplanted to the area of hair loss, but there is a problem to be solved, such as the limitation of the graft area and the effective control of engraftment and continuous growth after transplantation. There is a disadvantage that the satisfaction after the fall.

Recently, a treatment method using genes is expected as a treatment method. In January 1998, Dr. Angela Christiano of the University of Columbia, USA, discovered and announced a genetic factor involved in systemic alopecia disease, opening a new chapter in gene therapy for hair loss (Ahmad et. al . , Science , 30; 279: 720, 1998).

Gene therapy has been developed using methods for delivering DNA directly necessary to hair follicles using the above-described hair growth-related genes, and treatment methods such as blocking the expression of hair loss genes. However, gene therapy has a high degree of concern about the disadvantages that are not yet known about the efficacy, cost, stability, and future effects of the current treatment, and even though it reveals the genetic factors involved in hair loss, the treatment method using the gene may Due to technical problems, it is recognized that it will take a considerable time before it is realized safely.

Only two drugs are used: Propecia (Merck) and Minoxidil (Rogaine, Pfizer), which are recognized by the US Food Safety Agency (FDA). Two agents that have the effect of inhibiting type 2 5a-reductase (reductase) have been reported to have hair loss prevention and treatment effects of about 20-30% for minoxidil and 60-70% for propecia. Photographic evaluation showed that 66% of men treated with the drug showed marked hair reconstruction (Bouhanna, Dermatol . Surg . , 29: 1130, 2003; Thiboutot, Arch . Dermatol . , 135: 1417, 1999; Bouhanna, Dermatol . Surg ., 29: 1130, 2003; Messenger and Rundegren, Br . J. Dermatol ., 150: 186, 2004). However, such drug treatment is not fundamentally a treatment method by new hair follicle formation or hair regeneration, and its effect is effective at the time of administration, but when the medicine is stopped, hair loss resumes as it was before taking the medicine after a few months. In addition, many side effects such as sexual dysfunction or skin abnormalities have been reported (Messenger and Rundegren, Br J Dermatol. , 150: 186, 2004).

Alopecia, unlike normal hair growth in general, has lost its ability to produce hair, so regenerating the dermal papilla of hair follicles seems fundamentally impossible. However, active researches have been made to solve these problems, and attention is now being extended to the application of stem cells having hair regeneration ability in the treatment of hair loss. Thus, as well as studies on the differentiation ability of multipotent stem cells into hair follicle cells and the production of new hair follicles by hair follicle stem cells present in the scalp tissue, the fact that the correlation between epidermis and mesenchyme is an important factor for hair follicle formation. Was confirmed through animal experiments (Oshima et. al ., Cell , 104: 223, 2001; Lavker et al . , JID Symposium Proceedings 8:28, 2003; Kopen et al ., P roc . Nat . Acad . Sci USA , 96: 10711, 1999).

Bone marrow is a typical human body that can provide stem cells capable of differentiating into hair follicle cells, but since the bone marrow is not easily obtained, adipose tissue is a convenient site for obtaining stem cells. Zuk et al . , Tissue Eng ., 7: 211, 2001; Mizuno sod et al . , Plast . Reconstr . Surg . , 109: 199, 2002; Zuk et al . , Mol . Biol . Cel . , 13: 4279, 2002). Adipose tissue can be obtained in a large amount by non-invasive techniques such as liposuction, and the liposuction contains adipose stem cells (adipose precursor cells) and adipose tissue-derived stromal cells, which are the predecessor of adipocytes. For example, the cells can be easily obtained from crushed human adipose tissue by degradation by collagenase and differential centrifugation. It is also demonstrated that the expression of cell surface proteins is not the same as bone marrow stem cells, but can differentiate into adipocytes, chondrocytes, and osteoblasts under appropriate culture conditions (Erickson et al. al . , Biochem . Biophys . Res . Commun . , 290: 763, 2002; Halvorsen et al . , Tissue Eng . , 7: 729, 2001; Wickham et al . , Clin. Orthop . Relat . Res . , 412: 196, 2003; Dragoo et al . , J. Bone Joint Surg ., Br. , 85: 740, 2003). Stem cells present in adipose tissue are present in about 1000 times more than cells obtained from bone marrow, and stromal cells are present at less than 8.6 x 10 ⁴ cells per gram of adipose tissue, unlike stem cells. It can also be easily cultured in vitro can be supplied in large quantities, there is an advantage that can be stored at low temperatures.

In the reconstruction of hair follicle tissues, the interaction between the skin tissue and the mesenchyme of the tissue region to be reconstructed and the formation of neovascularization for smooth tissue regeneration are also recognized as necessary conditions for induction of hair growth.

Angiogenesis is closely regulated by several factors and homeostasis is maintained by a balance of factors that promote or inhibit vascular endothelial cell proliferation. Endothelial progenitor cells (EPCs), which are representative angiogenesis cells that can be conveniently obtained from bone marrow, umbilical cord blood and peripheral blood, respond to externally administered angiogenesis factors such as ischemia or VEGF. It is characterized by mobilizing circulating blood in and returning to ischemic tissue to participate in angiogenesis.

Therefore, the inventors of the present invention are not only adipose tissue-derived cells including autologous adipose tissue-derived adipose stem cells and mesenchymal stromal cells capable of differentiating into hair follicle cells, but also blood vessels to induce angiogenesis and improve hair growth effects. The present invention was completed by repeatedly transplanting a composition including autologous blood-derived monocytes containing endothelial progenitor cells and hematopoietic stem cells into an empty hair area, and remarkably improving the hair regrowth effect than conventional drug treatment. .

Disclosure of Invention An object of the present invention is to provide a composition for inducing hair growth, alopecia treatment and alopecia treatment, comprising autologous blood-derived monocytes and adipose stem cells and autologous adipose tissue-derived cells, including mesenchymal stromal cells. have.

In order to achieve the above object, the present invention is a foot containing autologous adipose-derived monocytes, including vascular endothelial progenitor cells and / or hematopoietic stem cells and autologous adipose tissue-derived cells, including adipose stem cells and mesenchymal stromal cells as an active ingredient It provides a composition for breast induction.

The present invention also provides an agent for treating hair loss or alopecia containing autologous adipose-derived cells, including autologous blood-derived monocytes and adipose stem cells and mesenchymal stromal cells as an active ingredient.

A composition for inducing hair growth, alopecia treatment and alopecia treatment, comprising autologous adipose tissue-derived cells, including blood-derived monocytes and adipose stem cells and mesenchymal stromal cells, according to the present invention, differentiate into hair follicle cells. It is possible to regenerate hair follicles without rejection after transplantation of the scalp tissue, and because it may have a synergistic effect on induction of hair growth by neovascularization by blood-derived monocytes. It is useful for hair loss treatment, alopecia treatment and hair shaping for cosmetic purposes.

In one aspect, the present invention relates to a composition for inducing hair growth comprising autologous adipose-derived cells, including autologous blood-derived monocytes and adipose stem cells and mesenchymal stromal cells as an active ingredient.

The composition for inducing hair growth of the present invention allows neovascularization to be formed by blood-derived monocytes, including vascular endothelial progenitor cells and / or hematopoietic stem cells, and thus, adipose stem cells and mesenchymal stromal cells contained in adipose tissue-derived cells. By inducing a synergistic effect on the differentiation of undifferentiated cells, such as stromal cells, into hair follicle cells, reconstruction of the hair loss or hair loss site is solved by solving two problems, namely, a blood circulation disorder that causes hair loss and the absence of hair follicle cells.

In the present invention, 'hair growth induction' refers to the ability to induce hair follicle reconstruction on the epidermis by forming a hair follicle in the hair loss site, the hair loss site or hair loss site.

'Monocyte' of the present invention is derived from autologous blood, can be obtained from bone marrow, umbilical cord blood, placental blood, spleen, and preferably can be isolated from peripheral blood. In addition, the monocytes may be used in the method of inducing hair growth in a mixed state of blood-derived lymphocytes or leukocytes such as monosite, a few epithelial progenitor cells, and hematopoetic stem cells. Only epithelial progenitor cells and hematopoetic stem cells can be isolated and used for transplantation.

In the present invention, 'stem cells' generally refers to primitive cells capable of differentiating into other specific tissues and the ability to self-renewal to continue to proliferate in an immature, undifferentiated state.

The monocytes of the present invention may be composed of 10 ³ to 10 ¹⁰ cells, and are generally used to treat all the cells obtained from less than 50 ml of autologous blood collected in the hair loss lesion site to prepare a hair growth inducing composition. Can be.

Adipose stem cells and mesenchymal stromal cells of the present invention are autologous adipose tissue-derived cells, which are decomposed by collagenase and crushed by differential centrifugation of adipose tissue obtained as a by-product of the lipolysis procedure. It can be easily obtained by known techniques such as methods (Halvorsen et al . , Tissue Eng . 7 (6): 729, 2001; Hauner et al . , J. Clin . Invest . 84: 1633, 1989; Rodbell et al . , J. Bio . Chem. , 241: 130, 1966). The adipose stem cells and mesenchymal stromal cells (mesenchymal stromal cells) is in a state of undifferentiation or differentiation, and when appropriate growth factors for cell differentiation, epithelial stem cells, matrix cells, clonogenic keratinocytes, skin cells, fat cells, etc. There is a characteristic of multipotentiality that can be differentiated into.

In the present invention, it is possible to induce hair growth into adipose tissue-derived cells including adipose stem cells and mesenchymal stromal cells. This is because fat stem cells or mesenchymal stromal cells with attached to this site can restore the structure of the lower hair follicles. Therefore, the present invention is characterized in that hair growth is induced by multipotent adipose stem cells and mesenchymal stromal cells, which are derived from autologous adipose tissue, together with monocytes.

In the present invention, in the reconstruction of the alopecia by the cells having differentiation ability derived from adipose tissue, it shows a synergistic effect in the induction of hair growth by angiogenesis, the cell therapy method for such angiogenesis is endothelial progenitors directly collected from bone marrow It is known that positive angiogenic effects can be obtained by administering cells to ischemic sites in cardiovascular or neurological diseases, or by implanting endothelial progenitor cells from peripheral blood vessels into animals with symptoms of ischemic lower limbs and myocardial infarction. Anklesaria P. et al ., Proc . Natl . Acad. Sci . USA , 84: 7681, 1987; Flax JD et al ., Nat . Biotechnol . 16: 1033, 1998; Kalka C. et al ., Proc . Natl . Acad . Sci . USA, 97: 3422, 2000; Lindvall O. et al ., Science , 247: 574, 1990; Murohara T. et al., J. Clin. Invest., 105: 1527, 2000; Kawamoto A. et al ., Circulation , 103: 634, 2005).

The composition for inducing hair growth of the present invention, in order to induce more excellent hair growth efficacy, in the step of transplanting the blood-derived monocytes TGF (transforming growth factor), IFG (insulin-like growth factor), platelet-derived growth factor (PDFG) EGF (epidermal growth factor), AFBF (acidic fibroblast growth factor), bFBF (basic fibrblast growth factor), HGF (hepatocytic growth factor), KFG (keratocyte growth factor), vascular endothelial growth factor (VEGF), Neuropeptides, proteiglycans, Growth factors capable of stimulating the smooth formation of hair follicle cells such as Noggin, Wnt Hedghog, TNF Family, and catenin may be additionally administered once or repeatedly at the same time or at different time intervals.

The composition for inducing hair growth of the present invention can be used in a single dose to contain 10 ³ to 10 ¹⁰ self-derived monocytes and 10 ⁵ to 10 ¹² adipose tissue-derived cells, preferably 10 ⁴ self-derived monocytes and 10 ⁷ adipose tissue derived cells can be used. The amount may be determined by a person skilled in the art, and may be variously prescribed by factors such as the hair loss state and the response sensitivity of the recipient.

The technique for administering the composition for inducing hair growth to the tissue can be performed by a person skilled in the art, and includes intravenous administration, intraperitoneal administration, intramuscular administration, subcutaneous administration, or topical administration. Parenteral administration is possible, preferably by subcutaneous or topical administration, and most preferably by direct injection into the area in need of hair loss or hair growth.

A key factor to be considered in the transplantation of adipose tissue-derived cells, including monocytes and adipose stem cells and mesenchymal stromal cells, is that the histocompatibility between the recipient of the cell and the cell to be transplanted must be consistent. The immune rejection reaction can be avoided, and the monocytes and adipose tissue-derived cells of the present invention can solve these problems in that they are autologous cells.

The composition for inducing hair growth of the present invention can be transplanted with adipose tissue-derived cells, including blood-derived monocytes and adipose stem cells and mesenchymal stromal cells, to the scalp or tissues in need of hair growth. Cells may be separated and administered sequentially to tissues to exhibit hair growth induction efficacy.

The hair growth guide composition of the present invention may be administered to a tissue once or several times, and the hair growth guide composition of the present invention may be administered several times, and preferably may be repeated less than four times. The number and duration can be adjusted by the expert knowledge of those skilled in the art according to the state of.

The composition for inducing hair growth of the present invention is carried out for a predetermined period of time so that the administered cells are settled smoothly in the tissue with each administration, and the period of leaving may be 2 days to 15 days, preferably in the art. It may be determined by a specialist belonging to it, and may be shortened or prolonged by factors such as the hair loss state and response sensitivity of the recipient.

In another aspect, the present invention relates to a hair loss treatment agent and alopecia treatment agent containing adipose tissue-derived cells including autologous blood-derived monocytes and adipose stem cells and mesenchymal stromal cells as an active ingredient.

Autologous adipose-derived cells, including autologous blood-derived monocytes and adipose stem cells and mesenchymal stromal cells for hair reconstruction, or hair loss-inducing compositions containing the cells as an active ingredient, hair loss treatment agents, and hair loss treatment agents Can be used for implantation further comprising a biocompatible carrier. The carrier is composed of a substance that has no side effects due to toxicity, rejection or irritation to mammals, including humans, and serves as a collector or support to prevent the cells from dispersing away from the implanted site. It is apparent in the art that those skilled in the art performing transplant procedures can readily determine.

The biocompatible carrier may be alginate, agarose, fibrin, collagen, gelatin, fibronectin, polyglycolic acid, polylactic acid, hyaluronic acid, polyethylene glycol, chondroital ), Dermatan, polysaccharides, mucopolysaccharides, hydrogels, dextran, amylose, proteins, glycoproteins and derivatives thereof, or platelet-containing serum, concentrated platelet mixed compositions, and the like. have.

The composition for inducing hair loss of the present invention, the hair loss treatment agent, and the hair loss treatment agent may further include a lubricant, a humectant, an emulsifier, a suspension agent, a preservative, and the like, in addition to the above components. Suitable pharmaceutically acceptable carriers and agents are Remington's Pharmaceutical Sciences (19th ed., 1995).

The composition for inducing hair loss of the present invention, the agent for treating hair loss, and the agent for treating alopecia may be prepared using a pharmaceutically acceptable carrier and / or excipient according to a method which can be easily carried out by a person of ordinary skill in the art. Can be prepared in unit dose form or incorporated into a multi-dose container, and may further comprise a dispersant or stabilizer.

The composition for inducing hair loss of the present invention, the alopecia treatment agent and the alopecia treatment agent can be applied to the scalp for the treatment of typical male alopecia and so-called alopecia which may occur after menopause or ovarian debridement, which are recognized as 'baldness', It can be applied to any part of the body that needs hair growth. For example, it can be used to improve the condition of hair damaged by trauma scar, wide forehead or M forehead, lashes or eyebrows and alopecia for the purpose of simple cosmetic effect.

The composition for inducing hair growth of the present invention, the alopecia treatment agent and the alopecia treatment agent may be used as monotherapy, but may also be used in combination with other conventional hair growth induction drug therapy or surgery therapy, and when the combination therapy is performed to maximize the efficacy Can be represented.

Pharmaceutical preparations that can be used together in the method of inducing hair growth and hair regrowth of the present invention include male hormones or DHT-inducing inhibitors including parasteride, dutasteride, thioctol and RU58841, parathyroid hormone (PTH) inhibitors, Minoxidil, Burserelin (Burserelin), blood circulation improvers and compositions applied to mesotherapy, and the like, the surgical treatment includes hair transplantation, scalp flap, scalp reduction and the like.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are only for describing the present invention in more detail and that the scope of the present invention is not limited by these embodiments in accordance with the gist of the present invention .

Example  1: Isolation of Adipose-derived Cells

According to the patient's consent, 1 ~ 5 times of sterile saline solution of tissue volume is added to the adipose tissue (clinic M hospital) obtained from subcutaneous fat removal for abdominal, thigh and other parts, and washed twice with 0.3 ~ 0.5 Physiological saline mixed with% collagenase type I was added 1: 1 to the tissue volume and treated at 36 to 38 ° C. for 30 to 60 minutes. The enzyme reaction was stopped by adding autologous serum 0.1-0.3 times the adipose tissue volume. Centrifuge the enzymatically digested adipose tissue at 1,000 to 5,000 rpm for 1 to 5 minutes, and then add 10 to 40 ml of sterile saline solution to free the precipitated cells and pass the 70-100 μm filter to remove solid components. Removed. By adding the same amount of sterile physiological saline and washing the cells by centrifugation, adipose tissue-derived cells including adipose stem cells and mesenchymal stromal cells were obtained.

Example  2: Isolating autologous peripheral blood-derived monocytes

25 ml of peripheral blood collected from the brachial vein, after consent of liposuction, was placed in a 50 ml centrifuge tube containing heparin, followed by the same amount of Ficoll-Pauqe PLUS (density: 20 to 1.076 to 1.078 g / ml, Amersham-Pharmacia Biotech, Sweden) is carefully dispensed to layer at the bottom of the blood and centrifuged at 3,000 rpm, 4 ° C. for 20-30 minutes to form a cell layer separated by density difference, from which a Pasteur pipette or syringe The intermediate cell layer containing monocyte cells was collected using to separate hematopoietic stem cells, vascular endothelial progenitor cells, and monocytes including platelets from peripheral blood.

Example  3: caused by monocyte and adipose tissue-derived cells Hair cell Proliferative capacity  Judo

The hair dermal papilla cells used in this experiment were pulverized under a microscope with anatomical knife as a sample on the back skin tissue of 1 day old rats, and then reacted by shaking at 37 ° C incubator for 4 hours by adding 200 unit / ml collagenase I solution. I was. The cells were blown for 1 minute to separate the cells from the finished tissue solution and allowed to stand at room temperature for 30 minutes. The supernatant was then carefully placed on 5 ml of Histopaque 1083 solution (Sigma-aldrich, Inc, USA), followed by centrifugation for 20 minutes at 2500 rpm. After centrifugation, the intermediate cell layer was taken to obtain Dulbecco's modified Eagle's medium (DMEN) containing 10% fetal bovine serum (FBS), 100 unit / ml penicillin, 100 ug / ml streptomycin, 0.25 ug / ml amphotericin B (Gibco BRL.USA). After the culture medium was added to the cell culture plate (Sarstedt, 100x20mm) and cultured by addition of a mixture of saline, minoxidil or monocytes and adipose tissue-derived cells under conditions of 5% CO ₂ , 37 ℃.

The cell culture state after 7 days of cultivation of hair dermal papilla cells was observed at 40 times magnification under the Nikon eclipse TS100 microscope, and the cell growth rate of the hair dermal papilla cells with the cell mixture was added to the other additives. In comparison, it was found to be excellent (FIG. 1).

Example  4: hair growth by monocytes and adipose tissue-derived cells Induction  Confirm

1 x 10 ⁷ adipocyte-derived cells comprising adipocytes and mesenchymal stromal cells obtained from Example 1 and 1 X 10 ⁴ autologous blood-derived monocytes obtained from Example 2 were mixed with saline Thus, it was evenly administered at intervals of 1 cm to the scalp having four hair follicles at weekly intervals. In order to verify the hair growth efficacy, the results of applying the minoxyl (50mg, Hyundai drug) in the same period, the same number of times.

As a result, it was shown that the hair condition after one month was significantly different in the thickness of the hair and the number of hairs derived from one hair follicle than before the cell treatment (FIG. 2). In addition, the administration of the adipocyte-derived cells, including adipocytes and mesenchymal stromal cells, treated by Leader's dermatology alone (Fig. 3), was administered with the blood-derived monocytes of the present invention. When the hair growth effect appeared to be better (Fig. 4 and Fig. 5).

1 is a photograph showing the results of confirming the cell proliferation induction of the hair growth induction composition of the present invention for hair cells (A: saline treatment; B: minoxyl 50mg / ml; C: autologous blood-derived monocytes and adipose tissue derived Composition containing cells)

Figure 2 is a photograph of the hair root area showing the hair growth induction effect by the composition for inducing hair growth of the present invention on the scalp of the subject (A: Before Treatment; B: composition containing autologous blood-derived monocytes and adipose tissue-derived cells).

Figure 3 is a head photograph showing the hair growth induction effect on the scalp of the subject using only adipose stem cells treated by Readers Dermatology

Figure 4 is a top photo of the head showing the hair growth induction effect on the scalp of the subject 1 using the composition for inducing hair containing the blood-derived monocytes and adipose tissue-derived cells of the present invention.

Figure 5 is a top photo of the head showing the hair growth induction effect on the scalp of the subject 2 using the composition for inducing hair containing blood-derived monocytes and adipose tissue-derived cells of the present invention.

Claims (11)

A composition for inducing hair growth comprising adipose tissue-derived cells including autologous blood-derived monocytes and adipose stem cells and mesenchymal stromal cells as an active ingredient. The composition for inducing hair growth according to claim 1, wherein the blood-derived monocytes are one or more of epithelial progenitor cells and hematopoetic stem cells. The composition for inducing hair growth according to claim 1, wherein the adipose tissue-derived cells including adipose stem cells and mesenchymal stromal cells are derived from autologous adipose tissue. The composition of claim 1, wherein the composition comprises transforming growth factor (TGF), insulin-like growth factor (IFG), platelet-derived growth factor (PDFG), epidermal growth factor (EGF), acidic fibroblast growth facotr (AFBF), and bFBF. one or more growths selected from the group consisting of basic fibrblast growth factor (HGF), hepatocytic growth factor (HGF), keratocyte growth factor (KFG), vascular endothelial growth factor (VEGF), Neuropeptides, proteiglycans, Noggin, Wnt Hedghog, TNF Family and catenin A composition for inducing hair growth, further comprising a factor. A hair loss treatment agent containing autologous blood-derived monocytes and adipose tissue-derived cells including adipose stem cells and mesenchymal stromal cells as an active ingredient. The agent for treating hair loss according to claim 5, wherein the blood-derived monocytes are one or more of epithelial progenitor cells and hematopoetic stem cells. The hair loss treatment agent according to claim 5, wherein the adipose tissue-derived cells including the adipose stem cells and mesenchymal stromal cells are derived from autologous adipose tissue. An alopecia therapeutic agent containing autologous blood-derived monocytes and adipose tissue-derived cells including adipose stem cells and mesenchymal stromal cells as an active ingredient. The agent for treating alopecia according to claim 8, wherein the blood-derived monocytes are one or more of epithelial progenitor cells and hematopoetic stem cells. The method of claim 8, wherein the adipose tissue-derived cells including the adipose stem cells and mesenchymal stromal cells are derived from autologous adipose tissue. The method of claim 5 or 8, wherein the therapeutic agent is transforming growth factor (TGF), insulin-like growth factor (IFG), platelet-derived growth factor (PDFG), epidermal growth factor (EGF), and acidic fibroblast growth (AFBF). facotr), basic fibrblast growth factor (bFBF), hepatocytic growth factor (HGF), keratocyte growth factor (KFG), vascular endothelial growth factor (VEGF), Neuropeptides, proteiglycans, Noggin, Wnt Hedghog, TNF Family and catenin A therapeutic agent further comprising at least one growth factor selected.

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