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In vitro and in vivo studies of tissue engineering in reconstructive plastic surgery
Linköping University, Department of Clinical and Experimental Medicine, Burn Unit . Linköping University, Faculty of Health Sciences.
2005 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

To correct, improve, and maintain tissues, and their functions, are common denominators in tissue engineering and reconstructive plastic surgery. This can be achieved by using autolo-gous tissues as in flaps or transplants. However, often autologous tissue is not useable. This is one of the reasons for the increasing interest among plastic surgeons for tissue engineering, and it has led to fruitful cross-fertilizations between the fields. Tissue engineering is defined as an interdisciplinary field that applies the principles of engineering and life sciences for development of biologic substitutes designed to maintain, restore, or improve tissue functions. These methods have already dramatically improved the possibilities to treat a number of medical conditions, and can arbitrarily be divided into two main principles:

> Methods where autologous cells are cultured in vitro and transplanted by means of a cell suspension, a graft, or in a 3-D biodegradable matrix as carrier.

> Methods where the tissue of interest is stimulated and given the right prerequisites to regenerate the tissue in vivo/situ with the assistance of implantation of specially designed materials, or application of substances that regulate cell functions - guided tissue regeneration.

We have shown that human mammary epithelial cells and adipocytes could be isolated from tissue biopsies and that the cells kept their proliferative ability. When co-cultured in a 3-D matrix, patterns of ductal structures of epithelial cells embedded in clusters of adipocytes, mimicking the in vivo architecture of human breast tissue, were seen. This indicated that human autologous breast tissue can be regenerated in vitro.

The adipose tissue is also generally used to correct soft tissue defects e.g. by autologous fat transplantation. Alas 30-70% of the transplanted fat is commonly resorbed. Preadipocytes are believed to be hardier and also able to replicate, and hence, are probably more useful for fat transplantation. We showed that by using cell culture techniques, significantly more pre-adipocytes could survive and proliferate in vitro compared to two clinically used techniques of fat graft handling. Theoretically, a biopsy of fat could generate enough preadipocytes to seed a biodegradable matrix that is implanted to correct a defect. The cells in the matrix will replicate at a rate that parallels the vascular development, the matrix subsequently degrades and the cell-matrix complex is replaced by regenerated, vascularized adipose tissue.

We further evaluated different biodegradable scaffolds usable for tissue engineering of soft tissues. A macroporous gelatin sphere showed several appealing characteristics. A number of primary human ecto- and mesodermal cells were proven to thrive on the gelatin spheres when cultured in spinner flasks. As the spheres are biodegradable, it follows that the cells can be cultured and expanded on the same substrate that functions as a transplantation vehicle and scaffold for tissue engineering of soft tissues.

To evaluate the in vivo behavior of cells and gelatin spheres, an animal study was performed where human fibroblasts and preadipocytes were cultured on the spheres and injected intra-dermally. Cell-seeded spheres were compared with injections of empty spheres and cell suspensions. The pre-seeded spheres showed a near complete regeneration of the soft tissues with neoangiogenesis. Some tissue regeneration was seen also in the ‘naked’ spheres but no effect was shown by cell injections.

In a human pilot-study, intradermally injected spheres were compared with hyaluronan. Volume-stability was inferior to hyaluronan but a near complete regeneration of the dermis was proven, indicating that the volume-effect is permanent in contrast to hyaluronan which eventually will be resorbed. Further studies are needed to fully evaluate the effect of the macroporous gelatin spheres, with or without cellular pre-seeding, as a matrix for guided tissue regeneration. However, we believe that the prospect to use these spheres as an injectable, 3D, biodegradable matrix will greatly enhance our possibilities to regenerate tissues through guided tissue regeneration.

Place, publisher, year, edition, pages
2005.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 905
Keyword [en]
Tissue engineering, plastic surgery, guided tissue regeneration
National Category
Surgery
Identifiers
URN: urn:nbn:se:liu:diva-8504ISBN: 91-85299-06-5 (print)OAI: oai:DiVA.org:liu-8504DiVA: diva2:23287
Public defence
2005-09-02, Berzeliussalen, Hälsouniversitetet, Linköping, 09:00 (English)
Opponent
Supervisors
Note
On the day of the defence date the status of article V was In Press.Available from: 2007-03-16 Created: 2007-03-16 Last updated: 2009-08-23
List of papers
1. Mammary epithelial cell and adipocyte co-culture in a 3-D matrix: The first step towards tissue-engineered human breast tissue
Open this publication in new window or tab >>Mammary epithelial cell and adipocyte co-culture in a 3-D matrix: The first step towards tissue-engineered human breast tissue
2001 (English)In: Cells Tissues Organs, ISSN 1422-6405, Vol. 169, no 4, 361-367 p.Article in journal (Refereed) Published
Abstract [en]

Reconstruction of the female breast after cancer surgery is a demanding task where the methods used today suffer from several disadvantages. In the present study we have investigated the possibility to use tissue engineering methods to regenerate human autologous breast tissue. Human mammary epithelial cells and preadipocytes were derived from breast tissue biopsies from healthy women undergoing reduction mammoplasty, and the two celltypes were co-cultured with conventional cell culture methods as well as in 3-D matrices. The study shows that it is possible to harvest both human mammary epithelial cells and preadipocytes in a single session, propagate several subcultures, and that the cells maintain a normal intercellular distribution and growth-pattern when co-cultured in a 3-D collagen gel. We propose that growth and formation of a tissue closely resembling normal human breast tissue be readily obtained in the described in vitro cell culture set-up using basic tissue engineering principles. This concept may be of great importance in the development of new methods for reconstruction of the human breast.

Keyword
Tissue engineering, Adipocyte, Mammary epithelial cell, Breast tissue
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-14344 (URN)10.1159/000047903 (DOI)
Available from: 2007-03-16 Created: 2007-03-16 Last updated: 2009-08-18
2. Adipose tissue processed for lipoinjection shows increased cellular survival in vitro when tissue engineering principles are applied: Culture techniques and survival of fat
Open this publication in new window or tab >>Adipose tissue processed for lipoinjection shows increased cellular survival in vitro when tissue engineering principles are applied: Culture techniques and survival of fat
2002 (English)In: Scandinavian Journal of Plastic and Reconstructive Surgery and Hand Surgery, ISSN 0284-4311, Vol. 36, no 3, 166-171 p.Article in journal (Refereed) Published
Abstract [en]

Correcting soft tissue defects by autologous fat grafting is a routine procedure in plastic surgery. Its efficacy and safety has been discussed extensively and several techniques of lipoinjection have been developed. However, one is bound to overcorrect by 30%-70% or need to repeat the procedure because of resorption of the transplant. The reasons are that many of the transplanted cells are already differentiated, and also that there is no nutritional support to the inner cell layers when they are transplanted as fragments. By culturing autologous adipocytes one can ensure that only non-differentiated, but committed, preadipocytes are transplanted and the procedure can be done in a way that ensures optimal nutritional support for the cells. In the present study we have compared our cell culture technique with two common clinical ways of processing liposuction material and found that (pre)adipocytes survive and proliferate significantly better in cell culture.

Keyword
words, cell culture, fat grafting, tissue engineering
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-14345 (URN)
Available from: 2007-03-16 Created: 2007-03-16 Last updated: 2009-08-18
3. New degradable polymer scaffold for regeneration of the dermis In vitro and in vivo studies
Open this publication in new window or tab >>New degradable polymer scaffold for regeneration of the dermis In vitro and in vivo studies
Show others...
Manuscript (Other academic)
Identifiers
urn:nbn:se:liu:diva-14346 (URN)
Available from: 2007-03-16 Created: 2007-03-16 Last updated: 2010-01-13
4. Growth of cultured human ecto- and mesodermal cells on macroporous biodegradable gelatin spheres
Open this publication in new window or tab >>Growth of cultured human ecto- and mesodermal cells on macroporous biodegradable gelatin spheres
Show others...
Manuscript (Other academic)
Identifiers
urn:nbn:se:liu:diva-14347 (URN)
Available from: 2007-03-16 Created: 2007-03-16 Last updated: 2010-01-13
5. Macroporous gelatine spheres as culture substrate, transplantation vehicle, and biodegradable scaffold for guided regeneration of soft tissues.: In vivo study in nude mice
Open this publication in new window or tab >>Macroporous gelatine spheres as culture substrate, transplantation vehicle, and biodegradable scaffold for guided regeneration of soft tissues.: In vivo study in nude mice
2007 (English)In: Journal of Plastic, Reconstructive, and Aesthetic Surgery, ISSN 1748-6815, Vol. 60, no 5, 543-555 p.Article in journal (Refereed) Published
Abstract [en]

In the course of development of a new type of filler for the correction of small defects in soft tissues we studied macroporous gelatine spheres as culture substrate, transplantation vehicle, and biodegradable scaffold for guided regeneration of soft tissues in vivo. We injected intradermally in nude mice gelatine spheres that had either been preseeded with human fibroblasts or preadipocytes, or left unseeded. We compared the extent of regenerated tissue with that found after injections of saline or single-cell suspensions of human fibroblasts or preadipocytes. Routine histological examinations and immunohistochemical staining for von Willebrand factor (indicating neoangiogenesis) were made after 7, 21, and 56 days. Injected saline or single-cell suspensions had no effect. However, a quick and thorough tissue regeneration with developing neoangiogenesis was elicited by the gelatine spheres and the effect of spheres preseeded with preadipocytes surpassed the effect of spheres preseeded with fibroblasts, which in turn surpassed the effect of unseeded gelatine spheres. We suggest that minor soft tissue defects such as wrinkles or creases can be corrected by injection of naked macroporous gelatine spheres, whereas larger defects are best corrected by injection of macroporous gelatine spheres preseeded with fibroblasts, or preadipocytes, or both.

Keyword
Filler, Soft tissue defect, Guided tissue regeneration, Tissue engineering, Animal, In vivo
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-14348 (URN)10.1016/j.bjps.2005.10.031 (DOI)
Available from: 2007-03-16 Created: 2007-03-16 Last updated: 2009-08-18
6. Use of macroporous gelatine spheres as a biodegradable scaffold for guided tissue regeneration in humans: An in vivo study
Open this publication in new window or tab >>Use of macroporous gelatine spheres as a biodegradable scaffold for guided tissue regeneration in humans: An in vivo study
2005 (English)In: Journal of Plastic, Reconstructive, and Aesthetic Surgery, ISSN 1748-6815Article in journal (Refereed) Submitted
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-14349 (URN)
Available from: 2007-03-16 Created: 2007-03-16

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Huss, Fredrik R.M.

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