Connective tissue and wound healing

1. Background and significance

Collagen is the most abundant protein of the skin, since it forms about 70% of the dry weight of the skin. Most of the skin collagen is composed of type I collagen and type III and others are the minor components. The biosynthesis of collagen is complex and it is vulnerable to disturbances in several genetic diseases (ie Ehlers-Danlos syndrome) and acquired diseases (ie scleroderma).

Figure 1

In addition many external and internal factors affect on the amount and biosynthesis of collagen. For example ultraviolet radiation increases degradation of collagen, and topical and systemic glucocorticoids decrease the synthesis rate of type I and III collagens. This is the major factor leading to steroid induced skin atrophy. Also other hormones such as thyroid hormones have profound effects on skin.

The collagen synthesis and degradation are also altered by aging and smoking.

The purpose of the present project is to study in vivo in patients collagen and collagen synthesis and degradation especially in smokers and those getting hormone treatments. An other part of the project is focusing on studying the wound healing in vivo employing methods developed in our laboratory. The mechanism of wound healing is essential for life, since not only the healing of visible wounds but also the curing of small, mostly epithelial defects take place continuously in a human being.

Wound healing is affected by external and internal factors and in many diseases such as diabetes.

The basic phenomena of cell motility and inflammation remain to be investigated, since they are not known in detail in humans in vivo. In addition non-healing wounds pose a significant burden to many patients.

2. Recent progress

In the recent studies we have investigated the connective tissue and collagen synthesis and degradation in heavy smokers. The study population consisted of 98 Finnsih males, of whom 47 were current smokers and 51 non-smokers. The collagen synthesis was measured from the skin using suction blister method; in which the suction blister are induced to the skin by a special device with vacuum (see figure 1) and after suction blisters (see figure 2) have developed the blister fluid is collected and used to analyze type I and III collagen propeptides. The method is very sensitive to detect minor changes in collagen synthesis rate.

Figure 2

The results revealed decreased synthesis of type I and III collagens in smokers as well as regulatory mechanisms which control the turnover of these and other extracellular matrix proteins. The level of matrixmetalloproteinase-8 (MMP-8) was increased in suction blisters in smokers in compared to controls. Also the level of tissue inhibitor of matrix metalloproteinase (TIMP-1) was lower in smokers in compared to controls.

These alterations could have deleterious action on skin connective tissue in smokers and cause wrinkling and delayed wound healing.

The effect of radiotherapy on human skin type I and III collagen synthesis was investigated in a group of women who were treated for breast cancer surgically and with radiotherapy. The synthesis of type I and III collagens was markedly increased as a result of radiotherapy. An increased amount of cross-linked type I collagen was detected in irradiated skin and collagen turnover was also increased in irradiated skin.

The results of this study suggest that alteration of collagen metabolism contributes to dermal side effects of therapeutic irradiation.

In collaboration with prof. Jan Faergemann (Sahgrenska University Hospital, Sweden) we have been able to show that topical application of thyroid hormone analogue is able to reverse glucocorticoid-induced skin atrophy in humans.

The wound healing was studied in diabetic patients employing suction blister model. The recovery of skin was followed by measuring the transepidermal water loss (TEWL). The microvascular reactivity was followed by laser Doppler and expression of cytokeratins and tight junctions by immunohistochemistry. The diabetic patients showed slower restoration of epidermal barrier and a weaker initial inflammatory response.

3. Future goals

We shall continue our studies on effects of irradiation on skin connective tissue. Especially our focus is to study the gene expression of several important mediators (TGF-beta, TNF-alfa etc) by gene array techniques. Also the studies on thyroid hormone analogue on skin connective tissue and wound healing will be investigated in detail.

We have started in collaboration with dr Vesa Koivukangas and prof Tero Ala-kokko to study the wound healing in patients who have severe sepsis. In this project large number of skin samples have been collected and will be analyzed using antibodies to cytokines, integrins, MMPs and collagens. Also the re-epithelialization rate and inflammation will be analyzed.

4. Research group members

4.1. Project leader

Aarne Oikarinen, Professor, M.D., Ph.D., University of Oulu

• MD degree 1974, University of Oulu
• PhD degree 1977, University of Oulu
• Specialist in dermatology and venereology, 1982
• Post-doctoral fellow 1982-84 UCLA, Los Angeles, USA
• Assistant professor, 1984-1986, University of Oulu
• Junior research fellow of the Academy of Finland, 1987-89
• Senior research fellow of the Academy of Finland, 1990-91
• Associate professor 1991-93, 95 University of Oulu
• Professor and chair of Dermatology and Venereology, 1997
• The current number of publications (original in scientific journals) is over 150 and they have been cited over 3000 times in scientific publications

4.2. Senior and post-doctoral investigators

Vesa Koivukangas, M.D., Ph.D.

4.3. Ph.D. students

Marjo Koskela M.D.

Fiia Gäddnäs, medical student

5. Project funding

The salaries, and laboratory reagents and analysis are provided by University of Oulu and by EVO a special research money for the University hospital provided by state.

6. Publications relevant to the project

• Oikarinen A et al. A new method to measure type I and III collagen synthesis in human skin in vivo. Demonstration of decreased collagen synthesis after topical glucocorticoid treatment. Journal of Investigative Dermatology, 98,220-225, 1992.
• Oikarinen A et al Demonstration of 72-kda and 92-kda forms of type IV collagenase in human skin: variable expression in various blistering diseases, induction during re-epithelialization and decrease by topical glucocorticoids. Journal of Investigative Dermatology, 101,205-210, 1993.
• Kainulainen T et al Laminin-5 expression is independent of the injury and microenvironment during re-epithelialization of wounds. Journal of Histochemistry and Cytochemistry, 46,353-360, 1998.
• Raitio Anina Smoking and skin. Comparison of the appearance, physical qualities, morphology, collagen synthesis and extracellular matrix turnover of skin in smopkers and non-smokers, thesis, Acta Univ. Oulu D 841, 2005.
• Riekki Riitta Late dermal effects of breast cancer radiotherapy, thesis, Acta Univ. Oulu D901, 2006.
• Koivukangas Vesa Wound healing in a suction blister model, Acta Univ. Oulu D 810, 2004.
• Yazdanparast P et al A thyroid hormone analogue, triiodothyroacetic acid, corrects corticosteroid-downregulated collagen synthesis. Thyroid 5,345-353, 2004.