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각막창상 치유에 대한 조직화학적 연구

Title
 각막창상 치유에 대한 조직화학적 연구
Other Titles
 Histochemical studies on corneal wound healing
Issue Date
1965
Publisher
 연세대학교 대학원
Description
의학과/박사
Abstract
[한글] Histochemical Studies on Corneal wound Healing Suck Joo Cho, M.D. Departments of Anatomy and Ophthalmology Yonsei University College of Medicine, Seoul, Korea (Director: Prof. K.D.Choi, M.D. and Prof. S.K.Hong, M.D.) In studies of corneal wound healing, Gutterbock (1871) and others proposed the theory of "mechanical sliding" of the corneal epithelium, and Wyss (1879) and others proposed the theory of epithelial cell multiplication. With regard to the appearance of inflammatory cells in the stroma, Cohnheim (1867), Robb and Kuwabara (1962) pointed out that leucocytes reached the corneal wound through the "conjunctival secretions". However, Maumenee and Kornblueth (1949), Heydenreich (1957) and Weimar (1958) observed the migration of leucocytes from perilimbal vessels into the corneal wound through stromal tissue. With regard to the demonstration of stromal metachromasia, Aurell(1954) pointed out that the greatest stromal metachromasia appeared of the end of the second week and then gradually decreased, with regard to content of stromal mucopolysaccharide, Francois and Rabaey (1956) stated that the quantity of mucopolysaccharide was not changed in spite of heavier local staining reaction. During corneal wound healing, Wolter (1958) differentiated the newly formed fibroblasts from the original activated stromal cells. Weimar (1958) emphasized the important role of fibroblasts in contributing to stromal regeneration, and Dunnington (1958) noted that the newly developed or covered corneal epithelium of the wound exerted a profound effect upon fibroblast formation in the healing corneal wound. In its absence fibroblast formation was decreased and did not reach a normal rate until the epithelium had grown across the anterior surface of the incision. Recently many histochemicaol studies of the cornea in experimental animals and human being have been presented. Wislocki (1952) presented in detail the histochemical characteristics of the eye of the normal rhesus monkey. Ichikawa (1954) observed the fluctuation of nucleic acids and mucopolysaccharide of the cornea during corneal wound healing. Since deConcillis (1934) first reported the enzymatic activity of alkaline phosphatase in the intact retina of rabbit, Bonting et al. (1961) demonstrated adenosine triphosphat ase in the intact ocular tissue of several animals and Lessell and Kuwabara (1964) have established in detail the action and presence of various phosphatases including adenosi ne di and triphosphatases in intact ocular tissues of various animals. The author was interested the fact that in wound healing of the avascular tissue of the cornea there is a different patteern from that of the ordinary tissue. In this experiments the author attempted to study histological and histochemical reactions of the cornea in the healing processes of artificially produced corneal wound. Materials and Methods The animals used in this experiment were 70 mature house rabbits weighing approximately 2.0 to 2.5kg. In the experimental groups, by 6 mm trephine the rabbits were given an artificial incision at the central zone of the cornea extending from a superficial to a perforating wound. The animals were killed at various intervals from 6 hours to 30 days. Pieces of intact and wounded cornea were fixed in Zenker's and Orth's fluid before the hematoxylin and eosin preparation, Weigert's resorcin fuchsin, toluidin blue, and methylen blue stains. Then 5 micra paraffin sections were made. When the periodic acid Shiff method was used, pieces of tissue were fixed in cold Rossman's fluid and then prepared in the same way of previously. In order to demonstrate desoxyribonucleic and ribonucleic acid, pieces of tissue were fixed in Lillie's fluid and stained in methyl green pyronin method after paraffin embedding. Nonspecific esterases were demonsterated using the coupling azo dye method for acid phosphatase and Gomori technique for alkaline phosphatase. For the demonstration of specific esterase, the Wachstein and Meisel techinique of adenosine di and triphosphatases was used. As a control part of the slides were stained with the periodic acid Schiff method after preincubation in saliva and diluted diastase solution. Also as a control part of the slides were stained with methyl green pyronin method after digestion in a diastase solution. The slides were compared with which were not digested preparations. Each incubating media were used as its own control. The main substrate for each enzyme activity was omitted to determine whether or not any artificial product was present and to exclude any artificial factor. Results and Conclusions 1. In the nonperforating wound spindle-shaped fibroblasts appeared early in the 6 hour group, their activity was recognized by the cellular multiplication or hyperplasia and hypertrophy of the corneal stroma which occurred near the wound edge. 2. In perforating wounds endothelium covered the wound that is the surface facing the anterior chamber completely in the 48 hour group and became cuboidal in shape. Healing was accompanied by a marked deposition of pyroninophilic substance in the cytoplasm. This phenomenon suggested an active contribution of endothelium to the healing process of the corneal wound. 3. As to meatachromasia, in the early stages, there was no significant incidence. It was evident in the 7 day group and at near the newly formed or loaded tissue of the wound cavity. Stainability was gradually reduced until it reached the normal state in the 3? day group. A significant metachromasia was observed at the end the first week, this indicates a quantitative increase of the local mucopolysaccharide associated with scar formation of the wound healing. 4. In the demonstration of glycogen and other polysaccharides by the periodic acid Schiff method, the early stage revealed a slight reducion of glycogen in the corneal epithelium near or at the wound. In the 24 or 48 hour group a moderate deposition of glycogen granules was observed in the epithelial cytoplasm of the whole layers covering the wound. The heavier deposition of glycogen granules in the epithelial cytoplasm covering the wound continued almost to the end of the third week, and the gradually decreased, the normal picture was present at the end of one month. A persistently heavier occurrance of glycogen in the covered epithelium of the wound would indicate a prolonged participation of the carbohydrate metabolism in the newly covered or developed epithelium raning from the early stages to the 3rd week after injury. 5. The nonspecific esterases of acid phosphatase and alkaline phosphatase were demonstrated. The former was confined to the epithelial layer except of r an indefinite linear short positive reaciton which occurred in the stromal immediately beneath the epithelial layer. The early stages revealed a progressive decrease of enzyme activity in the epithelial layer near the wound. However, the fibroblasts appearing in the wound did reveal some slight enzyme activity in their cytoplasm. Alkaline phosphatase activity did show slight impariment in the early stages. After 24 hours some parts of the cornearl epithelial layer near the wound or at the wound edge revealed an almost normal enzyme activity showing a tendency to rapid recovery. The fibroslasts which appeared in the wound showed early positive alkaline phosphatase enzyme activity. Demonstration of these enzyme activities in the stromal fibroblasts of the wound may indicate an active protein synthesis in the healing process of the stroma. 6. With regard to enzyme activities of specific esterases, as adenosine di and triphosphatase in the early stages these were absent and later the appeared in the wounded corneal epithelium. In the wound stroma, the fibroblasts near the wound showed a remarkable increase of these specific esterases and a moderate increase in the enzyme activities of the leucocytes in the wound. The appearance of positive enzyme activities probably indicates an active metabolic process which is related to the phosphate donor and energy supply.
[영문] In studies of corneal wound healing, Gutterbock (1871) and others proposed the theory of "mechanical sliding" of the corneal epithelium, and Wyss (1879) and others proposed the theory of epithelial cell multiplication. With regard to the appearance of inflammatory cells in the stroma, Cohnheim (1867), Robb and Kuwabara (1962) pointed out that leucocytes reached the corneal wound through the "conjunctival secretions". However, Maumenee and Kornblueth (1949), Heydenreich (1957) and Weimar (1958) observed the migration of leucocytes from perilimbal vessels into the corneal wound through stromal tissue. With regard to the demonstration of stromal metachromasia, Aurell(1954) pointed out that the greatest stromal metachromasia appeared of the end of the second week and then gradually decreased, with regard to content of stromal mucopolysaccharide, Francois and Rabaey (1956) stated that the quantity of mucopolysaccharide was not changed in spite of heavier local staining reaction. During corneal wound healing, Wolter (1958) differentiated the newly formed fibroblasts from the original activated stromal cells. Weimar (1958) emphasized the important role of fibroblasts in contributing to stromal regeneration, and Dunnington (1958) noted that the newly developed or covered corneal epithelium of the wound exerted a profound effect upon fibroblast formation in the healing corneal wound. In its absence fibroblast formation was decreased and did not reach a normal rate until the epithelium had grown across the anterior surface of the incision. Recently many histochemicaol studies of the cornea in experimental animals and human being have been presented. Wislocki (1952) presented in detail the histochemical characteristics of the eye of the normal rhesus monkey. Ichikawa (1954) observed the fluctuation of nucleic acids and mucopolysaccharide of the cornea during corneal wound healing. Since deConcillis (1934) first reported the enzymatic activity of alkaline phosphatase in the intact retina of rabbit, Bonting et al. (1961) demonstrated adenosine triphosphat ase in the intact ocular tissue of several animals and Lessell and Kuwabara (1964) have established in detail the action and presence of various phosphatases including adenosi ne di and triphosphatases in intact ocular tissues of various animals. The author was interested the fact that in wound healing of the avascular tissue of the cornea there is a different patteern from that of the ordinary tissue. In this experiments the author attempted to study histological and histochemical reactions of the cornea in the healing processes of artificially produced corneal wound. Materials and Methods The animals used in this experiment were 70 mature house rabbits weighing approximately 2.0 to 2.5kg. In the experimental groups, by 6 mm trephine the rabbits were given an artificial incision at the central zone of the cornea extending from a superficial to a perforating wound. The animals were killed at various intervals from 6 hours to 30 days. Pieces of intact and wounded cornea were fixed in Zenker's and Orth's fluid before the hematoxylin and eosin preparation, Weigert's resorcin fuchsin, toluidin blue, and methylen blue stains. Then 5 micra paraffin sections were made. When the periodic acid Shiff method was used, pieces of tissue were fixed in cold Rossman's fluid and then prepared in the same way of previously. In order to demonstrate desoxyribonucleic and ribonucleic acid, pieces of tissue were fixed in Lillie's fluid and stained in methyl green pyronin method after paraffin embedding. Nonspecific esterases were demonsterated using the coupling azo dye method for acid phosphatase and Gomori technique for alkaline phosphatase. For the demonstration of specific esterase, the Wachstein and Meisel techinique of adenosine di and triphosphatases was used. As a control part of the slides were stained with the periodic acid Schiff method after preincubation in saliva and diluted diastase solution. Also as a control part of the slides were stained with methyl green pyronin method after digestion in a diastase solution. The slides were compared with which were not digested preparations. Each incubating media were used as its own control. The main substrate for each enzyme activity was omitted to determine whether or not any artificial product was present and to exclude any artificial factor. Results and Conclusions 1. In the nonperforating wound spindle-shaped fibroblasts appeared early in the 6 hour group, their activity was recognized by the cellular multiplication or hyperplasia and hypertrophy of the corneal stroma which occurred near the wound edge. 2. In perforating wounds endothelium covered the wound that is the surface facing the anterior chamber completely in the 48 hour group and became cuboidal in shape. Healing was accompanied by a marked deposition of pyroninophilic substance in the cytoplasm. This phenomenon suggested an active contribution of endothelium to the healing process of the corneal wound. 3. As to meatachromasia, in the early stages, there was no significant incidence. It was evident in the 7 day group and at near the newly formed or loaded tissue of the wound cavity. Stainability was gradually reduced until it reached the normal state in the 3? day group. A significant metachromasia was observed at the end the first week, this indicates a quantitative increase of the local mucopolysaccharide associated with scar formation of the wound healing. 4. In the demonstration of glycogen and other polysaccharides by the periodic acid Schiff method, the early stage revealed a slight reducion of glycogen in the corneal epithelium near or at the wound. In the 24 or 48 hour group a moderate deposition of glycogen granules was observed in the epithelial cytoplasm of the whole layers covering the wound. The heavier deposition of glycogen granules in the epithelial cytoplasm covering the wound continued almost to the end of the third week, and the gradually decreased, the normal picture was present at the end of one month. A persistently heavier occurrance of glycogen in the covered epithelium of the wound would indicate a prolonged participation of the carbohydrate metabolism in the newly covered or developed epithelium raning from the early stages to the 3rd week after injury. 5. The nonspecific esterases of acid phosphatase and alkaline phosphatase were demonstrated. The former was confined to the epithelial layer except of r an indefinite linear short positive reaciton which occurred in the stromal immediately beneath the epithelial layer. The early stages revealed a progressive decrease of enzyme activity in the epithelial layer near the wound. However, the fibroblasts appearing in the wound did reveal some slight enzyme activity in their cytoplasm. Alkaline phosphatase activity did show slight impariment in the early stages. After 24 hours some parts of the cornearl epithelial layer near the wound or at the wound edge revealed an almost normal enzyme activity showing a tendency to rapid recovery. The fibroslasts which appeared in the wound showed early positive alkaline phosphatase enzyme activity. Demonstration of these enzyme activities in the stromal fibroblasts of the wound may indicate an active protein synthesis in the healing process of the stroma. 6. With regard to enzyme activities of specific esterases, as adenosine di and triphosphatase in the early stages these were absent and later the appeared in the wounded corneal epithelium. In the wound stroma, the fibroblasts near the wound showed a remarkable increase of these specific esterases and a moderate increase in the enzyme activities of the leucocytes in the wound. The appearance of positive enzyme activities probably indicates an active metabolic process which is related to the phosphate donor and energy supply.
URI
http://ir.ymlib.yonsei.ac.kr/handle/22282913/117037
Appears in Collections:
2. 학위논문 > 1. College of Medicine (의과대학) > 박사
Yonsei Authors
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