Neutral Salt-soluble Collagen Research Articles

Maturational alterations in gap junction expression and associated collagen synthesis in response to tendon function

Energy-storing tendons including the equine superficial digital flexor tendon (SDFT) contribute to energetic efficiency of locomotion at high-speed gaits, but consequently operate close to their physiological strain limits. Significant evidence of exercise-induced microdamage has been found in the SDFT which appears not to exhibit functional adaptation; the degenerative changes have not been repaired by the tendon fibroblasts (tenocytes), and are proposed to accumulate and predispose the tendon to rupture during normal athletic activity. The anatomically opposing common digital extensor tendon (CDET) functions only to position the digit, experiencing significantly lower levels of strain and is rarely damaged by exercise. A number of studies have indicated that tenocytes in the adult SDFT are less active in collagen synthesis and turnover than those in the immature SDFT or the CDET. Gap junction intercellular communication (GJIC) is known to be necessary for strain-induced collagen synthesis by tenocytes. We postulate therefore that expression of GJ proteins connexin 43 and 32 (Cx43; Cx32), GJIC and associated collagen expression levels are high in the SDFT and CDET of immature horses, when the SDFT in particular grows significantly in cross-sectional area, but reduce significantly during maturation in the energy-storing tendon only. The hypothesis was tested using tissue from the SDFT and CDET of foetuses, foals, and young adult Thoroughbred horses. Cellularity and the total area of both Cx43 and Cx32 plaques/mm 2 of tissue reduced significantly with maturation in each tendon. However, the total Cx43 plaque area per tenocyte significantly increased in the adult CDET. Evidence of recent collagen synthesis in the form of levels of neutral salt-soluble collagen, and collagen type I mRNA was significantly less in the adult compared with the immature SDFT; procollagen type I amino-propeptide (PINP) and procollagen type III amino-propeptide (PIIINP) levels per mm 2 of tissue and PINP expression per tenocyte also decreased with maturation in the SDFT. In the CDET PINP and PIIINP expression per tenocyte increased in the adult, and exceeded those in the adult SDFT. The level of PINP per mm 2 was greater in the adult CDET than in the SDFT despite the higher cellularity of the latter tendon. In the adult SDFT, levels of PIIINP were greater than those of PINP, suggesting relatively greater synthesis of a weaker form of collagen previously associated with microdamage. Tenocytes in monolayers showed differences in Cx43 and Cx32 expression compared with those in tissue, however there were age- and tendon-specific phenotypic differences, with a longer time for 50% recovery of fluorescence after photobleaching in adult SDFT cells compared with those from the CDET and immature SDFT. As cellularity reduces following growth in the SDFT, a failure of the remaining tenocytes to show a compensatory increase in GJ expression and collagen synthesis may explain why cell populations are not able to respond to exercise and to repair microdamage in some adult athletes. Enhancing GJIC in mature energy-storing tendons could provide a strategy to increase the cellular synthetic and reparative capacity.

Effect of succinic acid monoethyl ester on hemoglobin glycation and tail tendon collagen properties in type 2 diabetic rats

Succinic acid monoethyl ester (EMS) was recently proposed as an insulinotropic agent for the treatment of type 2 diabetes. The aim of the study was to investigate the effect of EMS and metformin administration on tail collagen content and its characteristics in streptozotocin-nicotinamide-induced type 2 diabetic rats. EMS was administered intraperitoneally for 30 days to normal and diabetic rats. In the diabetic rats, a significant increase in the levels of glucose, glycated hemoglobin, hydroxyproline, collagen content, extent glycation, fluorescence, neutral salt, acid and pepsin soluble collagen content was absorbed with a significant decrease in the level of insulin, hemoglobin in streptozotocin-nicotinamide diabetic rats. Moreover, a daily administration of nonglucidic nutrient EMS and metformin significantly decreased the levels of glucose, glycated hemoglobin, hydroxyproline, collagen content, extent glycation, fluorescence, neutral salt, acid and pepsin soluble collagen content, whereas it increased insulin, hemoglobin levels in diabetic rats. The positive influence of nonglucidic nutrient on both collagen content and its properties suggests a potential mechanism for the ability of EMS to delay diabetic complications.

AGE-related cross-linking of collagen is associated with aortic wall matrix stiffness in the pathogenesis of drug-induced diabetes in rats

Diabetes mellitus is major risk factor for cardiovascular disease, and atherosclerosis accounts for most of the morbidity and mortality of diabetic patients. To examine the effects of diabetes on the vessel wall, we examined the association of collagen cross-linking in relation to matrix stiffness of the descending aorta in streptozotocin-induced diabetic rats. The matrix stiffness of the vessel was determined by measuring the tensile properties of the tissue. Seven weeks following the establishment of diabetes, both control and diabetic rats were killed and the descending aortas were excised and analyzed. The findings from biomechanical analysis indicated a significant increase in maximum load (26%), stress (22%), Young's modulus of elasticity (60%), and toughness (32%) in diabetic aortas compared to control. In contrast, the maximum strain of the diabetic rat aorta was significantly reduced by 20% compared to control rats, suggesting stiffening of the blood vessel. The results from biochemical analysis showed that the amount of total collagen increased by 21% in diabetic tissues compared to the control. The sequential extractions of collagen showed that the diabetic specimens yielded 34% more neutral salt-soluble collagen (NSC) than the control. The amount of pepsin-soluble collagen was 31% less in diabetic tissues than in the control group, whereas the amount of insoluble collagen (ISC) increased by 56%. A significant accumulation in advanced glycation end products (AGEs) were seen in pepsin- and collagenase-soluble collagen in diabetic vessel. Furthermore, the altered biomechanical properties of the vessel wall were strongly correlated with the biochemistry of collagen. Overall, these results provide evidence that the diabetic state is associated with the changes in collagen biochemistry and in the biomechanics of the blood vessel.

Changes in collagen fractions of the myocardium induced by mechanical unloading of the left ventricle

Improvement of cardiac function by mechanically (assist device) unloading the left ventricle is frequently observed. The composition of different collagen fractions in the extracellular matrix may be a predictor for improvement. We examined the Neutral Salt Soluble Collagen (NSC) and the Acid Soluble Collagen (ASC) content in the myocardium taken from three groups of patients. Group 1 (N 18): healthy myocardium (unused donor hearts). Group 2 (N 8) comprises two subgroups, 2a: myocardium of patients taken at device implantation; 2b: myocardium of the same patients taken at device removal (transplantation). Group 3 (N 10): myocardium taken at device placement from later on weaned patients (no transplantation). The mean amount of NSC of group 1 (3.61 g/mg) did not significantly differ from that of group 2a (3.16 g/mg), P 0.507, however, after unloading it showed a significant increase in group 2b (4.69 g/mg), P 0.2000. At device placement, group 3 showed significant higher amounts of NSC (5.63 g/mg) than in group 1 (P 0.0001) and than in group 2a and 2b (P 0.002; P 0.028). As compared to group 1 (1.77 g/mg), the mean amount of ASC was significantly reduced in group 2a (0.20 g/mg), P 0.0001 and after unloading in group 2b (0.62 g/mg), P 0.001. Groups 1, 3 (1.31 g/mg) and 2b (0.62 g/mg) did not differ statistically (P 0.114, P 0.060), but group 2a showed a statistically lower ASC content than group 3, P 0.002. Ejection fraction (EF) before device placement was below 20% in all patients. At device removal, the EF of group 2 patients (transplanted) was below 35% and group 3 patients (weaned off ) above 50%. Mechanical unloading leads to an increase of NSC and ASC. It leads to NSC increasing above the level of the control myocardium. ASC remains significantly below the control level. The increase in collagen fractions could be regarded as an expression of the reverse of the maladaptive myocardial interstitial remodeling. The weaned patients already showed a hyper-normal NSC and a normal ASC fraction at implantation. This collagen constellation seemed to cope better with the disease than that of the other patients.

Glycation-Induced Matrix Stability in the Rabbit Achilles Tendon

Connective tissue susceptibility to nonenzymatic glycation was examined following 0, 2, 4, 6, 8, and 10 weeks of incubating the rabbit Achilles tendon in phosphate-buffered saline containing ribose (glycated). The biomechanical integrity of the glycated tendons was then compared to control tendons incubated in phosphate-buffered saline (nonglycated) at each time interval, while the biochemical stability of both groups of tendons was determined by examining collagen extractability and the formation of pentosidine at 8 weeks. Whereas there were no significant biomechanical differences between control and glycated tendons at 0- and 2-week intervals (P > 0.05), moderately significant increases in maximum load, energy to yield, and toughness of glycated tendons were observed at 4 weeks. Beyond 4 weeks of incubation, the differences between glycated and nonglycated tendons became highly significant, as glycated tendons withstood more load and tensile stress (P < 0.01 for each variable), attained significantly higher modulus of elasticity (P < 0.01), absorbed more energy (P < 0.01), and became tougher (P < 0.01) than controls. These differences in the biomechanical indices of the effects of glycation were stable between the 6th and 10th week of glycation. The maximum increases in the biomechanical measurements as a result of glycation were 29% for maximum load, 125% for stress, 19% for strain, 106% for Young's modulus of elasticity, 14% for energy to yield, and 57% for toughness. Biochemical analysis showed a 61% reduction in the extractability of neutral salt-soluble collagen, a 48% decrease in acid-soluble collagen, and a 29% decline in pepsin-soluble collagen in glycated tendons (P < 0.01). In contrast, there was a 28% increase in the amount of insoluble collagen and significantly higher amounts of pentosidine (P < 0.01) in glycated tendons. Collectively, these biomechanical and biochemical results suggest that nonenzymatic glycation may explain the altered stability of connective tissue matrix induced by the processes of diabetes and aging.

Laser photostimulation accelerates wound healing in diabetic rats

In this study, we examined the hypothesis that laser photostimulation can facilitate healing of impaired wounds in experimental diabetes using a rat model. Diabetes was induced in male rats by streptozotocin injection and two 6 mm diameter circular wounds were created on either side of the spine. The left wound of each animal was treated with a 632.8 nm He:Ne laser at a dose of 1.0 J/cm2 for five days a week until the wounds closed (three weeks). Measurements of the biomechanical properties of the laser-treated wounds indicated there was a marginal increase in maximum load (16%), stress (16%), strain (27%), energy absorption (47%) and toughness (84%) compared to control wounds of diabetic rats. Biochemical assays revealed that the amount of total collagen was significantly increased in laser treated wounds (274 +/- 8.7 microg) over the control wounds (230 +/- 8.4 microg). Sequential extractions of collagen from healing wounds showed that laser treated wounds had significantly greater concentrations of neutral salt soluble (15%) and insoluble collagen (16%) than control wounds, suggesting accelerated collagen production in laser treated wounds. There was an appreciable decrease in pepsin soluble collagen (19%) in laser treated wounds over control wounds, indicating higher resistance to proteolytic digestion. In conclusion, the biomechanical and biochemical results collectively suggest that laser photostimulation promotes the tissue repair process by accelerating collagen production and promoting overall connective tissue stability in healing wounds of diabetic rats.

Characterization of dermal type I collagen of C3H mouse at different stages of the hair cycle.

Hair follicles develop or regress in accordance with the hair cycle. In this study, we partially characterized fibrillar type I collagen, the predominant component in the dermis, at two stages of the hair cycle: anagen and telogen. Skin samples were obtained from the backs of two groups of 11-week-old C3H mice: one at anagen stage induced by shaving and the other at telogen stage. The amount of neutral salt-soluble (newly synthesized) collagen obtained from anagen skin was about twofold that from telogen skin, while the level of acid-soluble collagen was not significantly different between the two groups. The degree of lysine hydroxylation of pepsinized type I collagen obtained from anagen skin was significantly higher than that in telogen (5.0% higher in alpha1 chain, and 15.6% higher in alpha2 chain). Proline hydroxylation at the anagen stage was also slightly higher than in the telogen stage. Two major collagen cross-links were found in both groups of skin; dehydro-hydroxylysinonorleucine and dehydro-histidinohydroxymerodesmosine. The concentration of the latter, a complex tetravalent cross-link, was significantly lower in anagen skin when compared with telogen skin (mean +/- SD 0.64 +/- 0. 07 vs. 0.78 +/- 0.06 mol/mol collagen). The former showed no significant difference between the two groups. In addition, a significant amount of lysyl-aldehyde (a cross-link precursor) was found in anagen (0.16 +/- 0.02 mol/mol collagen), while it was 0.12 mol/mol collagen in telogen. These results indicate that the remodelling of collagen is more active in anagen skin than in telogen, and that characteristic post-translational modifications of dermal collagen seen in anagen may play a part in facilitating an environment around hair follicles for their migration and growth.

Laser photostimulation of collagen production in healing rabbit Achilles tendons.

Low energy laser photostimulation at certain wavelengths can enhance tissue repair by releasing growth factors from fibroblasts and stimulate the healing process. This study was designed to evaluate the influence of laser photostimulation on collagen production in experimentally tenotomized and repaired rabbit Achilles tendons. A total of 24 male New Zealand rabbits, ages 10-12 weeks, were used. Following tenotomy and repair, the surgical hind limbs of the rabbits were immobilized in customized polyurethane casts. The experimental animals were treated with a 632.8 nm He:Ne laser daily at 1.0 J cm(-2) for 14 days. Control animals were sham treated with the laser head. On the fifth day after repair, the casts were removed to allow the animals to bear weight on the lower extremity. The animals were euthanized on the 15th postoperative day, then, the Achilles tendons were excised, processed and analyzed. Biochemical analyses of the tendons revealed a 26% increase in collagen concentration with laser photostimulation indicating a more rapid healing process in treated tendons compared to controls. Sequential extractions of collagen from regenerating tissues revealed that the laser photostimulated tendons had 32% and 33% greater concentrations of neutral salt soluble collagen and insoluble collagen, respectively, than control tendons suggesting an accelerated production of collagen with laser photostimulation. A significant decrease (9%) in pepsin soluble collagen was observed in laser-treated tendons compared to controls. There were no statistically significant differences recorded in the concentrations of hydroxypyridinium crosslinks and acid soluble collagen between treated and control tendons. This study of laser photostimulation on tendon healing in rabbits suggests that such therapy facilitates collagen production in a manner that enhances tendon healing.

Regulation of Collagen Matrix Reconstruction by Phosphated Polysaccharides.

Phosphated polysaccharides were used to regulate the rate of matrix reconstruction from a neutral-salt soluble collagen (NSC) solution and the stability of the matrix in comparison with various chemically modified polysaccharides. Non-charged polysaccharides such as soluble starch, dextran, acetyl starch, hydroxypropyl starch and starch propylsulfonate had no substantial effect on matrix reconstruction. The positively charged polysaccharides, 3-trimethylamino-2-hydroxypropyl starch, slightly accelerated the molecular rearrangement, but was little incorporated in the matrix and had no effect on the thermal denaturation behavior of the matrix. Sulfated polysaccharides, polyanions, caused reconstruction without a lag phase because of too-rapid aggregation, and resulted in the reconstruction of the matrix with a less stable and heterogenous macrostructure. However, starch phosphate and dextran phosphate markedly accelerated the reconstruction of the matrix with uniform intermolecular cohesion similar to that of the control. This regulatory function of phosphated polysaccharides was enhanced with decreasing the pH from 9 to 5.

The Effect of Prenatal Protein-Energy Malnutrition on Collagen Metabolism in Fetal Bones

We analyzed various biochemical variables of the bones in fetal rats whose dams were protein-energy malnourished. Dams were randomly divided into two groups and fed either a 6% protein diet as a malnourished group or a 20% protein diet as a control, from day 13 of gestation to day 22, when fetuses were removed. Hexosamine and hydroxyproline contents of the calvaria and hexosamine contents of long bones were greater in the malnourished group than in the controls. Sequential extractability of collagen differed among various bones in the malnourished group and controls. The ratio of alpha:beta obtained from SDS-polyacrylamide gel of neutral salt-soluble collagen tended to increase in the long bones and mandible, and decrease in the calvaria and ribs in the malnourished group. Also, the ratio of alpha 1:alpha 2 tended to be lower in the malnourished group than in the control group in all bones. Protein-energy malnutrition during pregnancy has shown to affect biochemical composition of various fetal bones.

Levels of calcium and soluble collagen in turkey egg shell membranes

1. 1. This experiment examined the effects of weeks in egg production and type of housing confinement of turkey hens on calcium and soluble collagen levels in egg shell membranes; and discussion was given to their apparent relationship to gas exchange in turkey eggs. 2. 2. The high level of acid-soluble collagen in inner and outer egg shell membranes of aging caged hens compared with the same aged floor-penned hens may have a relationship with the low hatchability generally recognized in caged hens. 3. 3. The levels of calcium found in the outer shell membrane are low and appeared to decrease with the age of the hen. 4. 4. There were no differences over time in levels of total collagen and neutral salt-soluble collagen (newly formed collagen) found in egg shell membranes of turkey hens confined in cages or floor pens. 5. 5. It is suggested that the acid-soluble collagen levels found in inner shell membranes may have a relationship in limiting respiratory gas exchange during latter incubation time, and thus limit embryo survival.

A subcutaneous implant for wound healing studies in humans

A method for studying wound healing in humans is described. The technique is based on the production of a standard subcutaneous injury during implantation of a retrievable sponge. The injury is produced by introducing a small device (PVA implant) which consists of a 5.7-cm piece of perforated silicone tubing containing two pieces of polyvinyl alcohol sponge. The sponge provides a site for attracting inflammatory cells with subsequent fibroblast infiltration. The sterile PVA implant is inserted subcutaneously in the upper arm by means of a 12-gauge needle and remains there for 14 days. Upon removal, one sponge has hydroxyproline deposition quantitated using a high-performance liquid chromatography technique. The other sponge may be processed for light and electron microscopy or specialty staining. Other collagen determinations such as percentage neutral salt-soluble collagen are also possible. Using the PVA implant has made it possible to follow the kinetics of collagen deposition in the rat. There was a marked increase in collagen accumulation from Day 2 (0.89 nmole/mg sponge) to Day 14 (18 nmole/mg sponge) in the rat. Collagen deposition was also measured in human control subjects (5.07 nmole/mg sponge, n = 12) and compared to trauma patients (2.04 nmole/mg sponge, n = 5). Histologic staining showing fibroblast infiltration and collagen deposition correlated well with the biochemical findings. This implant, coupled with recent HPLC technology, provides a safe, acceptable technique to study human would healing parameters and overcomes many of the limitations of previous methods.

歯髄コラーゲンおよびその分解産物の単球走化誘起能について

The purpose of this study was to investigate the chemotactic activity of the bovine dental pulp collagen and collagen peptide fragments using human blood monocytes. Chemotactic assays were performed with the modified Boyden's method. The results obtained were as follows: 1) Non-collagenous proteins and neutral salt-soluble collagen from the bovine dental pulps did not show a high chemotactic activity. 2) Neutral salt-soluble collagen from the skin of fetal bovine had a greater chemotactic activity than that from the dental pulp. 3) Peptide fragments obtained by digesting the above three proteins with CNBr showed a greater chemotactic activity than those proteins themselves. 4) Peptide fragments obtained by digesting theinsoluble proteins, most of which consisted of insoluble collagen, with CNBr had a great chemotactic activity on the human monocytes. The molecular weight of these peptide fragments was thought to be less than 14, 000. 5) Solubilized proteins obtained by digesting insoluble proteins with trypsin and demineralizing them with EDTA showed a greater chemotactic activity on the human monocytes than the peptide fragments derived from the insoluble collagen digested with trypsin.

Studies on collagen in the experimental myocardial infarction.

The healing process of myocardial infarction was studied with special reference to the production of collagen by the determinations of prolylhydroxylase (PHase) activity and contents of hydroxyproline (Hyp) of the collagen subfractions, i.e., neutral salt soluble (NSC), acid soluble (ASC) and insoluble collagens (ISC) in comparison with histologic observation of the proliferation of connective tissue in the canine heart. The contents of Hyp in NSC and ASC increased in the infarcted tissue 5 days after coronary ligation prior to its increase in ISC. The content of Hyp in NSC, which is a precursor of ASC and ISC, reached the maximal value at 7 days and that reached the maximal value in ASC 7 to 14 days after coronary ligation. ISC increased markedly after 14 days. PHase activity increased on the second day and attained to the highest at 5 to 7 days. Proliferation of fibroblasts paralleled with the increases in PHase activity and NSC, and an increase in collagen fiber did with the contents of ISC. From these results it is concluded that in the healing stage of myocardial infarction, PHase was activated earlier than the increase in immature collagen in the infarcted area which was observed 5 to 7 days after coronary ligation and was followed by the production of mature collagen after 14 days.

Kinetic Analysis of Collagen Fibrillogenesis: II. Corneal and Scleral Type I Collagen

Fibril formation of neutral salt soluble and pepsin-treated type I collagen from rabbit corneal stroma or sclera was compared using a turbidimetric analysis which permits the determination of apparent rate constants and activation energies for the lag and growth phase of collagen fibrillogenesis. Information regarding the lateral growth of fibrils was obtained from the final turbidity values. Neutral salt soluble corneal collagen had smaller rate constants for both the lag and growth phases of fibrillogenesis than scleral collagen. Pepsin treatment decreased the rate constants for both collagens proportionately. The activation energies were higher for type I collagen from cornea than sclera. Pepsin treatment increased the activation energy for both phases of corneal fibril formation but only the growth phase of scleral collagen fibrillogenesis was affected. The extent of lateral fibril growth was compared using the intrinsic turbidity values which are related to the mass per unit fibril length. Neutral salt soluble scleral type I collagen had a significantly higher intrinsic turbidity than did neutral salt soluble corneal collagen indicating that scleral collagen formed thicker fibrils; however, this difference was not retained after pepsin treatment, demonstrating that a helical-telopeptide interaction occurs in corneal type I collagen which influences fibril diameter. The observed differences in the rate constants, activation energies and intrinsic turbidity values indicates that there are molecular differences which are responsible for fibrillar differences of corneal and scleral type I collagens.

Increases in lung prolyl hydroxylase and superoxide dismutase activities during bleomycin-induced lung fibrosis in hamsters

Intratracheal administration of bleomycin causes pulmonary fibrosis in hamsters. Using this model the activities of lung prolyl hydroxylase and superoxide dismutase and the accumulation of neutral salt soluble and insoluble collagens have been determined. One unit of bleomycin was injected intratracheally to hamsters, whereas control animals received an equivalent volume of sterile saline by the same route. Total lung prolyl hydroxylase activity was significantly elevated at all times following bleomycin treatment. The activity was increased as early as 2 days, peaked to a maximum value of 400% of the control at 14 days, followed by a sharp decline to 235% and 180% of the control activity at 21 and 28 days after bleomycin treatment, respectively. Except for the earliest time (2 days), lung prolyl hydroxylase specific activity was also significantly elevated at all times after bleomycin treatment. A significant increase in both total and specific activities of lung superoxide dismutase was also observed at all times after bleomycin treatment. Total activity peaked to a maximum value of 315% of the control activity at 14 days and the specific activity to a maximum value of 190% of the control at 21 days after bleomycin treatment. Thereafter, both activities declined, but were still significantly elevated over the control at 28 days after the treatment. Lung proline pool size was significantly increased at all times and attained a maximum value of 372% of the control at 14 days after bleomycin treatment. Increases in the lung prolyl hydroxylase and superoxide dismutase activities and in the proline pool size preceded the significant increases in neutral salt soluble and insoluble collagens which occurred at 7 days after bleomycin treatment and continued to be significantly elevated for the remaining period of the study.

Effect of Protein Malnutrition on the Cross-linking of Gingival and Uterine Collagen in Rats

The effect of protein malnutrition on gingival and uterine collagen cross-linking was studied in female albino rats. The gingival and uterine samples were taken on the 28th day of the experimental period in both groups; the percent reversibility of neutral salt-soluble collagen, and the susceptibility of insoluble collagen to denaturing agents (to KSCN, urea, or pronase) were increased in gingival and uterine samples of protein-deficient animals. The analysis of gingival and uterine samples showed α1 and α2 subunits of neutral salt-soluble collagen appreciably increased, but β11 and β12 chains and the aldehyde content were significantly decreased in protein-deficient animals compared to controls. The results indicate that in protein deficiency the cross-linking and maturation of collagen are impaired.

Influence of bleomycin on the metabolism of dermal collagen in albino rats

The metabolism of collagen in male rats by treatment with bleomycin was studied following the injection of [ 3H]proline and the determination of specific and total activity of [ 3H]hydroxyproline in skin collagen fractions and urine. In the case of the bleomycin-treated animals, there was found to be an increase in the neutral salt soluble collagen content with no change in insoluble collagen content as compared to the control group. The specific and total radioactivity of [ 3H]hydroxyproline in soluble and insoluble collagen fractions was also increased. Examination of [ 3H]hydroxyproline activity in soluble and insoluble collagen showed that the conversion of soluble to insoluble collagen was improved by the bleomycin-treated group. It was found that this was accompanied by a decrease in urinary excretion of total hydroxyproline and [ 3H]hydroxyproline during the first 12 hr after the administration of [ 3H]proline. Therefore, the results of the present investigation clearly indicate that the maturation of soluble to insoluble collagen is promoted and accompanied by a decrease in the catabolism of soluble collagen in the bleomycin-treated animals. In addition, administration of bleomycin increased the synthesis of collagen.

Bioflavonoid-mediated stabilization of collagen in adjuvant-induced arthritis.

In rats with adjuvant-induced arthritis, the effect of (+)-catechin (CA) and 0-(beta-hydroxyethyl) rutosides (HR) on the crosslinking of collagen was studied. Compared with controls the arthritic group showed an increase in the reversibility of neutral salt-soluble collagen gel, solubility of acid-insoluble collagen to denaturing agents and the ratio of alpha/beta subunits of neutral salt-soluble collagen. These results suggest an impaired maturation of collagen in arthritic animals. Administration of CA or HR to the arthritic animals caused a decrease in the reversibility of collagen gel and in the solubility of acid-insoluble collagen to denaturing agents and to pronase. In addition, the electrophoretic patterns of neutral salt-soluble collagen on SDS polyacrylamide gels also showed a decrease in the alpha/beta ratio. All these results may collectively indicate that both flavonoids promote the crosslinking of collagen in arthritic animals.

Influence of streptozotocin- and alloxan-induced diabetes on the crosslinking of dermal collagen.

The influence of experimentally induced diabetes on rat dermal collagen crosslinking was investigated in male albino rats. In comparison to the normal, the diabetic group demonstrated decrease in percent reversibility of neutral salt-soluble collagen gel and susceptibility of insoluble collagen to denaturing agents and pronase whereas the aldehyde content was significantly increased. The electrophoretic gels revealed a marked decrease of alpha/beta ratio in diabetic animals. The results indicated that both the intra- and intermolecular crosslinkings of collagen were increased in experimentally induced diabetes.