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PubMed1969

The quantitative relationship of urinary peptide hydroxyproline excretion to collagen degradation.

To determine the quantitative relationship of urinary hydroxyproline peptide excretion to collagen breakdown, known quantities of radioactive hydroxyproline peptides were administered to unlabeled animals and excertion of radioactivity in respiratory carbon dioxide, urine, and feces was measured. The major routes of excretion of collagen peptide metabolites were respiratory carbon dioxide (75%) and urine, as hydroxyproline-containing peptides (25%). Since the predominant urine hydroxyproline peptide linkage is proly-hydroxyproline, L-prolyl-L-hydroxyproline-(3)H was administered to unlabeled animals. Greater than 80% of the administered dipeptide was excreted in urine, suggesting that this peptide linkage is not hydrolyzed to a significant extent in vivo. These data suggest that urinary hydroxyproline excretion is a "fairly" sensitive indicator of collagen breakdown and can be used at the clinical level to quantitate changes in collagen breakdown.

Animals↗
PubMedDate not supplied

Elevated plasma hydroxyproline. A possible risk factor associated with connective tissue injuries during overuse.

Basal plasma hydroxyproline was measured in 104 male Navy Seal candidates 1 week into their intense physical training program, which lasted 7 weeks, and correlated to the incidence of connective tissue injuries incurred later in the training program. Eleven subjects (10.6%) were diagnosed as having connective tissue injuries. Those subjects with connective tissue injuries had a significantly higher (P less than 0.05) mean plasma hydroxyproline value (4.02 micrograms/ml) than subjects without injury (3.10 micrograms/ml). The majority of graduates (75%) had plasma hydroxyproline values less than 3.3 micrograms/ml. These graduates represented the strongest and most enduring injury-free subjects. Of the subject pool who incurred connective tissue injuries, only 27% had plasma hydroxyproline values less than 3.3 micrograms/ml. The majority of the injured subjects (73%) had plasma hydroxyproline values greater than or equal to 3.3 micrograms/ml. In conclusion, there is a relationship between initial training basal plasma hydroxyproline levels and connective tissue injuries later incurred in an intense physical training program. These data suggest that elevated plasma hydroxyproline levels may represent a risk factor associated with connective tissue injuries.

Adolescent↗
PubMed1999

Effects of Sho-saiko-to extract on liver fibrosis in relation to the changes in hydroxyproline and retinoid levels of the liver in rats.

To examine the effects of Sho-saiko-to extract on liver fibrosis, the drug was administered to rats with dimethylnitrosamine-induced liver-injury at various doses. Hydroxyproline and retinoid levels in the liver were measured as indicators of liver function. In liver-injured rats, the hydroxyproline level in the liver (957+/- 154nmol g(-1)) was about 4.16-times that found in normal liver (230+/-11 nmol g(-1)), but administration of Sho-saiko-to extract (0.75%, 1.5% or 3%) reduced the hydroxyproline level significantly (554+/-58, 356+/-51, 374+/-66nmol g(-1), P<0.01). Single administration of the active constituents of Sho-saiko-to extract, glycyrrhizin, baicalin or baicalein, decreased the hydroxyproline level significantly compared with the ordinary food group (P < 0.05), but the decrease was smaller compared with the Sho-saiko-to extract group. The liver retinoid level was higher in the Sho-saiko-to extract group than the ordinary food group and the value increased dose-dependently. A significant negative correlation, r=-0.814 (P<0.001) was detected between the hydroxyproline level and retinoid level in the liver of liver-injured rats. Significant negative correlations, r =-0.728 (P < 0.001) and r= -0.873 (P < 0.001), were also detected between the liver hydroxyproline level and the liver concentrations of the active constituents (glycyrretic acid, baicalin and baicalein) in the liver-injured rats. From these findings, it was considered that the liver concentrations of hydroxyproline and retinoid as well as the active constituents were involved in the improvement of liver fibrosis in the liver-injured rats administered Sho-saiko-to extract. Administration of Sho-saiko-to extract inhibited collagen production while an increase in retinoid level inhibited activation of Ito cells leading to inhibition and prevention of liver fibrosis.

Administration, Oral↗
PubMed1982

[The influence of diet on the urinary excretion of total, free and conjugated non-dialysable hydroxyproline in the normal adult (author's transl)].

Having established reference values for the different forms of urinary excretion of hydroxyproline on the basis of the results obtained in 12 normal subjects aged from 20 to 43 years, the authors studied the influence of three diets on the excretion of total, free and conjugated hydroxyproline. Urinary levels of total and peptide dialysable hydroxyprolines were found to be highly sensitive to the dietary collagen content, whilst free hydroxyproline increased only when the dietary collagen intake was very high. Peptide non-dialysable hydroxyproline, corresponding to peptides of molecular weight 6000-8000 was not influenced by diet. The ratio dialysable hydroxyproline/non-dialysable hydroxyproline varied from reference values if the diet was loaded with collagen. Only the ratio free hydroxyproline/non-dialysable hydroxyproline seemed to be independent of diet in the subjects studied.

Adult↗
PubMed1994

Spectroscopic study of hydroxyproline transport in rat kidney mitochondria.

Hydroxyproline uptake by rat kidney mitochondria is here first shown by monitoring the reduction of the intramitochondrial pyridine nucleotides which occurs as a result of metabolism of imported hydroxyproline via hydroxyproline oxidase and 3-hydroxy-pyrroline-5-carboxylate dehydrogenase. Widely used criteria for demonstrating the occurrence of carrier-mediated transport were applied to this process. Hydroxyproline uptake shows saturation features (Km and Vmax values, measured at 20 degrees C and at pH 7.20, were found to be about 1.4 mM and 5 nmoles/min x mg mitochondrial protein, respectively) and proves to be inhibited by the impermeable compound phenylsuccinate, but insensitive to externally added methylglutamate. Difference found in the Km and Vmax values, a different inhibitor sensitivity and the failure of hydroxyproline to cause efflux of glutamate from the mitochondria show that hydroxyproline enters mitochondria by means of a translocator different from those which transport proline.

Animals↗
PubMed1978

[Analysis of the hydroxyproline containing serum proteins in connective tissue diseases (author's transl)].

The serum proteins were fractionated on Sephadex G 200, their hydroxyproline content determined. Three protein peaks (I-III) containing different hydroxyproline concentrations could be separated. In connective tissue disorders accompanied by increased collagen synthesis or collagen degradation, an elevation of hydroxyproline was found in peak II. An increase of hydroxyproline always was associated with an increase of the serum proteins in the same fractions. Hydroxyproline serum levels above normal obviously are due to a certain capacity of the serum proteins binding free hydroxyproline and collagen metabolites.

Blood Proteins↗
PubMed1984

A criterion to determine whether cis-4-hydroxyproline is produced in animal tissues.

Hydrolyzates of tissues that had been labeled with [14C]proline often contain significant amounts of cis-4-hydroxy[14C]proline. Since animal cells do not contain an enzyme which can effect formation of cis-4-hydroxyproline, there are only two possible explanations for its presence. Either it is formed during acid hydrolysis of trans-4-hydroxyproline (which is synthesized by cells and is a common constituent of connective tissues), or it is produced by a nonenzymatic mechanism such as attack by oxygen radicals. It is important to resolve this issue because if a nonenzymatic mechanism is active in connective tissues, then it will be necessary to reevaluate currently accepted ideas about production of hydroxyproline. This communication describes a method for distinguishing between the two alternate explanations. Tissues or cells are labeled with [14C]proline, and then a known amount of trans-4-hydroxy[3H]proline is added to each sample before hydrolysis; the relative amounts of [14C]- and [3H]-cis-4-hydroxyproline are compared after hydrolysis. It is known from a separate series of measurements with mixtures of [14C]- and [3H]-trans-4-hydroxyproline standards that there is a very high correlation (r = 0.998) between acid-induced formation of the [14C]- and [3H]-cis epimers. One can thus compare the amount of cis-4-hydroxy[14C]proline in a hydrolyzate from a biological system with the amount that would be expected if it were all formed during acid hydrolysis. This method was used to show that fibroblasts cultured under conditions commonly used to study collagen metabolism do not produce cis-4-hydroxyproline. This result strongly suggests that nonenzymatic hydroxylation does not normally occur in cell culture systems.

Carbon Radioisotopes↗
PubMed1981

Plasma hydroxyproline fractions in patients with dialysis osteodystrophy.

Plasma hydroxyproline fractions were measured in 17 normal subjects and in 54 patients on maintenance haemodialysis therapy (MHT) with various degrees of dialysis osteodystrophy. On the basis of both radiological and histological findings these patients were divided into three groups: radiologically normal, histologically normal and those with osteitis fibrosa. The mean total plasma hydroxyproline concentrations were significantly elevated in all groups of MHT patients. However, these increases were mainly due to peptide-bound and free hydroxyproline fractions. The highest values for these two fractions were found in patients with osteitis fibrosa. The free to peptide-bound hydroxyproline ratio was not significantly altered in the majority of patients on dialysis; the mean ratio was significantly lower in patients with osteitis fibrosa when compared with patients with no histological evidence of bone disease. This finding would suggest that there is no inhibition of hydroxyproline catabolism in patients on haemodialysis and the measurements of both free and peptide-bound hydroxyproline were equally sensitive in identifying patients with osteitis fibrosa.

Bone Diseases↗
PubMed1996

A simplified method for the analysis of hydroxyproline in biological tissues.

A critical study of the different steps involved in previous procedure for hydroxyproline assay allows the direct measurement of collagen content in tissue' homogenates without losing the advantages of the method. The procedure is based on alkaline hydrolysis of the tissue homogenate and subsequent determination of the free hydroxyproline in hydrolyzates. Chloramine-T was used to oxidize the free hydroxyproline for the production of a pyrrole. The addition of Ehrlich's reagent resulted in the formation of a chromophore that can be measured at 550 nm. Optimal assay conditions were determined using tissue homogenate and purified acid soluble collagen along with standard hydroxyproline. Critical parameters such as the amount of chloramine-T, sodium hydroxide, p-dimethylaminobenzaldehyde, pH of the reaction buffer, and length of oxidation time were examined to obtain satisfactory results. The method has been applied to samples of tissue homogenate and purified acid soluble collagen, with recovery of added hydroxyproline of 101 +/- 6.5 and 104 +/- 6.0 (SD) percent, respectively. The method is highly sensitive and reproducible when used to measure the imino acid in tissue homogenates. The modified hydroxyproline assay presented in this communication will be useful for routine measurement of collagen content in extracts of various tissue specimens. In addition, the modified method can be used for batch processing of column fractions to monitor the collagen concentrations during purification.

Achilles Tendon↗
PubMed2000

The hydroxyproline content of amnion and prelabour rupture of the membranes.

OBJECTIVE: To determine whether prelabour rupture of the membranes is caused by a generalised reduction in amniotic collagen. SETTING: Leeds General Infirmary, UK. STUDY DESIGN: The hydroxyproline content of amnion from 55 women with prelabour rupture of the membranes was compared with that from 50 women whose membranes ruptured during labour and 28 women who were delivered by elective caesarean section. RESULTS: No association was found between hydroxyproline content and the timing of membrane rupture. Hydroxyproline content was inversely correlated with gestation. The hydroxyproline content per cm(2) amnion was significantly increased in cases which had laboured. There was a strong correlation between the mechanical properties and the hydroxyproline content of amnion. CONCLUSIONS: PROM is not associated with a generalised reduction in the hydroxyproline content of amnion.

Amnion↗
PubMed1976

Relevance of hydroxyproline excretion to bone metastasis in breast cancer.

In 181 consecutive patients with breast cancer, urinary hydroxyproline excretion has been critically evaluated in conjunction with clinical, biochemical, radiological and scintigraphic parameters. The urinary hydroxyproline/creatinine ratio is a sensitive index of the presence of bone metastases. Urinary hydroxyproline excretion is a reliable method of selecting those patients whose elevated serum alkaline phosphatase is secondary to bone disease rather than liver idsease. The estimation of hydroxyproline excretion furthermore gives information on the activity of bone metastasis, and its response to treatment, which cannot be given by radiological or scintigraphic methods. It is doubtful whether urinary hydroxyproline estimation will help to detect bone metastases before they are apparent on scintigrams. When the bone scan is doubtful, as often occurs in older subjects, hydroxyproline excretion has been found to be helpful in classifying the patient. When scintigraphy is not available, an elevation of hydroxyproline excretion, together with an elevation of Ca/cr ratio or alkaline phosphatase activity, may pre-date by several months the radiological demonstration of osseous metastases.

Adult↗
PubMed1985

Clinical significance of free plasma hydroxyproline measurement in metabolic bone disease.

Free hydroxyproline was measured in plasma of 67 normal subjects and in 70 patients with bone disease including primary hyperparathyroidism (n = 19), osteoporosis (n = 18), Paget's disease (n = 14), cancer involving bone (n = 8), chronic renal failure (n = 6), and osteomalacia (n = 6), and osteomalacia (n = 5). A good correlation was found between plasma and urinary values of the amino acid in normal subjects (r = 0.66; p less than 0.001). In patients with skeletal disorders a highly significant direct correlation was observed between free plasma hydroxyproline on the one hand and urinary hydroxyproline (r = 0.92; p less than 0.001) and serum alkaline phosphatase activity (r = 0.86; p less than 0.001) on the other, even though there were a few examples of dissociations among these parameters. Free plasma hydroxyproline decreased in the patients with Paget's disease following chronic administration of salmon calcitonin. Following successful parathyroidectomy, free plasma levels of hydroxyproline decreased in all the cases studied. Measurement of free plasma hydroxyproline thus appears to provide a specific index of bone metabolism that may be usefully employed as an alternative to the assay of other markers of bone turnover.

Adult↗
PubMed1978

Urinary hydroxyproline excretion in acromegaly.

The urinary hydroxyproline excretion of 8 acromegalics kept on a collagen free diet has been studied. Urinary hydroxyproline showed a circadian rhythm with maximum excretion rates at night and early morning and minimum excretion rates in the late afternoon and evening. When mean values are compared, there is a significant correlation between fasting GH-levels and 24 h urinary hydroxyproline excretion (Rs = 0.74 and between GH and fasting hydroxyproline:creatinine ratios (Rs = 0.76). No significant correlation could be demonstrated between collagen content of bone biopsies and GH-levels or urinary hydroxyproline excretion. It is suggested that a practical assessment of disease activity in acromegaly can be achieved through a simplified hydroxyproline assay.

Acromegaly↗
PubMed1975

Determination of hydroxyproline content in human heart muscle.

Hydroxyproline content was chemically determined on 45 human hearts obtained at autopsy in order to study whether hydroxyproline contents vary in proportion to heart weight or not. There were no significant differences in the mean hydroxyproline content between the epicardium and the endocardium, between the left ventricle and the right ventricle, or among cardiovascular and renal disease group, malignant disease group and miscellaneous disease group. Total hydroxyproline content per mg dry weight of the left ventricle increased proportionally with heart weight. Hydroxyproline content in neutral salt soluble fraction increased more proportionally than that in acid soluble fraction. This tendency was more evident in the endocardium than in the epicardium of the left ventricle. These results seem to be well consistent with those of histometrical estimation of scar tissue in human heart muscle. Hydroxyproline content in acid soluble fraction of the right ventricle was increased proportionally with heart weight.

Adolescent↗
PubMed1983

Peplomycin sulfate and pulmonary fibrosis: hydroxyproline, uronic acid, proline hydroxylase and glucosamine 6-phosphate synthetase in lungs of hamsters treated with peplomycin.

Effect of peplomycin sulfate (PLM) on pulmonary fibrosis was examined. Hydroxyproline, uronic acid, proline hydroxylase (EC 1.14.11.2) and glucosamine 6-phosphate synthetase (EC 2.6.1.16) in lungs of hamsters treated with PLM were studied and compared with those of hamsters treated with bleomycin (BLM). PLM, when administered intraperitoneally, one injection daily for 10 consecutive days, at either a high- (5 mg/kg) or low- (2.8 mg/kg) dosage-level, caused no significant increase of lung hydroxyproline and uronic acid as compared with controls. BLM on the other hand effected a significant increase in lung hydroxyproline on the high-dosage level (5 mg/kg) but not on the low-dosage level (2.8 mg/kg). In contrast, when administering PLM intratracheally, the concentrations of hydroxyproline in lungs increased 20% over the control levels. A transient increase of proline hydroxylase and glucosamine 6-phosphate synthetase also occurred shortly after the instillation. These increases were also observed in the corresponding groups treated with BLM, which confirmed the previous observations by other investigators. However, the magnitude of the increase was relatively lower in those values of PLM as compared with those of BLM. These data suggested that (1) PLM, when administered with multiple dosages intraperitoneally, showed no significant effect on the elevation of lung hydroxyproline; (2) PLM, when administered with a dose intratracheally, induced pulmonary fibrosis similar to that caused by BLM. However, the hydroxyproline accumulation in lungs of PLM-treated hamsters was less than in those of the BLM-treated; (3) The fibrotic effect on the lungs caused by either PLM or BLM was probably attributed to acceleration of the syntheses of collagen and acidic glycosaminoglycans.

Animals↗
PubMed1991

[Concentration of total and protein-bound hydroxyproline in serum and urine in patients with laryngeal neoplasms].

The concentration of total and protein-bound hydroxyproline in the serum and total hydroxyproline in the urine was measured in the group of 52 patients with larynx carcinoma by means of Prockop's and Udenfriend's colorimetric method modified by Tomaszewski. The mean level of total and protein-bound serum hydroxyproline in the group of clinical patients was statistically significantly raised when compared with control group. The mean level of total hydroxyproline in the urine of patients with larynx carcinoma was statistically insignificantly raised when compared with control group. The degree of clinical or organic progress of larynx carcinoma was not found to influence essentially the level of hydroxyproline in the serum of the examined patients. It was shown however, that statistically essential increase in the total hydroxyproline level in the urine depends on the degree of organic progress of larynx carcinoma.

Adult↗
PubMed1979

Urinary hydroxyproline in healthy patients and in prostate patients with and without bone metastases.

An essential part of the classification of prostate carcinoma is the diagnosis of bone metastases. This was done with 70 patients using x-ray analysis, scintography, determination of the acid and alkaline phosphatase, and pelvic crest biopsy, as well as aspiration of the pelvic and sternal bone marrow. In addition, the hydroxyproline concentration was determined in the 24-hour-urine. The study, which was initially undertaken on a sample group (n = 145), yielded a high correlation between age and sex and hydroxyproline values. Women before menopause show significantly lower values than do men of the same age. The data on patients with prostata cancer (n = 70) showed that patients with and without bone metastases, who had been treated with estrogens, had a significantly lower quantity of hydroxyproline than did patients who had not received estrogen therapy. Patients with skeletal metastases (n = 24) showed significantly higher hydroxyproline excretion in the urine than did those with prostate cancer without metastases, or healthy men of the same age (n = 35). Comparison of the results of hydroxyproline determination with the other diagnostic methods for demonstrating bone metastases showed that hydroxyproline determination was diagnostically on par with the scintigram. Pelvic crest biopsy, pelvic and sternal marrow aspiration can be considered valuable supplementary diagnostic procedures.

Bone Neoplasms↗
PubMed1979

Changes in connective tissue metabolism due to bone fractures in children aged 10--14 years. III. Urinary hydroxyproline and glycosaminoglycan excretion following bone fractures treated in different ways.

Urinary hydroxyproline and glycosaminoglycan excretion was studied in the course of fracture healing in twenty 10--14-year-old children with fracture of the femur or of the tibia and fibula. In 11 children reduction and plaster cast were used, in 5 osteosynthesis, and in 4 extension of 6--21 days duration combined with immobilization by plaster cast. In the state of resorption, an increase was observed in the urinary excretion of hydroxyproline and total glycosaminoglycans and in the six glycosaminoglycan fractions studied. As a result of the soft tissue damage accompanying the fracture, the excretion of hyaluronic acid, heparan sulphate, and dermatan sulphate showed the most marked increase. No statistically significant relationship could be demonstrated between the mode of treatment and the amounts of urinary hydroxyproline and glycosaminoglycan, but the stage of resorption was prolonged in the case of treatment with traction. At the beginning of regeneration, a decrease in hydroxyproline excretion and then an increase corresponding to the intensive synthesis of collagen were demonstrated. Glycosaminoglycan excretion during the development of fibrous callus was considerably below the control value. Excretion of the metabolites decreased markedly in the five operated patients. in the case of extension, the stage of regeneration prolonged by 4 to 10 days. In the stage of calcification, urinary hydroxyproline excretion was increased while the excretion of glycosaminoglycan and its fractions was the same as in the controls except in the patients treated with traction. No statistically significant relationship was found between the urinary amounts of hydroxyproline and glycosaminoglycan and the mode of treatment. In the case of traction, fracture repair and the formation of connective tissue callus were prolonged.

Adolescent↗