The Course of Selected Bone Resorption Marker Concentrations in Response to Short‐term Hypocalcemia Experimentally Induced with Disodium EDTA Infusions in Dairy Cows

The collagen metabolites hydroxyproline (HYP), deoxypyridinoline (DPD), and the carboxyterminal telopeptide of type I collagen (ICTP) are suitable markers for bone resorption in humans and several animal species. The purpose of this study was to describe the course of bone resorption markers during short‐term hypocalcemia induced with disodium ethylenediaminetetraacetic acid (Na₂EDTA) and to investigate whether bone resorption is increased in dairy cows under these conditions. EDTA infusions have been used as a model for periparturient paresis in dairy cows and to estimate the calcium mobilization rate from body reserves in ruminants. In this study, hypocalcemia was induced by means of a 5 % Na₂EDTA infusion (0.55 mg/kg/min Na₂EDTA for 5 h = total dose of 100.6 g). Two experiments were conducted: (1) Six 4–11 years‐old Brown Swiss cows were infused intravenously with EDTA for 5 h. Blood and urine samples were taken repeatedly from 1 day before until 10 days after infusion. (2) Towards the end of the lactation, the experiment was repeated with the same animals after a 14‐day‐period of feeding a low calcium diet (26 g/animal per day). The EDTA‐infusion induced hypocalcemia and hypophosphatemia. The HYP‐, DPD‐ and ICTP‐concentration remained mainly unaffected during both infusions. Only DPD showed an increase during infusion and HYP an increase 2 days after the infusion. In conclusion, the EDTA infusion had little effect on the concentrations of the measured bone markers, which may be due to the fact that the serum calcium pool was refilled by increased absorption of Ca via the gastrointestinal tract. From these results, it can be concluded that bone resorption was not influenced by EDTA infusion.     

Type I and III collagen degradation products in serum predict patient survival in head and neck squamous cell carcinoma

Cancer invasion induces extracellular matrix remodeling and collagen degradation. The aim of this study was to assess whether serum levels of type I and III collagen degradation products were associated with patient survival in head and neck squamous cell carcinoma (HNSCC). A novel enzyme immunoassay was developed for measuring type III collagen N-terminal telopeptide (IIINTP) in human serum samples. In addition, type I collagen C-terminal telopeptide (ICTP), matrix metalloprotease-8 (MMP-8) and tissue inhibitor of metalloproteases-1 (TIMP-1) were assessed in 205 blood samples from HNSCC patients. High levels of serum ICTP and IIINTP and plasma TIMP-1 were associated with poor survival. The concentration of ICTP was associated with levels of IIINTP and TIMP-1. The plasma concentration of MMP-8 was associated with tumor stage, but not with survival or levels of ICTP, IIINTP or TIMP-1 suggesting that other collagenases/proteases are responsible for the cleavage of type I and type III collagens. The rate of type I and type III collagen degradation is associated with patient survival and can be used as a prognostic marker in HNSCC.     

Circulating type I collagen degradation products: a new serum marker for clinical severity in patients with scleroderma?

Systemic sclerosis (SSc; scleroderma) results in the excessive deposition of extracellular matrix components in affected organs. This is partly due to enhanced synthesis; however, the role of degradative processes in this disease is still poorly understood. Sera of 32 patients with SSc (22 with the diffuse, 10 with the limited form) and of six patients with morphoea were assessed using radioimmunoassays for the cross-linked carboxy terminal telopeptide of type I collagen (ICTP) and for the amino terminal propeptide of type I procollagen (PINP) reflecting type I collagen degradation and synthesis, respectively. In 27 of the 32 patients with SSc, the concentration of ICTP was above the upper limit of the normal value (4.6 μg/L) and the mean level was clearly elevated at 7.92 μg/L. The ICTP concentration correlated with the skin score measuring the extent of the lesions, whereas no such correlation was found for PINP. The ICTP antigen in serum, studied by immunoblotting, had a molecular weight of about twice that of the trypsin-generated fragment isolated from human bone collagen. The mean concentration of serum PINP was 43.9 μg/L and no patient exceeded the upper limit of the normal range (80 μg/L).We report here for the first time that the concentration of the type I collagen degradation product ICTP in serum shows a close correlation with the extent of skin fibrosis in patients with SSc. We conclude that the increased deposition of type I collagen in this disease is accompanied by an increased turnover of this molecule, indicating a more complex derangement of synthetic and degradative processes than previously acknowledged.     

Serum markers of collagen synthesis and degradation in skin diseases. Altered levels in diseases with systemic manifestation and during systemic glucocorticoid treatment

Serum concentrations of the markers of collagen synthesis and degradation, collagen I propeptide (PICP), collagen III propeptide (PIIINP) and the cross-linked telopeptide of type I collagen (ICTP) were measured in young male dermatological patients and in control subjects. No significant differences were noted between patients suffering from atopic eczema (n=24), other eczemas (n=11), acne (n=8), psoriasis (n=7) or tinea (n=9) and the control subjects (n=24). In the total study population representing patients with common skin diseases and control subjects there was a significant correlation between the serum concentrations of PICP and PIIINP and between the concentrations of PICP and ICTP. This suggests that synthesis of type I and III collagens in vivo is coordinated and that the degradation and synthesis of type I collagen is balanced. These markers were also measured in older patients suffering from psoriasis, eczema and various connective tissue diseases. It was noted that the degree of skin involvement in these diseases was not related to the serum concentrations of the markers of collagen metabolism. The highest levels of PICP and PIIINP were observed in a patient with systemic mastocytosis (PICP 309 g/1 and PIIINP 8.0 g/1). Increased levels of PIIINP were also found in patients with a high alcohol consumption. We have previously demonstrated that systemic glucocorticoids reduce collagen propeptide levels in serum. In the present study we also proved that systemic gluocorticoids have no effect on collagen degradation. Thus the side effects of glucocorticoids, such as growth inhibition, skin atrophy and decrease in bone mass, are a result of inhibition of the synthesis of collagen and other macromolecules. The results indicate that local or generalized skin diseases do not markedly alter serum markers of collagen synthesis or degradation. The alterations in collagen metabolism determined by measurements in serum are thus mostly related to systemic involvement and medication (especially glucocorticoids).     

Basement membrane laminin and type IV collagen in various benign and malignant adnexal tumors of the skin: an immunohistochemical study

Thirty benign and seven malignant adnexal tumors of the skin and one lymph node metastasis were stained for laminin and type IV collagen with rabbit antibodies against the human basement membrane (BM) proteins using the immunoperoxidase technique. Fifteen of the benign sweat gland, sebaceous gland, and hair follicle tumors showed a continuous and distinct BM around the tumor aggregates. The cylindromas and eccrine spiradenomas seemed to produce excessive amounts of BM material, part of which was seen as amorphic patches within the tumor cell clusters, whereas the trichofolliculomas, trichoepitheliomas, and pilomatrixomas showed an absence of BM from many areas. In syringomas, in addition to the tubular structures surrounded by a continuous BM, undifferentiated cell nests containing granular BM material were present. They probably represent primitive structures obtaining during early development into tubules. The seven malignant tumors and the only metastasis studied here all contained small, narrow strips of BM material extracellularly between the infiltrating tumor clusters. Only in two cases was faint staining for laminin found within the cells. The pepsin pretreatment of the formalin-fixed, paraffin-embedded samples had most probably degraded the intracytoplasmic BM material in most cases. The BM defects were found to be associated with malignancy and low differentiation of the adnexal skin tumors, as reported previously for other tumor types, but a partial loss of BM was also associated with high differentiation in some benign adnexal tumors.     

Increased amount of type III pN-collagen in AAA when compared with AOD.

OBJECTIVE: the extent of the processing of type III procollagen to type III collagen was determined in nine human abdominal aortic aneurysms (AAA), and compared with ten samples of aortoiliac occlusive disease (AOD). METHODS: the aminoterminal propeptide (PIIINP) and telopeptide (IIINTP) of type III procollagen and collagen, respectively, were immunologically measured in the soluble and insoluble fractions of the extracellular matrix. The assay for PIIINP in the insoluble matrix was further validated. RESULTS: the insoluble matrices of AAAs contained at least 12 times more incompletely processed type III pN-collagen than AOD specimens (0.74% and 0.061%, respectively). Also, the soluble extracts of AAAs tended to contain more non-processed type III pN-collagen than free, properly cleaved aminoterminal propeptide. CONCLUSIONS: the larger amount of type III pN-collagen suggests an alteration in the metabolism of type III collagen in AAAs. This may partially explain the decreased tensile strength of the aortic tissue.     

Familial high serum PICP: high even in skin interstitial fluid, suppressible by topical corticosteroid treatment.

Circulating C-terminal propeptide of type I procollagen (PICP), mostly originating from bone, is mainly cleared by mannose receptors (MRs) in liver endothelial cells (LECs). We hypothesized that skin macrophage MRs could also play a role in local (in situ) clearance of PICP originating from skin type I procollagen synthesis. We tested this hypothesis in a male subject with a genetic systemic clearance defect, apparently due to an abnormality in MR function in LECs (or in PICP structure). Since skin macrophages may express the same MRs as LECs do, the genetic defect could affect them as well; hence, if elevated PICP concentrations even in skin interstitial fluid (IF) were found in our subject, it would suggest a role for local MR-mediated PICP clearance in skin. Since glucocorticoids (GCs) upregulate MRs in vitro, we measured the effect of topical GC on suction blister fluid (SBF)-PICP of the test person as compared with normal subjects. SBF-PICP was elevated in the case, which was consistent with the hypothesis. Furthermore, the GC-induced decrease was accentuated. The results suggest that skin macrophage MRs can have a role in skin PICP clearance in situ.     

Type I collagen turnover and cross-linking are increased in irradiated skin of breast cancer patients.

BACKGROUND AND PURPOSE: The effects of radiation therapy on the turnover and structure of type I collagen were studied in irradiated and contralateral skin of 18 breast cancer patients without clinically evident fibrosis. MATERIALS AND METHODS: The rates of on-going type I collagen synthesis and degradation were assessed by the aminoterminal propeptide of type I procollagen (PINP) and by two different assays (ICTP and SP4) for the carboxyterminal telopeptide of type I collagen in the soluble tissue extracts, respectively. Also, TIMP-1, TIMP-2 and the MMP-2/TIMP-2 complex were measured in the tissue extracts. Insoluble skin matrices, containing the cross-linked type I collagen fibres, were heat-denatured and digested with trypsin. Then, the variants of the carboxyterminal telopeptide of type I collagen were separated by high performance liquid chromatography (HPLC). The major histidinohydroxylysinonorleucine (HHL)-cross-linked variant was quantified by the SP4 assay, and the minor pyridinoline analogue (PA)-cross-linked telopeptide was quantified by the ICTP assay. RESULTS: Both the synthesis and degradation of type I collagen were increased (r=0.906; P<0.001) on the irradiated side, whereas the concentration of the MMP-2/TIMP-2 complex was decreased. In the insoluble tissue digests, the HHL-cross-linked telopeptides of type I collagen, also, when expressed/tissue hydroxyproline, were increased in the irradiated skin. TIMP-1, TIMP-2 or PA-cross-linked telopeptides of type I collagen showed no differences between the two sides. CONCLUSIONS: Radiotherapy induces a long-term increase in the turnover of type I collagen and leads to the accumulation of cross-linked type I collagen in skin.     

Amniotic fluid laminin and type IV collagen in normal and pathological pregnancies

Specific radioimmunoassays for the fragment P1 of human laminin and the 7-S collagen domain of human type IV collagen were used to quantify these basement membrane proteins in second trimester amniotic fluid samples from 21 normal and 41 pathological pregnancies, the latter group being defined by elevated amniotic fluid alpha-foetoprotein (AFP) or abnormal foetal karyotype, or both. The mean laminin P1 concentration in the normal 15 to 18-wk pregnancies was 36 micrograms/l (range 10-77) and that of the 7-S collagen was 46 micrograms/l (range 7-152). The molecular size of the antigens in amniotic fluid from both normal and pathological pregnancies, when assessed by gel filtration was very large, probably representing intact laminin and type IV collagen. Pathological pregnancies, e.g. cases of Turner syndrome, Meckel syndrome and anencephaly often had elevated amniotic fluid laminin and type IV collagen concentrations. A weak, but nevertheless significant, correlation was found between the amniotic fluid laminin and type IV collagen concentrations and also between type IV collagen and AFP, but none between laminin and AFP. In eight pregnancies with foetuses suffering from the congenital nephrotic syndrome of the Finnish type, a genetic disease assumed primarily to involve some component of the glomerular basement membrane, the amniotic fluid concentrations of both laminin and type IV collagen were within normal limits in spite of an elevated amniotic fluid AFP.