Absence of the DNA repair enzyme human 8-oxoguanine glycosylase is associated with an aggressive breast cancer phenotype.

BACKGROUND: 8-Oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) is the most abundant marker of DNA damage and it reflects oxidative stress. Human 8-oxoguanine glycosylase (hOGG1) is a DNA-repair enzyme that participates in 8-oxodG removal. METHODS: hOGG1 protein expression was immunohistochemically studied in 96 patients with local or locally advanced breast cancer and in 20 lesions of non-malignant breast disease. 8-OxodG levels had been previously determined in all patients. RESULTS: hOGG1 was overexpressed in invasive vs non-invasive lesions (P=0.006). 8-OxodG and hOGG1 had a significant inverse association (P=0.046). Lack of hOGG1 expression was associated with the most poor prognostic factors of breast cancer. In addition, all triple-negative breast carcinomas (TNBCs) were hOGG1 negative (P=0.027 vs non-TNBCs). Patients with a lack of both hOGG1- and 8-oxodG immunostaining showed extremely poor breast cancer-specific survival compared with those with either 8-oxodG- or hOGG1-positive tumours (P<0.000005). CONCLUSION: The current results imply that absence of hOGG1 expression is associated with features of aggressive breast cancer. Tumours lacking both 8-oxodG and hOGG1 seem to indicate especially poor prognosis.

Genitourinary prolapse and joint hypermobility are associated with altered type I and III collagen metabolism.

PURPOSE: The aim of this study was to determine whether benign joint hypermobility (BJH) is associated with urogenital prolapse and altered collagen metabolism. METHODS: 43 postmenopausal women with previous vaginal hysterectomy operated due to genitourinary prolapse were recruited. Each patient was also evaluated for joint hypermobility. The collagen metabolism was studied measuring serum concentrations of type I and III procollagen aminoterminal propeptides and trivalently cross-linked carboxyterminal telopeptide of type I collagen. RESULTS: Clinical joint hypermobility was found in 35% patients. Women with joint hypermobility had higher concentration of aminoterminal propeptide for type I procollagen and the values were statistically significant (P < 0.0178). Recurrent prolapse was found in 47% of the patients with BJH as compared to non-hypermobile group (25%). In this subgroup the results were statistically significant (P < 0.0085) for type III collagen. Also, the mean serum concentration for type III procollagen was significantly increased above the reference limit. CONCLUSIONS: Women with joint hypermobility have more recurrent genital prolapse as compared to women with normal joint mobility. Plain hypermobility was associated with higher concentrations for type I procollagen. Patients with recurrent prolapse and joint hypermobility have significantly high concentrations for type III procollagen.

8-Hydroxydeoxyguanosine: a new potential independent prognostic factor in breast cancer.

BACKGROUND: 8-Hydroxydeoxyguanosine (8-oxodG) is the commonly used marker of oxidative stress-derived DNA damage. 8-OxodG formation is regulated by local antioxidant capacity and DNA repair enzyme activity. Earlier studies have reported contradictory data on the function of 8-oxodG as a prognostic factor in different cancer types. METHODS: We assessed pre-operative serum 8-oxodG levels with an enzyme-linked immunosorbent assay in a well-defined series of 173 breast cancer patients. 8-OxodG expression in the nuclei of cancer cells from 150 of these patients was examined by immunohistochemistry. RESULTS: The serum 8-oxodG levels and immunohistochemical 8-oxodG expression were in concordance with each other (P<0.05). Negative 8-oxodG immunostaining was an independent prognostic factor for poor breast cancer-specific survival according to the multivariate analysis (P<0.01). This observation was even more remarkable when ductal carcinomas only (n=140) were considered (P<0.001). A low serum 8-oxodG level was associated statistically significantly with lymphatic vessel invasion and a positive lymph node status. CONCLUSIONS: Low serum 8-oxodG levels and a low immunohistochemical 8-oxodG expression were associated with an aggressive breast cancer phenotype. In addition, negative 8-oxodG immunostaining was a powerful prognostic factor for breast cancer-specific death in breast carcinoma patients.

Immunohistochemical study of matrix metalloproteinase 9 and tissue inhibitor of matrix metalloproteinase 1 in benign and malignant breast tissue--strong expression in intraductal carcinomas of the breast.

OBJECTIVE: Matrix metalloproteinases (MMPs) are involved in carcinogenesis due to their tissue remodeling capability, and there is convincing evidence linking gelatinase B (MMP-9) with malignant cell invasion. Tissue inhibitor 1 of MMP (TIMP-1) is a strong inhibitor of MMP-9 but has also tumor-enhancing effects. Only few data exist on MMP-9 or TIMP-1 expression in tissue samples of different breast histology. METHODS: MMP-9 and TIMP-1 immunoreactivity was examined in a wide range of breast tissue samples differing in histology from usual ductal hyperplasia (UDH) to fully developed ductal breast carcinoma. Immunohistochemical expression of MMP-9 was studied in 178 samples: 31 UDH samples, 29 atypical ductal hyperplasia (ADH) samples, 28 ductal carcinoma in situ (DCIS) samples and 90 ductal invasive carcinoma samples (30 samples of malignancy grades I, II and III, respectively). TIMP-1 expression was also analyzed in 178 breast tissue samples: 41 UDH, 21 ADH and 34 DCIS lesions, and 82 invasive ductal breast carcinomas (25 in grade I, 30 in grade II and 27 in grade III). RESULTS: A significantly distinctive pattern of MMP-9 protein expression was shown in DCIS samples, where 85.7% of the cases showed moderate or strong positivity and negative staining was rare (p = 0.021). Negative or weakly positive MMP-9 staining was the most prominent finding in UDH (71%), ADH (69%) as well as in invasive carcinoma samples (64.4%). Various degrees of TIMP-1 expression were seen in 86.5% of all cases. DCIS and invasive carcinoma samples revealed similar immunostaining: at least some positivity was seen in 91.1% of the DCIS samples and 91.5% of infiltrative carcinomas. Thus, TIMP-1 negativity (22.2%) was significantly associated with hyperplastic lesions (p = 0.026). CONCLUSIONS: These results suggest that MMP-9 and TIMP-1 overexpression are early markers of breast carcinogenesis preceding tumor invasion. Apparently, DCIS carries the risk to evolve into a malignant phenotype according to these markers. The clinical importance of these findings is discussed.

Aminoterminal propeptide of the alpha1-homotrimer variant of human type I procollagen (hotPINP) in malignant pleural effusion.

The aminoterminal propeptide (hotPINP) of type I homotrimer, a putative malignancy-associated type I collagen variant, was purified for the first time and a method was established for its detection in pleural fluid. Samples of 58 patients, with malignant or benign disease, were studied with specific immunoassays for the two propeptides of type-I procollagen (PICP and PINP) and with HPLC-DEAE chromatography to separate the two PINP variants. HotPINP was present in 64% of both benign and malignant pleural effusion fluids, with the exception of malignant mesotheliomas, none of which showed the presence of hotPINP. Also the PICP to PINP ratios were lower than normal in both benign and malignant samples (altogether in 69% of samples), although this deviation was greater in malignancy. These two phenomena were independent of each other. As synthesis of the alpha1-homotrimer-variant of type-I collagen seems to be relatively common during the formation of pleural effusion, it may be generally related to a fibroproliferative reaction in the pleural wall.

Cross-linked telopeptides of type I and III collagens in malignant ovarian tumours in vivo.

Malignant tumours often induce a fibroproliferative response in the adjacent stroma, characterized by increased expression of type I and type III procollagens. In normal tissues, fibrillar collagens normally undergo extensive intermolecular cross-linking that provides tensile strength to the tissue. Here we set out to characterize collagen cross-linking in human ovarian carcinoma tissue in vivo. Biochemical and immunochemical methods were used for cross-linked telopeptides of type I and III collagens in samples of benign and malignant serous tumours. The locations and staining patterns of these proteins were visualized immunohistochemically. The contents of both total collagen and the cross-linked type I and type III collagens in the malignant samples were only about 20% of those in the benign tumours. The cross-linked telopeptide antigens derived from the collagens were smaller and more heterogeneous in size in the malignant than in the benign tumours, indicating a defective cross-linking process scarcely leading to the formation of mature cross-links in the collagen fibres in malignancy. Immunostaining revealed disorganized type I and type III collagen bundles in carcinomas. These findings suggest that the collagen cross-linking process is aberrant in malignant tumours, possibly resulting in increased susceptibility of tumour collagens for the proteolysis often associated with tumour invasion.

Characterization of type I collagen synthesis and maturation in uterine carcinosarcomas.

BACKGROUND: Epithelial malignancies often induce an enhanced expression of interstitial collagens in the fibroblasts within the tumor tissue and the surrounding non-neoplastic stroma. In uterine carcinosarcomas (malignant mixed müllerian tumors [MMMTs]) both the stroma and the epithelium are malignant. METHODS: In this investigation, both in situ hybridization and immunohistochemical staining were applied with two different antibodies that were capable of distinguishing between newly synthesized and mature, trivalently cross-linked Type I collagen to define Type I procollagen mRNA expression and the synthesis and maturation of the corresponding protein in MMMTs. RESULTS: In the better differentiated parts of these tumors, in which anticytokeratins stained only clearly carcinomatous cells, Type I procollagen mRNA expression was limited to stromal fibroblasts; mature Type I collagen bundles were abundant and regular. In poorly differentiated areas, in which anticytokeratins stained only a few individual cells, Type I procollagen mRNA was expressed peculiarly by three morphologically different cell types. In addition to benign mesenchymal cells, Type I procollagen mRNA was present in atypical epithelial and mesenchymal cells. In these tumors, the collagen bundles close to the malignant cells were comprised of newly synthesized Type I collagen, with only little evidence of the presence of mature, fully cross-linked collagen. CONCLUSIONS: These results strongly suggest that the undifferentiated cells of MMMTs are capable of producing their own stroma with irregularly arranged collagen bundles.

The extracellular matrix in skin tumor development-a morphological study.

The development of cancer involves epithelial-stromal interactions. Alterations in the synthesis and deposition of type I and III collagens are related to the tumor morphology. Skin carcinogenesis in experimental animals provides a reliable model for the development of neoplasia. Ultraviolet (UV) irradiation is the main etiological factor for epidermal dysplasia and malignant tumors in man, but also for dermal degeneration. Non-neoplastic dermal changes and skin tumors induced by ultraviolet irradiation and 7,12-dimethylbenz(a)anthracene were investigated in various mouse strains with different susceptibilities to tumor formation. UVB irradiation resulted in an increased immunoreactivity of collagens in the dermis, in comparison with 7,12-dimethylbenz(a)anthracene. Increased synthesis and deposition of type I and III collagens were found in the stroma adjacent to benign alterations. In well-differentiated squamous cell carcinomas, a similar induction of collagen synthesis and deposition was observed. The destruction of fibrillary structures was more pronounced during the decrease of differentiation from moderately to poorly differentiated squamous cell carcinomas. Anaplastic carcinomas with spindle cell morphology displayed a delicate meshwork of reticular fibers and collagen III, and abnormal expression of mRNA for collagens in some malignant cells with epithelial characteristics. The underlying stroma reacts to the development of epithelial tumors in a reproducible way, which is related to the carcinogenic agent involved.

Type I and type III procollagen propeptides in amniotic fluid of normal pregnancies and in a case of mild osteogenesis imperfecta.

BACKGROUND: The propeptides derived from type I and III procollagens, PICP, PINP and PIIINP, indicate the synthesis of the corresponding collagens. Their circulating concentrations reflect the growth velocity in infants and children METHODS: We measured these propeptides in 145 samples of amniotic fluid of normal pregnancy. In addition, we have analysed an amniotic fluid and serum sample from a mother with osteogenesis imperfecta, and later the infant's serum sample was also collected for procollagen propeptide analysis. RESULTS: High concentrations of propeptides, 100-1000 times higher than those in adult serum, were found in early second trimester, decreasing significantly towards term, reflecting the decreased foetal growth rate. Interestingly, the amino-terminal propeptide of type I procollagen, PINP, decreased more that the corresponding carboxy-terminal propeptide, PICP, although both are in principle derived from the same protein. At both stages of pregnancy, the discrepant ratio of PICP to PINP indicated a molar excess of PINP. Abnormally low concentrations of PICP and PINP with normal PIIINP concentrations measured in amniotic fluid and in the serum indicated decreased synthesis of type I procollagen in a foetus/infant with mild osteogenesis imperfecta. CONCLUSIONS: Our data show a decrease in collagen synthesis with the stage of pregnancy and lower values of type I procollagen propeptides in a case of OI.

New lethal disease involving type I and III collagen defect resembling geroderma osteodysplastica, De Barsy syndrome, and Ehlers-Danlos syndrome IV.

We describe the clinical findings and biochemical features of a male child suffering from a so far undescribed lethal connective tissue disorder characterised by extreme hypermobility of the joints, lax skin, cataracts, severe growth retardation, and insufficient production of type I and type III procollagens. His features are compared with Ehlers-Danlos type IV, De Barsy syndrome, and geroderma osteodysplastica, as these disorders show some symptoms and signs shared with our patient. The child died because of failure of the connective tissue structures joining the skull and the spine, leading to progressive spinal stenosis. The aortic valve was translucent and insufficient. The clinical symptoms and signs, together with histological findings, suggested a collagen defect. Studies on both skin fibroblast cultures and the patient's serum showed reduced synthesis of collagen types I and III at the protein and RNA levels. The sizes of the mRNAs and newly synthesised proteins were normal, excluding gross structural abnormalities. These findings are not in accordance with any other collagen defect characterised so far.

Aberrant type I and type III collagen gene expression in human breast cancer in vivo.

Increased synthesis and degradation of extracellular matrix components are associated with breast cancer development. This study evaluated type I and type III procollagen mRNA expression and the corresponding protein synthesis and maturation, as well as the tissue distribution of these collagens, in benign breast lesions, infiltrating ductal carcinomas, and their metastases by in situ hybridization and immunohistochemistry. In the benign lesions, the type I and type III collagen bundles were regularly organized and the expression of the corresponding mRNA was weak, indicating a relatively slow collagen turnover. In the malignant tumours, increased expression of type I and type III procollagen mRNAs was observed in the fibroblastic cells of the stroma; the malignant epithelial cells did not participate. The staining of corresponding newly-synthesized pN-collagens showed aberrant bundles in the invasive front of the malignant tumours. Newly-synthesized type I and type III procollagens were occasionally observed in fibroblastic cells, particularly in grade 2 and grade 3 tumours. Metastases of breast carcinoma resembled poorly differentiated primary tumours with respect to their collagen synthesis and deposition. The increased synthesis of fibrillar type I and type III procollagens may serve as a pathway for tumour invasion. The enhanced synthesis is associated with the formation of aberrant collagen bundles, which may be more readily degradable and may thus facilitate breast tumour invasion.

Immunoassay for intact amino-terminal propeptide of human type I procollagen.

We have developed quantitative immunoassays for the intact, trimeric amino-terminal propeptide of human type I procollagen (PINP) and its Col1 domain. Intact PINP was isolated from the pleural fluids of cancer patients by a combination of ion-exchange, gel-filtration, and reversed-phase chromatographies. The amino-terminal Col1 domain of PINP was isolated after bacterial collagenase treatment of the heat-denatured trimeric propeptide. For the intact PINP assay we used a polyclonal antibody with only 1.2% cross-reaction with the monomeric Col1 domain. In human serum, this assay detects only one peak of PINP antigenicity that has the size of known intact PINP. Under similar conditions, an assay for the Coll domain of PINP recognized two circulating antigens. The biological relevance was further verified in wound fluid. Interassay and intraassay CVs were 3.1-9.3% for values within the reference intervals (mean +/- 2SD) for intact PINP in serum, which were 19-84 microg/L for women and 20-76 microg/L for men.

Expression of mRNAs for type I and type III procollagens in serous ovarian cystadenomas and cystadenocarcinomas.

Malignant ovarian tumors induce a strong fibro-proliferative reaction characterized by the active production of type I and type III procollagen both locally in the ovary as well as more remotely in the peritoneal cavity. Our purpose was to determine the origin of the increased collagen production observed in serous ovarian tumors with different histological grades of malignancy, ie, whether the malignant cells or the stromal fibroblasts are responsible for the synthesis of collagen fibers. We visualized the mRNAs corresponding to the pro alpha 1(I) and pro alpha 2(I) chains of type I procollagen and the pro alpha 1(III) chain of type III procollagen by in situ hybridization. Strong signals for both chains of type I procollagen were seen in stromal fibroblasts next to tumor cell islets, whereas the reaction was weak or absent near benign ovarian cysts. In poorly differentiated tumors, the signals were particularly abundant and occasionally also seen in the neoplastic cells themselves. Type III procollagen mRNA expression was similar, although somewhat less distinct. These findings indicate that the production of interstitial procollagens is related to the degree of malignancy and neoplastic activity of tumors. The formation of collagen in well differentiated ovarian tumors is a function of stromal fibroblasts, whereas in poorly differentiated tumors, aberrant expression of one or several chains of type I and type III procollagens in the neoplastic cells is also likely to take place.

Cardiovascular stress in isokinetic trunk strength test.

STUDY DESIGN: In this experimental study, the participants served as their own controls. OBJECTIVE: To estimate cardiovascular stress in isokinetic trunk extension-flexion strength test. SUMMARY OF BACKGROUND DATA: Thirty-one women with low back or neck pain who participated in the physical reconditioning program were the study participants. METHODS: Trunk extension and flexion strength was measured at three angular velocities with heart rate recording and standard graded submaximal bicycle ergometer test. Power output at the maximal age-predicted heart rate level was estimated separately by trunk extension-flexion and bicycle ergometer tests. RESULTS: Torque decreased and heart rate and power increased with angular velocity (P < 0.0001). Power and heart rate values at the highest angular velocity of 120 degrees s-1 in isokinetic trunk strength testing were comparable to those measured during bicycle ergometer test at the submaximal level. In trunk strength testing 13 subjects exceeded the power and 23 subjects exceeded the heart rate they achieved in the bicycle ergometer test. Power output at the maximal age-predicted heart rate level estimated by isokinetic trunk strength test and bicycle ergometer test had a clear correlation 0.76 (confidence interval 95% 0.55-0.88). CONCLUSIONS: The cardiovascular capacity is an important factor-limiting performance in isokinetic trunk strength testing, particularly at high velocities. Patients with suspected heart problems need special attention. Heart rate recording may help estimate degree of effort.