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Background: This study aimed to determine human papillomavirus (HPV) status and to investigate p16(INK4A) and Ki-67 expression and their correlation with clinical parameters and survival in women with primary carcinoma of the vagina (PCV).Methods: The presence of HPV DNA was evaluated by PCR. Genotyping was performed by Luminex in 68 short-term (2 years) and long-term (8 years) PCV survivors. p16(INK4A) and Ki-67 expression was evaluated by immunohistochemistry.Results: Human papillomavirus DNA was detected in 43% of patients, the majority (63%) of whom were HPV16 positive. High p16(INK4A) expression was significantly correlated with low histopathological grade (P=0.004), HPV positivity (P=0.032), and long-term survival (P=0.045). High Ki-67 expression was negatively correlated with histopathological grade (P<0.001) and tumour size (P=0.047). There was an association between HPV positivity and low histopathological grade, but not between HPV positivity and survival.Conclusion: High p16(INK4A) expression was associated with long-term survival, but the only independent predictors for survival were tumour size and histopathological grade. Our results indicate that p16(INK4A) and Ki-67 expression might be useful in tumour grading, and that it might be possible to use p16(INK4A) expression as a marker for HPV positivity, but this has to be further elucidated. [ABSTRACT FROM AUTHOR]

Background:This study aimed to determine human papillomavirus (HPV) status and to investigate p16INK4A and Ki-67 expression and their correlation with clinical parameters and survival in women with primary carcinoma of the vagina (PCV).Methods:The presence of HPV DNA was evaluated by PCR. Genotyping was performed by Luminex in 68 short-term (2 years) and long-term (8 years) PCV survivors. p16INK4A and Ki-67 expression was evaluated by immunohistochemistry.Results:Human papillomavirus DNA was detected in 43% of patients, the majority (63%) of whom were HPV16 positive. High p16INK4A expression was significantly correlated with low histopathological grade (P=0.004), HPV positivity (P=0.032), and long-term survival (P=0.045). High Ki-67 expression was negatively correlated with histopathological grade (P<0.001) and tumour size (P=0.047). There was an association between HPV positivity and low histopathological grade, but not between HPV positivity and survival.Conclusion:High p16INK4A expression was associated with long-term survival, but the only independent predictors for survival were tumour size and histopathological grade. Our results indicate that p16INK4A and Ki-67 expression might be useful in tumour grading, and that it might be possible to use p16INK4A expression as a marker for HPV positivity, but this has to be further elucidated. [ABSTRACT FROM AUTHOR]

Background: Genomic stability is one of the crucial prognostic factors for patients with endometrioid endometrial cancer (EEC). The impact of genomic stability on the tumour tissue proteome of EEC is not yet well established.Methods: Tissue lysates of EEC, squamous cervical cancer (SCC), normal endometrium and squamous cervical epithelium were subjected to two-dimensional (2D) gel electrophoresis and identification of proteins by MALDI TOF MS. Expression of selected proteins was analysed in independent samples by immunohistochemistry.Results: Diploid and aneuploid genomically unstable EEC displayed similar patterns of protein expression. This was in contrast to diploid stable EEC, which displayed a protein expression profile similar to normal endometrium. Approximately 10% of the differentially expressed proteins in EEC were specific for this type of cancer with differential expression of other proteins observed in other types of malignancy (e.g., SCC). Selected proteins differentially expressed in 2D gels of EEC were further analysed in an EEC precursor lesion, that is, atypical hyperplasia of endometrium, and showed increased expression of CLIC1, EIF4A1 and PRDX6 and decreased expression of ENO1, ANXA4, EMD and Ku70.Conclusion: Protein expression in diploid and aneuploid genomically unstable EEC is different from the expression profile of proteins in diploid genomically stable EEC. We showed that changes in expression of proteins typical for EEC could already be detected in precursor lesions, that is, atypical hyperplasia of endometrium, highlighting their clinical potential for improving early diagnostics of EEC. [ABSTRACT FROM AUTHOR]

Background: Cytology-based diagnostics of squamous cervical cancer (SCC) precursor lesions is subjective and can be improved by objective markers.Methods: IHC-based analysis of ANXA6, HSP27, peroxiredoxin 2 (PRDX2), NCF2, and tropomyosin 4 (TPM4) during SCC carcinogenesis.Results: Expression of ANXA6, HSP27, PRDX2, and NCF2 in the cytoplasm of dysplastic cells increased from cervical intraepithelial neoplasia 2/3 (CIN2/3) to microinvasive cancer. Invasive SCC showed lower expression of TPM4 than CIN and normal epithelium. CIN2/3 with the highest sensitivity and specificity differed from normal epithelium by cytoplasmic expression of HSP27. Patients with cytoplasmic HSP27 expression in SCC deviating from that observed in normal epithelium had worse relapse-free (P=0.019) and overall (P=0.014) survival. Invasive SCC with the highest sensitivity and specificity differed from normal epithelium by expression of PRDX2 and TPM4 in the cytoplasm, from CIN2/3 by the expression of ANXA6 and TPM4 in the cytoplasm, and from microinvasive SCC by the expression of PRDX2 and ANXA6 in the cytoplasm. The number of sporadic ANXA6+ cells between the atypical cells increased from CIN2/3 to invasive SCC.Conclusion: Detection of expression changes of the proteins ANXA6, HSP27, PRDX2, NCF2, and TPM4 in SCC precursor lesions may aid current cytological and pathological diagnostics and evaluation of prognosis. [ABSTRACT FROM AUTHOR]

The objective was to identify proteins differentially expressed in vaginal cancer to elucidate relevant cancer-related proteins. A total of 16 fresh-frozen tissue biopsies, consisting of 5 biopsies from normal vaginal epithelium, 6 from primary vaginal carcinomas and 5 from primary cervical carcinomas, were analysed using two-dimensional gel electrophoresis (2-DE) and MALDI-TOF mass spectrometry. Of the 43 proteins identified with significant alterations in protein expression between non-tumourous and tumourous tissue, 26 were upregulated and 17 were downregulated. Some were similarly altered in vaginal and cervical carcinoma, including cytoskeletal proteins, tumour suppressor proteins, oncoproteins implicated in apoptosis and proteins in the ubiquitin-proteasome pathway. Three proteins were uniquely altered in vaginal carcinoma (DDX48, erbB3-binding protein and biliverdin reductase) and five in cervical carcinoma (peroxiredoxin 2, annexin A2, sarcomeric tropomyosin kappa, human ribonuclease inhibitor and prolyl-4-hydrolase beta). The identified proteins imply involvement of multiple different cellular pathways in the carcinogenesis of vaginal carcinoma. Similar protein alterations were found between vaginal and cervical carcinoma suggesting common tumourigenesis. However, the expression level of some of these proteins markedly differs among the three tissue specimens indicating that they might be useful molecular markers. [ABSTRACT FROM AUTHOR]

Protein patterns in six samples from primary vaginal cancers, in five from normal vaginal tissue and in five primary cervical cancers, were analysed using two-dimensional polyacrylamide gel electrophoresis (2-DE). Protein expression profile was evaluated by computer-assisted image analysis (PDQUEST) and proteins were subsequently identified using matrix-assisted laser desorption/ionisation mass spectrometry. The aim was to analyse the protein expression profiles using the hierarchical clustering method in vaginal carcinoma and to compare them with the protein pattern in cervical carcinoma in order to find a helpful tool for correct classification and for increased biomedical knowledge. Protein expression data of a distinct set of 33 protein spots were differentially expressed. These differences were statistically significant (Mann-Whitney signed-Ranked Test, P<0.05) between normal tissue, vaginal and cervical cancer. Furthermore, protein profiles of pairs of primary vaginal and cervical cancers were found to be very similar. Some of the protein spots that have so far been identified include Tropomyosin 1, cytokeratin 5, 15 and 17, Apolipoprotein A1, Annexin V, Glutathione-S-transferase. Others are the stress-related proteins, calreticulin, HSP 27 and HSP 70. We conclude that cluster analysis of proteomics data allows accurate discrimination between normal vaginal mucosa, primary vaginal and primary cervical cancer. However, vaginal and cervical carcinomas also appear to be relatively homogeneous in their gene expression, indicating similar carcinogenic pathways. There might, further, be a possibility to identify tumour-specific markers among the proteins that are differentially expressed. The results from this study have to be confirmed by more comprehensive studies in the future. [ABSTRACT FROM AUTHOR]

Acetaldehyde (AcH) at preincubationconcentrations of 447, 89.4, and 17.9 mM potentiates theeffects of heparin on the clotting time of plasma. Whilecontrol plasma clotted in the range of 12.6 ± 0.1 to 13.8 ± 0.1 sec, and heparin-treatedplasma clotted in a range from 131.5 ± 2.5 to168.2 ± 1.2 sec, heparin that was preincubated atroom temperature for 30 min with 89.4 or 447 mM AcH didnot clot plasma in 300 sec. Heparin exposed to 17.9 mMAcH clotted plasma in 193 ± 1.1 sec. Ethanol ata 404 mM concentration also prolonged the clotting timeof heparin-treated plasma >300 sec, while 202 mM ethanol prolonged the clotting time ofheparin-treated plasma from 149.0 ± 2.0 sec to219.5 ± 1.7 sec. It is suggested that AcH altersthe tertiary structure of heparin by adduct formation,possibly by formation of cyclic acetals with iduronicand glucuronic acids, thereby more readily affectingbinding of the glycosaminoglycan to antithrombin IIIand/or thrombin, prolonging clotting time. Ethanol, which does not react covalently with heparin,might affect its conformation as a consequence of anorganic solvent effect. Protamine sulfate prolonged theclotting time of plasma from 13.6 ± 0.1 sec to 17.9 ± 0.2 sec. Protaminesulfate-treated heparin clotted plasma in 21.0 ±0.4 sec relative to heparin-treated plasma (160.4± 1.7 sec). In subsequent experiments,AcH-treated protamine sulfate extended the clotting time of protamine sulfate from17.9 ± 0 sec to 33.7 ± 0.6 sec. Prioraddition of protamine sulfate to AcH- heparin mixturesor heparin to protamine sulfate-AcH mixtures beforeaddition to plasma resulted in clotting times of 22.0± 0.4 sec and 24.1 ± 0.5 sec,respectively, relative to control clotting times of162.3 ± 2.6 sec for plasma-heparin mixtures.These results confirm both the reduction in coagulation time ofheparin-treated plasma by protamine sulfate and theprolongation of clotting time of plasma by protaminesulfate. Furthermore, and importantly, they indicatethat acetaldehyde-treated protamine sulfate is a more effectiveanticoagulant than protamine sulfate. It is suggestedthat reversible adduct formation between acetaldehyde,heparin, and protamine sulfate may occur as a meansexplaining the essentially identical coagulation time ofthese mixtures when added to plasma regardless of theorder of premixing. Ethanol (404 mM) did not influenceprotamine sulfate effects. Lastly, the potentiation of the anticoagulant function of heparin byacetaldehyde suggests that a structural modification ofthe glycosaminoglycan may occur in alcoholics. [ABSTRACT FROM AUTHOR]

The anticoagulant activity of antithrombin III (ATIII), as observed in a plasma-free system consisting of thrombin and fibrinogen, is readily reduced by acetaldehyde (AcH) at concentrations of 447, 89.4, and 17.9 mM. Whereas control thrombin-fibrinogen mixtures clotted in 17.7+/-0.75 sec, ATIII prolonged clotting time to 55.0+/-1.75 sec on preincubation with thrombin for 30 min at room temperature. On subsequent preincubation of ATIII with the AcH for 30 min at room temperature and passage of the mixture through Sephadex G-25 minicolumns to remove excess AcH, the eluates were tested for anticoagulant activity. Clotting times of 20.9+/-1.0, 32.3+/-1.0, and 45.3+/-1.6 sec were obtained with 447, 89.4, and 17.9 mM AcH-ATIII mixtures, respectively. These data suggest that functional groups on ATIII, such as guanidiniums, aminos, and others are susceptible to adduct formation with AcH, thereby altering the shape and charge of the anticoagulant. As a consequence of this type of reaction, an altered molecule of reduced biological activity may be produced. These experimental results may explain, in part, the reduction in ATIII levels reported by others in patients with alcoholic liver disease. [ABSTRACT FROM AUTHOR]

Evidence is presented to indicate that there exists in lymphoid tissue, as a result of transforming lymphocytes, a new lymphokine which is chemotactically specific for lymphocytes, called 'lymphotactin'. Lymphotactin has been purified to electrophoretic homogeneity; has a molecular weight of 10,500 D and an isoelectric point of 5.9. Its role in amplifying the immune defense system by recruitment of naive lymphocytes into propinquity with the challenging antigen is suggested. Purification of macrophage migration inhibitory factor from thymus extracts to electrophoretic homogeneity leads to a compound of molecular weight of 36,500 D and an IEP of 6.9. Chemically it contains sialic acid and o-methyl glucopyranoside as its only carbohydrates. Purified MIF activates the macrophage phagocytically. Skin reactive factor and lymph node permeability factor have been isolated and purified and are found to be inhibited by pepstatin and antihistamine and to have an isoelectric point of pH 4.2 and a molecular weight of 50,000-100,000 D. It is believed that this anionic permeability increasing agent actually arises from the lysosomes of macrophages and lymphoblasts (the normal small lymphocyte having essentially no lysosomal organelles). The mononuclear cell infiltration characteristic of crude SRF and LNPF may proceed from their being contaminated with lymphotactin. [ABSTRACT FROM AUTHOR]

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