Localization of hyaluronan with a hyaluronan-specific hyaluronic acid binding protein in the placenta in pre-eclampsia.

Hyaluronan (HA), a high molecular weight polysaccharide, is a major component of connective tissue and is thus present in the extracellular matrix of most tissues. Increased serum concentrations have been reported in association with pre-eclampsia and liver malfunction, amongst other disorders. We have performed histochemical investigations with a HA-specific hyaluronic acid binding protein in placentas from uncomplicated pregnancies and from patients with pre-eclampsia. Staining for HA was found in the stroma and blood vessel walls of stem villi in all the placentas investigated. The syncytiotrophoblast and cytotrophoblast cells usually remained unstained. In addition, reactivity for HA was found within and on the surface of intervillous and perivillous fibrinoid deposits. Since fibrinoid deposits are increased in pre-eclampsia, our findings suggest that the increased HA serum concentrations in cases of pre-eclampsia could result from the stroma of the infarcted villi and from the fibrinoid deposits. HA may reach the maternal blood through fibrinoid gaps.

[Expression of galectin-3 in thyroid gland and follicular cell tumors of the thyroid. A critical study of its possible role in preoperative differential diagnosis]. / Untersuchungen zur Expression von Galectin-3 im Schilddrüsengewebe und in Follikelzelltumoren der Schilddrüse. Kritische Bewertung einer möglichen präoperativen differenzialdiagnostischen Nutzung.

Galectin-3 belongs to a group of endogenous lectins with an affinity to glycoconjugates containing beta-galactoside residues. It has been detected in numerous tissues and studied in connection with tumor growth, dedifferentiation, and metastasis. Only few studies have dealt with galectin-3 detection in tumors of the thyroid gland and with its possible role for differential diagnosis. We studied 118 cases of thyroid gland tumors with a monoclonal antibody against galectin-3; we compared the preparations by a semiquantitative score to determine differences in expression. Normal thyroid gland tissue, goiter tissue, and tissue with functional enhancement were largely negative for galectin-3. Adenomas with a typical cytological pattern were predominantly negative, but a focal positive reaction in single cells and cell groups or follicles was possible. Almost all papillary carcinomas showed a distinct galectin-3 expression. While findings in follicular carcinomas and oxyphilic adenomas and carcinomas were very uneven, with both positive and negative tumors, the galectin-3 reaction can be helpful in recognizing follicular variants of the papillary carcinoma. Investigation of the biological significance of tumors should always be cautious and consider known histological criteria for malignancy.

Increase of segments of elastic-type blood vessel walls in fetal placental stem villi during pre-eclampsia at term.

In recent studies we described the presence of elastic-type blood vessels within trunci and rami chorii of human placental stem villi. For systemic and pulmonary hypertension it is known that elastic fibres are enhanced in arteries. The aim of our study was, therefore, to examine whether pre-eclampsia may lead to an increase of elastic tissue fibres in blood vessel walls of placental stem villi and whether there are differences in the thickness of blood vessel walls within these villi when compared to normotensive pregnant women. Twenty-six women with uncomplicated pregnancies and 25 patients with pre-eclampsia were investigated. Unfixed cryostat serial sections were processed for conventional orcein staining and for the demonstration of alpha-actin-immunoreactivity. The intensity of orcein staining of stem villus blood vessel walls was evaluated by a semiquantitative score method. Significant higher intensities of orcein staining (P<0.00001) were calculated for blood vessel walls of placentae with pre-eclampsia. The amount of thick stem villus vessels (>41 microm) increased during pre-eclampsia from 39 gestational weeks onwards. Our study demonstrates that segments of thick blood vessel walls and elastic-type vessel walls are increased in placental stem villi of patients with pre-eclampsia. This reaction may protect the fetal placental vessels and avert an increase of the fetal hypertension.

Histochemical evaluation of placental dipeptidyl peptidase IV (CD26) in pre-eclampsia: enzyme activity in villous trophoblast indicates an enhanced likelihood of gestational hypertensive disorders.

PROBLEM: To determine whether differences in placental dipeptidyl peptidase IV (DPP IV, CD26) activities occurred in hypertensive complicated pregnancies as compared with uncomplicated pregnancies. METHOD: DPP IV activity was detected with H-Gly-Pro-4M2NA as the substrate in placental cryostat sections from 65 patients with gestational hypertension and 67 patients with uncomplicated pregnancies. The graduated intensities of the reaction product in the villous trophoblast were scored semiquantitatively by light microscopy and were related to the relative frequencies of hypertensive disorders (proportional odds model). After detection of enzyme activity, the same tissue samples were homogenized and used for kinetic fluorometric measurements. RESULTS: Enhanced villous trophoblastic DPP IV activity was significantly associated with an increased frequency of proteinuric hypertension in pregnant women (cumulative odds ratio theta1 = 1.6; P < 0.01). CONCLUSION: This study demonstrates for the first time that increased villous trophoblastic DPP IV activity indicates an increased likelihood of the presence and of the severity of hypertensive disorders in pregnancy.

Binding of antibodies against high and low molecular weight cytokeratin proteins in the human placenta with special reference to infarcts, proliferation and differentiation processes.

Recent immunocytochemical studies have shown that placental villous trophoblasts contain the high molecular weight cytokeratin (CK) proteins 5/6 and 17. In the case of CK 17, trophoblastic immunostaining was positive in villi covered by fibrinoid. CKs 5/6 and 17 are expressed by hyperproliferative cells. The aim of this investigation was to examine the location of these CKs in placental infarcts, known to be demarcated by fibrinoid and hyperproliferative trophoblasts. The results were compared with those obtained by immunostaining against Ki-67, tenascin and alpha 1-, alpha 6- and beta 1-integrins, which are involved in cell proliferation, differentiation and regenerative processes. Furthermore, the expression of the single CKs 7, 8, 10, 13, 14, 18 and 19 was investigated by immunocytochemistry and immunoblotting. While low and high molecular weight CKs were present in villous and extravillous trophoblasts, only low molecular weight CKs were detected in vascular and extravascular placental smooth muscle cells. Placental infarcts revealed different immunoreactivities in the infarct margin and centre: high molecular CKs, tenascin, Ki-67 and oncofoetal fibronectin predominated in the infarct margin, low molecular CKs, fibrin and integrins in the centre. The expression of tenascin and a defined change in the expression of CK 17 indicates villous repair and hyperproliferative mechanisms in placental infarcts.

Molecular anatomy of the perivascular sheath in human placental stem villi: the contractile apparatus and its association to the extracellular matrix.

In previous studies, we have shown that smooth muscle cells and myofibroblast subpopulations of the perivascular stem villous sheath of the human placenta contain focal adhesion plaques and talin immunoreactivity. The close association of these cells to elastic and collagen fibres have led to the assumption of a functional myofibroelastic unit within the perivascular stem villous sheath. Interactions between the extracellular matrix and smooth muscle cells depend on a variety of structural protein assemblies. In the present study, we examined, by immunocytochemistry, whether the molecular assembly of extracellular matrix proteins and molecules of focal adhesions, known to be essential for signal transduction in smooth muscle cells, are also found in smooth muscle cells of the perivascular stem villous sheath of the human placenta. Vascular and extravascular smooth muscle cells were immunoreactive for alpha-actinin, vinculin, paxillin and tensin, the integrin chains alpha1 and beta1, and the basement membrane components laminin and heparan/-chondroitin sulfate proteoglycan perlecan. pp125(FAK) did not react. In the extracellular matrix of blood vessel walls and the perivascular stem villous sheath, we found immunoreactivity of fibronectin and collagen types I, VI and undulin (collagen type XIV). From our data we conclude that within the perivascular stem villous sheath, there exists a system of signal transduction molecules, indicating a cross talk between the smooth muscle cells of this sheath and their surrounding extracellular matrix.

Villous cytotrophoblast proliferating potential in complete and partial hydatidiform mole: diagnostic value of silver-stained nucleolar organizer region (AgNOR)-associated proteins.

Aside from their typical morphologic features, complete (CHM) and partial hydatidiform mole (PHM) are characterized by variable trophoblastic proliferation and/or atypia. CHM and PHM usually present little diagnostic difficulty. However, some may be extremely difficult to distinguish by morphologic features alone. Therefore, we investigated the diagnostic value of silver-stained nucleolar organizer region (AgNOR)-associated proteins in cytotrophblasts as compared to cytogenetic features of nine CHM, nine PHM and six non-molar spontaneous embryonic abortions (controls), as well as of two suspected CHM and two histologically suspected PHM. Tissue sections were submitted to autoclave pretreatment and to silver colloid solution. The proliferating potential of cytotrophoblasts was determined by the analysis of mean number and mean area of AgNORs per nucleus using a PC-based image analysis system. Mean values of AgNOR parameters were significantly different from each other (p < 0.001). Each of the four cases of tentative diagnosis could be assigned to the corresponding group of examined trophoblastic lesions. The evaluation of AgNORs in cytotrophoblasts contributes to a reliable discrimination of CHM and PHM; this fairly simple and economical method could serve as an useful addition to conventional methods of diagnosis in gestational trophoblastic disease.

Histochemical evaluation of placental angiotensinase A in pre-eclampsia: enzyme activity in villous trophoblast indicates an enhanced likelihood of gestational proteinuric hypertension.

The aim of this study was to examine whether differences in placental angiotensinase A (glutamyl aminopeptidase, EC 3.4.11.7) activities occurred in hypertensive complications of pregnancy compared with uncomplicated pregnancies. Biochemical and semiquantitative histochemical methods were used and compared for their applicability. Angiotensinase A activity was detected using L-alpha glutamyl-4-methoxy-2-naphthylamide (alpha-Glu-MNA) as substrate and Fast Blue B salt for simultaneous azo-coupling in cryostat sections of placental tissue samples from 32 patients with pre-eclampsia, 11 patients with pregnancy-induced hypertension and 44 participants with uncomplicated pregnancies. The graduated intensity of reaction product in the villous trophoblast and in fetal blood vessels was evaluated semiquantitatively in a double-blind study by light microscopy (semiquantitative score method). Score levels were related to relative frequencies of hypertensive disorders (proportional odds model) and correlated to the severity of gestational hypertension (Spearman's rank correlation). After detection of enzyme activity, the same tissue samples were homogenized and used for kinetic fluorometric measurements under the same substrate and buffer conditions as in enzyme histochemistry. Enhanced villous trophoblastic angiotensinase A activity was significantly associated with an increased frequency of pre-eclampsia in pregnant women (cumulative odds ratio x 0(1) 6.37; P < 0.001) and showed significant correlations with the severity of gestational hypertensive disorders, represented by systolic (r = 0.31; P < 0.05) and diastolic (r = 0.34; P < 0.05 blood pressure and by concomitant proteinuria (r = 044; P < 0.01). Histochemical evaluation of fetal blood vessels and biochemical measurements revealed no statistically significant results. In conclusion this study demonstrates for the first time that increased villous trophoblastic angiotensinase A activity indicates an increased likelihood of the presence of pre-eclampsia and the severity of hypertensive disorders in pregnancy.

Elastic fibres are an essential component of human placental stem villous stroma and an integrated part of the perivascular contractile sheath.

The stroma of human placental stem villi is believed to consist only of reticular and collagen fibres. In the present study we were able to show for the first time by light (orcein staining) and electron microscopy large amounts of elastic fibres in the stem villous stroma. Electron microscopically, homogeneous elastin was found alone or in association with microfibrils. In addition, microfibrils were observed forming long bands. These three structures, generally known to form elastic connective tissue, were seen in close connection with placental extravascular smooth muscle cells, which belong to the perivascular contractile sheath (PVCS) of stem villi. Elastin was associated with these smooth muscle cells and connected to collagen fibres via microfibrils. Collagen fibres were additionally interconnected by spike-like structures. Extravascular smooth muscle cells revealed numerous adhesion plaques which occupied conspicuously long cytoplasmic faces of the plasma membrane. In cryostat sections, immunoreactivity of talin, an attachment protein of adhesion plaques linking intracellular alpha-actin filaments with extracellular fibronectin, was detected in extravascular and vascular (media) smooth muscle cells. The arrangement of placental extravascular smooth muscle cells, elastic and collagen fibres suggests a functional myofibroelastic unit within the PVCS, which surrounds the large foetal blood vessels possibly contributing to elasticity and supporting tensile and/or contracting forces within the stem villi.

Antibodies to cytokeratins bind to epitopes in human uterine smooth muscle cells in normal and pathological pregnancies.

Cytokeratin antibodies have been widely used for the identification of trophoblast cells in the placental bed, following their invasion from the developing conceptus. Their identification centres upon the expression of cytokeratin in epithelial cells, from which trophoblast cells are derived. Our recent observations indicate that this strict relationship may be more complex than was thought. Cryostat and paraffin sections of human decidua and myometrium, taken from the placental bed and the uterotomy cut, were examined immunocytochemically for cytokeratins using ten antibody clones selected to identify different cytokeratin proteins and antigenic epitopes. Biopsy specimens were obtained from normal and pathological pregnancies (pre-eclampsia, fetal retardation, amnioninfection, hysterorrhexis, placenta praevia) at the time of caesarean section (26-41 weeks of pregnancy). Antibodies against nine clones, CAM 5.2, MNF 116, AE1/AE3, CK5, KS-B17.2, CY-90, M20, E3, and 34 beta E12 identified, as expected, syncytial giant cells and mononuclear trophoblasts within the placental bed and glandular epithelial cells throughout the uterus. In addition, they stained numerous fusiform cells that were classified by established criteria to represent smooth muscle cells, both within blood vessels and myometrium. No staining differences were observed between normal and pathological disorders. These results indicate that cytokeratin antibodies CAM 5.2, MNF 116 and AE1/AE3, and other antibodies targeting proteins 8 and 18, cross-react with epitopes expressed in cells other than giant trophoblastic cells and mononuclear trophoblasts in the uterus and, thus, caution has to be used when such antibodies are used for the diagnostic characterization of tissues related to the placental bed.

Monoclonal antibody against Klebsiella capsular polysaccharide reduces severity and hematogenic spread of experimental Klebsiella pneumoniae pneumonia.

Klebsiella pneumoniae is an important nosocomial pathogen causing severe pulmonary infections. The majority of clinical Klebsiella isolates produce a high-molecular-weight capsular polysaccharide (CPS) which is one of the dominant virulence factors. In the present study, we examined the potency of a murine immunoglobulin M monoclonal antibody (MAb) with specificity to Klebsiella type 2 CPS to protect rats against experimental Klebsiella pneumonia. The MAb did not prevent the invasion of virulent bacteria into the interalveolar space. However, the resolution of infection was accelerated in MAb-treated animals. This was demonstrated by (i) less severe weight loss and (ii) markedly reduced inflammatory reactions in the lung. The elimination of bacteria was significantly increased not only in the lungs but also in the livers of antibody-treated rats. This was reflected by reduced levels of circulating, soluble CPS and MAb-bound CPS. A mixture of human MAbs with specificity to CPS of clinically important Klebsiella serotypes may prove to be a useful tool for the prevention or supportive treatment of Klebsiella pneumonia.