R. Pijnenborg

Immunolocalization of tumour necrosis factor-α (TNF-α) in the placental bed of normotensive and hypertensive human pregnancies

Abstract To identify tumour necrosis factor (TNF)-γ immunopositive cells, third trimester human placental bed biopsies were selected from nine normotensive control women, 16 severely pre-eclamptic patients and seven patients with pre-existing hypertension with superimposed pre-eclampsia. In addition, five first and early second trimester specimens were included in the study. Immunostaining was performed with a mouse IgGI monoclonal antibody (J1 D9) reactive specifically with human TNF-α (1 : 300 ascitic fluid), using a biotin-streptavidin-peroxidase technique. Variable staining of stromal cells was noted in all biopsies. Specimens of early pregnancy showed marked immunostaining for TNF-a on proliferating tips of anchoring villi, invasive interstitial cytotrophoblast (but not the multinuclear giant cells), and endovascular trophoblast invading the spiral arteries. At term, weak staining was found in trophoblast incorporated within spiral artery walls. In biopsies from pre-eclamptic patients, spiral arteries without physiological change showed very little staining except in atherotic vessels where the infiltrated lipophages often showed intense immunolabelling. The marked presence of TNF-a in extravillous cytotrophoblast of young specimens is suggestive of a role in early invasion. Immunostaining of foam cells in non-invaded spiral arteries in pre-eclampsia at or near-term indicates a potential role of this cytokine in the development of atherotic lesions.

Expression of stromelysin-3 in the human placenta and placental bed

Human placentation is mediated by fetal trophoblastic cells which penetrate into the decidualized uterine endometrium. Trophoblast invasion requires the precisely regulated secretion of specific proteinases able to degrade the endometrial basement membranes and extracellular matrix. To document further the involvement of these proteinases during human placentation, we evaluated in vivo the expression of stromelysin-3, a member of the metalloproteinase family, during the first and third trimesters of pregnancy, by means of immunohistochemistry, in situ hybridization and Northern blot analysis. Human extravillous trophoblasts invading the maternal decidua produced stromelysin-3 during both, the first and third trimesters of pregnancy, but to a lesser extent during the latter. In floating villi, stromelysin-3 expression was restricted to the syncytiotrophoblasts that line intervillous vascular spaces. In conclusion, stromelysin-3 is expressed by differentiated, non-proliferative villous and extravillous trophoblastic cells in early and late placental beds and villi, and its pattern of expression evolves during pregnancy. Our observations suggest that stromelysin-3 could play a role in human placentation.

Pathogenesis of Fetal Hypomineralization in Diabetic Rats: Evidence for Delayed Bone Maturation

There is some evidence that fetuses of diabetic rats (FDR) are hypomineralized. To explore the pathogenic role of decreased maternal duodenal Ca absorption, fetal hypotrophy, and decreased placental calbindin-D9K, respectively, spontaneously diabetic rats fed a 1.0% Ca diet were compared with diabetic rats treated with 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] (15 ng/100 g) during week 3 of pregnancy, which restored duodenal calbindin-D9K concentrations to normal; with nondiabetic rats semistarved during week 3, which resulted in similar fetal hypertrophy; and with nondiabetic rats fed high cation diets (1.5% Ca-1.5% Sr and 1.5% Ca-3.5% Sr) during week 3, the latter of which repressed duodenal and placental calbindin-D9K toward concentrations measured in diabetic rats. In addition, fetal tibiae were studied histologically. Ca content was lower in 21.5-d-old FDR than in control fetuses. FDR had lower plasma osteocalcin (OC) levels and, on histomorphometry, increased hypertrophic cartilage width, indicating retarded bone maturation. Maternal 1,25(OH)2D3 treatment did not change Ca content and hypertrophic cartilage width in FDR. Fetuses of semistarved rats had plasma OC levels and hypertrophic cartilage width comparable to those of control fetuses. Fetuses of rats fed the 1.5% Ca-3.5% Sr diet were more severely hypomineralized than FDR but had higher plasma OC than both FDR and control fetuses, compatible with fetal Ca deficiency. Whereas diabetic placentas showed weak but homogeneous staining of calbindin-D9K in the labyrinth on immunohistology, degenerative zones were present in placentas of rats fed the 1.5% Ca-3.5% Sr diet. Thus, there is no mineralization defect in FDR caused by disturbed maternal duodenal Ca absorption or transplacental Ca transport, but a delay in bone maturation that is unexplained by their lower body weight.

Influence of a vitamin E-deficient diet on prostacyclin production by mesometrial triangles and aortic rings from nondiabetic and diabetic pregnant rats

We investigated the effect of a vitamin E-deficient diet on the synthesis of prostacyclin in nondiabetic and diabetic pregnant rats. In both groups the diet reduced fetal growth and prostacyclin production by mesometrial triangles; it also increased serum lipid peroxides, which are known to inhibit prostacyclin synthesis. Independently of the diet, mesometrial triangles of diabetic rats produced more prostacyclin than those of control rats, whereas an opposite tendency was found in the aortic rings. Our model seems useful for fundamental and pharmacologic studies of the vascular pathology of pregnancy.

Growth characteristics of diabetic rat ectoplacental cones in vivo and in vitro

Aims/hypothesis. To investigate the outgrowth of the ectoplacental cone in diabetic rats in vivo and in vitro.¶Methods. Female Wistar rats were injected intraperitoneally with streptozotocin (75 mg/kg body weight, n = 15), or with control buffer (n = 27) 3 days before mating. On day 9 (day 1 = copulation plug) decidual swellings were weighed and the volume and mitotic index of the embryo and ectoplacental cone were estimated. Also, ectoplacental cones were cultured either in the presence of decidual cells from pseudopregnant diabetic rats or in high glucose concentration media. Cultures were evaluated by the daily outgrowth and by the proportion of giant cells and proliferating cells on day 5.¶Results. In diabetic rats on day 9, the weight of the decidual swellings and the mitotic index in the ectoplacental cone were lower compared with controls (p < 0.0001 and p < 0.05, respectively). In vitro, control ectoplacental cones in the presence of decidual cells from diabetic rats showed a slight reduction in outgrowth on day 3 and 5 of culture. Outgrowth of diabetic ectoplacental cones in high glucose concentration medium was impaired on day 1 (p < 0.0005) compared with control ectoplacental cones in control medium, and on day 1 and 2 (both p < 0.005) compared with control ectoplacental cones in high glucose concentration medium. In control medium, the outgrowth of diabetic ectoplacental cones was impaired on day 1 (p < 0.05), compared with control ectoplacental cones. Proliferation was stimulated in diabetic ectoplacental cone cultures.¶Conclusion/interpretation. These data suggest that the outgrowth of diabetic ectoplacental cones is impaired by high glucose concentrations. [Diabetologia (2000) 43: 939–945]

Identification of ‘renin’-containing cells in the choriodecidua

Chorionic trophoblast, decidual cells, and macrophages have all been named as the site of renin in the placental membranes. To establish more clearly the nature of the renin-containing cells in the placental membranes, double immunostaining techniques were used to stain renin and specific cell markers in the same tissue sections. Cytokeratin was selected as an ectodermal cell marker and CD68 as a cytoplasmic macrophage marker. Cross-binding between antibodies was prevented by blocking species-related binding sites between the first and second sequence of the double-immunostaining procedures and by using highly selective immunostaining techniques in the second sequence. The results clearly show renin immunostaining in CD68-positive macrophages and not in cytokeratin-positive trophoblast. The anti-renal renin monoclonal antibody showed high affinity cross-reactivity with cathepsin D, another aspartic proteinase that can release angiotensin I from angiotensinogen. This should be seen in the context of earlier findings that only two of four anti-renal renin monoclonal antibodies showed staining in uterine and placental tissues and both cross-reacted with cathepsin D. The results indicate that differentiation between renin and cathepsin D and, possibly, other substances with shared properties and epitope homology deserves more attention than it has received thus far.

Prostacyclin and thromboxane in pregnancy

Normal pregnancy is characterized by a reduction of the peripheral vascular resistance, an increased plasmavolume and an increased cardiac output. Hypertension during pregnancy and pre-eclampsia occurs mostly in nulliparae after the 20th week of pregnancy. In this complication of pregnancy an incomplete vascular adaptation of pregnancy is present. The disorder is responsible for an increased maternal and perinatal morbidity and mortality. Since a few years information is available on the role of prostacyclin (prostaglandin It, PGI,) and thromboxane (TXA,) in the cardiovascular changes of pregnancy and in the pathogenesis of preeclampsia. PGI, and TXA, are both metabolites of arachidonic acid (AA). AA is mainly stored in esterform in phospholipids of the cellmembrane and is released by phospholipase A2 and phospholipase C. Free AA is metabolized by the enzyme prostaglandin-H-synthase to the cyclic endoperoxides prostaglandin Gz (PGG2) and prostaglandin H, (PGH2). In the platelets, under influence of thromboxaneA,-synthase, PGH2 is preferentially converted to TXA,, which is spontaneously hydrolised to the inactive TXB2. TXA, is a strong inductor of platelet aggregation and is also a vasoconstrictor. In the endothelial cells of the vascular wall PGHz in preferentially converted to PGI, by the enzyme prostacyclinsynthetase. PGI, is hydrolised to the inactive 6 keto PGF,a. PGI, is an inhibitor of plateletaggregation and a strong vasodilatator.

Differential effects of IL-11 on rat blastocysts and decidua during the peri-implantation period.

PROBLEM: To study effects of interleukin‐11 (IL‐11) on blastocyst development and decidualization.