Immunocytochemical Studies of Adhesion Molecules on Mouse UNK Cells and Their Extracellular Matrix Ligands During Mouse Pregnancy

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Uterine natural killer (uNK) cells are the dominant lymphocytes of pregnant mammals' uterus Studies have identified four differentiation stage of mouse uNK cells based on Dolichos biflorus lectin cytochemistry, and their distribution showed preferential domain in the uterus through out the pregnancy. This work was done to investigate the expression of alpha 5, alpha 6, and beta 7 integrins on uNK cells and their ligands distribution. Section of mouse uterus from sixth to seventeenth gestational days were submitted to immunocytochemistry and positive reactions for alpha 5, alpha 6, and beta 7 integrins were found on uNK from eighth to tenth gestational days but not after twelfth gestational days. Fibronectin reactions were seemed from sixth to tenth gestational days around uNK from the myometrium and endometrium close to the myometrium No reaction for fibronectin was seen in the decidualized and nondecidualized endometrium near the placenta Laminin reaction was seen Just in the antimesometrial side beta 7 integrin seems to be the active receptor to bind with VCAM-1. or MAdCA.M-1 of endothelial cells, promoting the uNK cross through the vessels. The absence of laminin in an uNK domain suggests these cells are not dependent of laminin and alpha 6 integrin for their establishment. However, fibronectin seems to support uNK migration, proliferation, differentiation, and survival in the uterus by binding with alpha 5 integrin. The loss of alpha 5 integrin ligation by the clown regulation or fibronectin could inhibits these events and further studies are need to investigate whether unligated alpha 5 can actively and initiate apoptosis, maybe in a caspase 8-dependent way that has been called integrin-mediated death. Anat Rec, 293 1081-1088,2010 (C) 2010 Wiley-Liss, Inc293610811088Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Subset classification of mouse uterine natural killer cells by DBA lectin reactivity

Uterine Natural Killer (uNK) cells are a transient lymphocyte population found in the pregnant uteri of human and rodents. The pregnant uterine environment appears to influence migration, differentiation and suppression of the cytolytic activation of uNK cells but the mechanisms involved in these processes are not well understood. Similarities to circulating NK (cNK) cells are limited. The present study sought to discrimate uNK cells from cNK cells in mice by identification of a unique uNK cell marker. Dolichos biflorus (DBA) lectin, which has high selectivity for glycoconjugates containing N-acetyl D-galactosomine in the terminal position, reacted with the plasma membranes of mouse uNK cells. DBA lectin did not react with other uterine lymphocytes or with cNK cell surfaces in Swiss, CBA-J, C57BL/6, SJL, BALB/c, DBA-2 mice strains. DBA lectin staining was useful for both light and electron microscopy and distinguished 4 uNK cell subtypes that appear to be stages of differentiation. Quantitative evaluation of these 4 uNK cell subtypes over early to late gestational times showed dynamic changes between immature and mature forms in different compartments of the implantation sites and indicated the occurrence of microdomains in the uterus capable of controlling uNK cell proliferation and differentiation. This is the first report showing mouse uNK cells expressing specific molecules not found in other NK cells. Use of this reagent should enhance studies of earlier, non-granulated forms of uNK cells and provide new strategies for purification of mouse uNK cells for functional and molecular studies. (C) 2003 Elsevier Science Ltd. All rights reserved.24547948

Functional analysis of murine uterine natural killer cells genetically devoid of oestrogen receptors

Uterine Natural Killer (uNK) cell differentiation in vivo requires oestrogen (E) priming prior to progesterone (P). Hybridomas between uNK precursor and SP2/0 cells express message for E receptor (ER)alpha but nor PR. However, mature, rodent and human uNK cells lack these receptors. To functionally assess requirements for uNK cell expression of ERalpha, or ERbeta during precursor differentiation, marrow was transplanted from either ERalphadegrees(/)degrees (alphaERKO) or ERbetadegrees(/)degrees (betaERKO) mice into alymphoid RAG-2degrees(/)degrees/gammacdegrees(/)degrees females. Recipients were mated and their implantation sites were examined by light microscopy, morphometry and ultrastructure. High numbers of uNK cells were established from each donor strain. Graft-derived uNK cells were similar in number and morphology to uNK cells of normal mice, suggesting that neither alpha- nor beta-ER is required for uNK precursor cell differentiation. Induction of spiral artery modification in the transplant recipients indicated that graft-derived uNK cells had functional properties. A novel technique for rapid isolation of highly purified uNK cells from normal mice using Dolichos biflorus agglutinin (DBA) lectin-conjugated magnetic beads was employed to obtain RNA. Expression of alpha- and beta-ER was absent by RT-PCR from NK cells isolated from the uterus, supporting the conclusions from the in vivo study. (C) 2003 Elsevier Science Ltd. All rights reserved.24440341

Hepatic morphological alterations, glycogen content and cytochrome P450 activities in rats treated chronically with N-omega-nitro-L-arginine methyl ester (L-NAME)

Chronic treatment of rats with N-omega stop-nitro-L-arginine methyl ester (L-NAME), an inhibitor of nitric oxide (NO) biosynthesis, results in hypertension mediated partly by enhanced angiotensin-I-converting enzyme (ACE) activity. We examined the influence of L-NAME on rat liver morphology, on hepatic glycogen, cholesterol, and triglyceride content, and on the activities of the cytochrome P450 isoforms CYP1A1/2, CYP2B1/2, CYP2C11, and CYP2E1. Male Wistar rats were treated with L-NAME (20 mg/rat per day via drinking water) for 2, 4, and 8 weeks, and their livers were then removed for analysis. Enzymatic induction was produced by treating rats with phenobarbital (to induce CYP2B1/2), beta-naphthoflavone (to induce CYP1A1/2), or pyrazole (to induce CYP2E1). L-NAME significantly elevated blood pressure; this was reversed by concomitant treatment with enalapril (ACE inhibitor) or losartan (angiotensin II AT(1) receptor antagonist). L-NAME caused vascular hypertrophy in hepatic arteries, with perivascular and interstitial fibrosis involving collagen deposition. Hepatic glycogen content also significantly increased. L-NAME did not affect fasting glucose levels but significantly reduced insulin levels and increased the insulin sensitivity of rats, based on an intraperitoneal glucose tolerance test. Immunoblotting experiments indicated enhanced phosphorylation of protein kinase B and of glycogen synthase kinase 3. All these changes were reversed by concomitant treatment with enalapril or losartan. L-NAME had no effect on hepatic cholesterol or triglyceride content or on the basal or drug-induced activities and protein expression of the cytochrome P450 isoforms. Thus, the chronic inhibition of NO biosynthesis produced hepatic morphological alterations and changes in glycogen metabolism mediated by the renin-angiotensin system. The increase in hepatic glycogen content probably resulted from enhanced glycogen synthase activity following the inhibition of glycogen synthase kinase 3 by phosphorylation.3291455