Meijin Tochigi

Murine fetal resorption and experimental pre-eclampsia are induced by both excessive Th1 and Th2 activation

It has been proposed that immune responses in mammalian normal pregnancy are Th2-like, thereby protecting the fetus and placenta from being rejected. Administration of exogenous Th1 cytokines into pregnant mice is reported to induce feto-placental resorption. However, the effects of exogenous Th2 cytokines and Th2 directed responses in pregnant animals have not been well studied. In this study, we examined IL-4 and IL-12, which play decisive roles in the development of Th2 and Th1 responses, respectively, in the induction of fetal resorption and development of experimental pre-eclampsia. Transfer of either IL-4 and/or IL-12 stimulated splenocytes from BALB/C virgin female mice into BALB/C pregnant mice mated with either C57BL/6 or BALB/C male mice resulted in fetal resorption and glomerular nephritis associated with hypertension and proteinuria. In mice treated with IL-12 stimulated splenocytes, fatty liver degeneration associated with bile retention was observed. These results indicate that both excessive Th1 and Th2 activation contribute to the development of fetal resorption and pre-eclampsia, but that Th1 is critical to the development of liver degeneration.

Effects of Paternal Lymphocyte Immunization on Peripheral Th1/Th2 Balance and TCR Vβ and VΓ Repertoire Usage of Patients with Recurrent Spontaneous Abortions

PROBLEM: The mechanism of immunotherapy for patients with recurrent spontaneous abortions is not well understood. In order to investigate the suppressor mechanism of paternal lymphocyte immunization, we examined peripheral blood lymphocyte subpopulations and the repertoire of T‐cell receptor (TCR) gene segments. METHOD OF STUDY: Twelve patients with recurrent miscarriage were treated with immunization with paternal lymphocyte vaccinations three times during 12–14 weeks. Before and 2 weeks after the final inoculation, lymphocyte subsets and intra‐cellular interferon (IFN)‐Γ and/or interleukin (IL)‐4 production were examined by flow cytometry. TCR Vβ and VΓ repertoires were examined by semi‐quantitative reverse transcription‐polymerase chain reaction (RT‐PCR). RESULTS: We found no significant difference in CD4/CD8 ratios, prevalence of CD56+CD3+ or CD57+CD3+ cells (possible extrathymic T cells), ΓΔT cells, and CD5+CD19+ (B‐1) cells. However, by in vitro activation with phorbol 12‐mytistate 13‐acetate (PMA) and ionomycin, peripheral CD4 cells demonstrated a significant decrease of IFN‐Γ‐producing T helper 1 (Th1) cells and an increase of IL‐4‐producing T helper 2 (Th2) cells after immunotherapy. Seven of nine patients who exhibited remarkable decreases in Th1/Th2 ratios became pregnant within 6 months after three courses of immunotherapy, and four women delivered healthy babies, while none of the three patients who exhibited an increased or unchanged Th1/Th2 ratio had full‐term pregnancies (χ2<0.0001). Further, changes in usage of TCR Vβ and VΓ gene segments were observed after immunotherapy in six patients examined. CONCLUSION: Our findings suggest a shift of Th1‐dominant to Th2‐dominant status by vaccination might play important roles in maintaining successful pregnancies. Induction of some T cells that utilize different TCR repertoires possibly suppresses maternal rejection reactions.

AROMATASE INHIBITORS IN CIGARETTE SMOKE, TOBACCO LEAVES AND OTHER PLANTS

A chance observation that cigarette smoke interferes with the aromatase assay led us to investigate tobacco leaf and smoke extracts for the presence of aromatase inhibitors. The highest inhibitory activity was found in the basic fraction of cigarette smoke. Further purification of this fraction led to the identification of N-n-octanoylnornicotine. Synthesis and testing of a series of acylated nornicotines and anabasines for their ability to inhibit aromatase showed an interesting correlation of activity with the length of the acyl carbon chain, with maximum activity at C-11. The acylated derivatives showed activity which was significantly greater than that of nicotine and anabasine. In vivo studies in rats indicated that administration of this inhibitor delayed the onset of NMU-induced breast carcinoma and altered the estrus cycle. These in vivo studies suggest that tobacco alkaloid derivatives exert their effects by suppression of the aromatase enzyme system. Toxicity studies indicated relatively low toxicity with LD50 for N-n-octanoylnornicotine = 367 mg/kg body weight. When extracts from thirty five varieties of vegetables, plant leaves, and fruits were analyzed, seventeen showed quantitatively significant aromatase inhibition which was comparable to that of green tobacco leaf, suggesting that naturally occurring substances may affect endocrine function through aromatase inhibition.

The abortifacient effect of 16,16-dimethyl-trans-δ2-PGE1 methyl ester, a new prostaglandin analogue, on mid-trimester pregnancies and long-term follow-up observations

The present clinical trials revealed that 16,16-Dimethyl-trans-delta 2-PGE1 methyl ester in the form of vaginal suppositories is highly effective in inducing mid-trimester termination of pregnancies. It also showed that prior treatment with laminaria and metreurynter may enhance the success rate while reducing the incidence and severity of side effects. It is easy and safe to use clinically, with minimal side effects, and in our series, revealed no deleterious effects on ensuing reproductive physiology. However, the definite mechanism involved in the action of this new analogue to cause myometrial contractions is still not completely understood, and requires further intensive investigation.

Expression of the recombinase-activating gene (RAG-1) in murine early embryogenesis

The recombinase activation genes, RAG‐1 and RAG‐2, are expressed together in immature T or B lymphocytes and possess activity to induce V(D)J rearrangement in T cell receptor (TCR) and Ig genes. In vertebrates, only Ig and TCR molecules are reported to have recombination in their development using multiple V, D, J component gene segments. Thus, expression of RAG genes are localized only in lymphoid organs and sites of extrathymic T cell differentiation. In this study, we have used RAG‐1 and RAG‐2 genes as markers of possible genetic recombination in developing murine preimplanation embryos, using the highly sensitive reverse transcriptase polymerase chain reaction (RT‐PCR) technique and in situ hybridization. From 40 preimplantation embryos of various developmental stages we extracted RNA. reverse‐transcribed it into cDNA and used it in RT‐PCR studies. A PCR of 35 cycles disclosed expression of RAG‐1 but not RAG‐2 in morulae and blastocysts. Southern blot hybridization using a specific synthetic oligonucleotide probe for RAG‐1 and RT‐PCR with another primer pair identified RAG‐1 expression in developing embryos. In situ hybridization using a cooled CCD camera also revealed localization of RAG‐1 mRNA in blastocysts. We propose possible genetic recombination during late preimplantation murine embryogenesis which may contribute to the loss of totipotency and differentiation of inner cell mass and trophoectoderm.

Role of eicosanoids in mouse ovulation

previous data are available on diffusion of calcium (Ca) and magnesium (Mg) through fetal membranes, in present study we have examined the permeability to these cations of chorioamniotic membranes obtained from women at term or preterm labor. Study Methods: Ca and Mg permeability was evaluated using a modified dual chamber method previously described by Lee (1994). Analysis of cations in the samples was done with ion chromatography using METROHM IC-690 chromatograph. The double compartment model (Godfrey, 1983) was modified to accommodate sample extractions Results: The diffusion of Ca across fetal membrane from term labor was markedly higher than in preterm labor. Perfusion rates for Ca and Mg were also higher in term than preterm and the differences were statistically significant. Conclusions: The data of the present study showing reduced permeability of Ca and Mg of fetal membranes from preterm labor suggest that this abnormality could be an important factor for the activation of myometrium in preterm labor.