Naohiko Kuno

Female sterility in mice lacking the basigin gene, which encodes a transmembrane glycoprotein belonging to the immunoglobulin superfamily

Basigin (Bsg) is a transmembrane glycoprotein belonging to the immunoglobulin superfamily. Bsg knock‐out mice exhibit infertility of both sexes. Based on limited results, defective implantation has been considered to be the cause of the female infertility. We demonstrate here that disruption of the Bsg gene produces the failure of female reproductive processes including not only implantation but also fertilization. Bsg mRNA expression in cumulus cells and basolateral localization of the Bsg protein in the endometrial epithelium further support the importance of Bsg in these processes.

Expression of basigin, a member of the immunoglobulin superfamily, in the mouse central nervous system

Basigin (Bsg) is a transmembrane glycoprotein belonging to the immunoglobulin superfamily. Chicken Bsg (HT7/neurothelin/ 5A11) is expressed in neuroblasts, but disappears from neurons after a specific stage of cytodifferentiation, and becomes restrictedly expressed in the capillary endothelium in the adult brain. We show herein by means of in situ hybridization that Bsg mRNA was expressed in neuroblasts in 13.5 day old mouse embryos. In the adult mouse, Bsg was differentially expressed in subregions of the brain. Strong Bsg expression was detected in the limbic system, including the olfactory system, hippocampal formation, septal area, amygdala, thalamic anterior nuclei, hypothalamus, mesencephalic tegmentum, entorhinal cortex, and cingulate gyrus. Bsg was also intensely expressed in the retinal neuronal layers, the Vth layer of the cerebral neocortex, Purkinje cells of the cerebellum, several nuclei of the brain stem, and the gray matter of the spinal cord. Although in situ hybridization showed a weak signal in the brain capillary endothelium, protein expression of Bsg was strong enough to be detected by immunohistochemistry. Northern blot analysis confirmed the strong expression of Bsg in the central nervous system. Taking into account that Bsg knockout mice exhibit abnormalities in behavior, but a normal blood-brain barrier function, the present findings suggest that Bsg functions actively in neuronal interactions in the central nervous system.

Tissue Distribution of Placental Leucine Aminopeptidase/Oxytocinase During Mouse Pregnancy

Placental leucine aminopeptidase (P-LAP), also called oxytocinase, is an enzyme responsible for hydrolyzing oxytocin. This enzyme is identical to cystine aminopeptidase. We examined the tissue distribution of P-LAP in normal adult mice and also in mothers and fetuses during mouse pregnancy using immunohistochemical (IHC) analysis. P-LAP-immunoreactive protein was expressed in various organs in a cell- and gestational stage-dependent manner. In the kidney, P-LAP was located in distal and collecting tubules but not in proximal tubules. The islet of Langerhans in the maternal pancreas stained positively for P-LAP in the periphery in early gestation. This staining pattern changed so that both the periphery and inner cells were positive during mid-gestation and finally only inner cells were positive in late gestation. Among the hematopoietic cells in the fetal liver, only megakaryocytes showed strong expression of P-LAP. The staining intensity increased with gestation on the apical surface of trophoblasts in the placental labyrinth. These data demonstrate that P-LAP is present in a variety of tissues, and its presence is affected by pregnancy and fetal development. Therefore, P-LAP may play a significant role in various physiological processes in non-pregnant, pregnant, and fetal mice.

Determination of the optimal time and dosage of all-trans retinoic acid for induction of murine exencephaly

Murine exencephaly corresponds to human anencephaly, and provides a model for studying the mechanism of development of the central nervous system. A system which induces exencephaly at an extremely high rate is required in order to examine embryos, before the stage of neural tube closure, as samples of future exencephaly. Herein, we report on a system which is close enough to the best conditions for induction of this malformation, involving ICR mice and all-trans retinoic acid. The intraperitoneal administration of 30 mg/kg of all-trans retinoic acid at 03:00 hr on day 8 (copulatory plug, day 0) induced exencephaly in 81.6% of live embryos, as evaluated on day 10, with a 41.7% embryonic death rate. Earlier administration (more than 3 hr before) greatly increased the rate of embryonic death, whereas later administration resulted in a reduction in the rate of exencephaly. These findings suggest that a specific time during early development is crucial for neural tube closure, and provide a system which may facilitate the study of neural development and the pathophysiology of human anencephaly.

L-Threo-3,4-dihydroxyphenylserine (DOPS) aldolase: A new enzyme cleaving DOPS into protocatechualdehyde and glycine

An enzyme which cleaves L-threo-3,4-dihydroxyphenylserine into protocatechualdehyde and glycine was demonstrated in extracts of human brains. Equimolar production of protocachualdehyde and glycine was quantitatively confirmed using high-performance liquid chromatography. In subcellular fractions of the brain, the highest enzyme activity was found in cytosol and soluble fraction. L-threo-DOPS proved to be the best substrate for this enzyme. The L-erythroisomer was less active and D-threo- and D-erythro-isomers were essentially inactive. The enzyme activity has an optimal pH around 7.4, and requires pyridoxal phosphate for maximal activity.

Possible involvement of aminopeptidase A in hypertension in spontaneously hypertensive rats (shrs) and change of refractoriness in response to angiotensin Ii in pregnant Shrs

Background Hypertension complicated with pregnancy is a major cause of maternal and fetal mortality, but its pathophysiology is unclear. Objective To investigate the pressor response to angiotensin II (Ang II) and the involvement of the Ang II degrading protease, aminopeptidase A, in spontaneously hypertensive rats (SHRs). Design Pregnant SHRs and Wistar–Kyoto (WKY) rats were studied. Angiotensin II (200 ng/kg per min) or saline was infused by osmotic pump from day of 15 gestation, and caesarean section was performed at day 20 of gestation. Blood pressure during pregnancy, weight of placentas and pups at caesarean section, and aminopeptidase A activity in placenta and renal cortex were measured. Results Ang II treatment induced increases in blood pressure that were greater in non-pregnant WKY rats than those in pregnant WKY rats, pregnant SHRs, and non-pregnant SHRs. Renal aminopeptidase A activity in SHRs was significantly lower than that in WKY rats. Renal aminopeptidase A activity in pregnant SHRs was significantly greater than that in non-pregnant SHRs, but there was no significant increase in pregnant WKY rats. Placental aminopeptidase A activity in SHRs was greater than that in WKY rats. Placental aminopeptidase A activity in WKY rats was increased by Ang II, but was not increased in SHRs. Weights of placentas and pups were significantly lower in SHRs than in WKY rats. Conclusions Renal aminopeptidase A may be involved in the development of hypertension and the regulation of blood pressure in SHRs. Placental aminopeptidase A may be upregulated in response to fetal stress in pregnant SHRs.

Proliferation and stratification of keratinocyte on cultured amniotic epithelial cells for tissue engineering

Human amniotic epithelial cells (HAECs) are formed from amnioblasts, separated from the epiblast at about the 8th day after fertilization. Recent studies suggest that HAECs can produce various biologically active substances. In this study, the effects of cultured HAECs on keratinocytes were investigated. First of all, the effect of the medium conditioned by cultured HAECs on the proliferation of keratinocytes was examined. The conditioned medium significantly enhanced the proliferation (P<0.05). Next, the effect of co-culture with HAECs was also examined. The keratinocytes formed a stratified epithelium on day 7 after the start of co-culture. The cultured epithelium formed by the co-culture was five to six layers thick, could be detached by dispase treatment, and had sufficient strength as a sheet. These results suggest that HAECs will be a novel supplemental material for the tissue engineering of skin.

Decrease in severity of intrauterine growth retardation in subsequent pregnancies

Objective: Intrauterine growth retardation (IUGR) is likely to recur ina subsequent pregnancy. We investigated the obstetric features of recurrent cases and the severity of IUGR by comparing initial and subsequent deliveries. Methods: From a total of 12 567 deliveries, 95 women who were delivered of small‐for‐gestational‐age (SGA) infants and who became pregnant again within 5 years, were enrolled. A retrospective, comparative study of recurrent and non‐recurrent groups was performed. Results: Twenty‐two of ninety‐five women gave birth to SGA infants again, and a relatively high risk of recurrence was confirmed, but no single recurrence‐associated features were revealed. Within the recurrent group, the degree of IUGR was more severe in only five cases in the subsequent pregnancy. Conclusions: IUGR tends to recur, but does not increase in severity in most cases. We conclude that there is no need for excessive concern about the recurrence of IUGR.