Hiroshi Chisaka

Human Parvovirus B19 Induces Cell Cycle Arrest at G 2 Phase with Accumulation of Mitotic Cyclins

ABSTRACT Human parvovirus B19 infects specifically erythroid progenitor cells, which causes transient aplastic crises and hemolytic anemias. Here, we demonstrate that erythroblastoid UT7/Epo cells infected with B19 virus fall into growth arrest with 4N DNA, indicating G2/M arrest. These B19 virus-infected cells displayed accumulation of cyclin A, cyclin B1, and phosphorylated cdc2 and were accompanied by an up-regulation in the kinase activity of the cdc2-cyclin B1 complex, similar to that in cells treated with the mitotic inhibitor. However, degradation of nuclear lamina and phosphorylation of histone H3 and H1 were not seen in B19 virus-infected cells, indicating that the infected cells do not enter the M phase. Accumulation of cyclin B1 was persistently localized in the cytoplasm, but not in the nucleus, suggesting that B19 virus infection of erythroid cells raises suppression of nuclear import of cyclin B1, resulting in cell cycle arrest at the G2phase. The B19 virus-induced G2/M arrest may be the critical event in the damage of erythroid progenitor cells seen in patients with B19 virus infection.

The incidence of, and factors leading to, parvovirus B19-related hydrops fetalis following maternal infection; report of 10 cases and meta-analysis

OBJECTIVES to clarify the approximation of the frequency of B19-related nonimmune hydrops fetalis (NIHF), and to know the critical period during which maternal infection led to NIHF. METHODS we investigated the characteristics of 10 cases of antenatal B19 infection diagnosed over the past 10 years in Miyagi prefecture, Japan, and performed a meta-analysis of these cases and those previously reported in the literature. RESULTS NIHF caused by intrauterine B19 infection was diagnosed between 11 and 23 weeks of gestation in 10 women over the past 10 years in Miyagi prefecture, Japan. The source of infection was the mothers older child in six out of 10 cases, and children at a kindergarten where the mothers worked in two cases. The interval between the onset of infection and the diagnosis of NIHF ranged from 2 to 6 weeks. B19 infection was responsible for 10 (15.2%) in 66 cases of aetiology unknown NIHF in this study, and for 57 (19.1%) of 299 cases of non-malformed or aetiology-unknown NIHF by meta-analysis of the literature. Meta-analysis of the 165 reported cases of antenatal B19 infection, including the 10 cases described above, showed that there was a 10.2% excess risk of fetal death in women infected with B19 during pregnancy and a 12.40% excess risk in women infected during the first 20 weeks of pregnancy. Transplacental transmission was confirmed in 69 (24.1%) of 286 cases. The mean gestational age at diagnosis of NIHF was 22.8 +/- 5.1 weeks. The mean interval between the onset of maternal infection and diagnosis of NIHF was 6.2 +/- 3.7 weeks. CONCLUSIONS these approximations will be useful for counselling and management for pregnant women. The critical period during which maternal infection led to NIHF correlated with the hepatic period of hematopoietic activity. These findings suggest that parvovirus B19 may have an affinity for erythroid lineage cells at the hepatic stage of hematopoiesis, which may strongly influence the clinical features of feto-maternal B19 infection.

Fetal signals and parturition

John R. G. Challis, Frank H. Bloomfield, Alan D. Bocking, Valentina Casciani, Hiroshi Chisaka, Kristin Connor, Xuesen Dong, Peter Gluckman, Jane E. Harding, Jim Johnstone, Wei Li, Stephen Lye, Kunihiro Okamura and Marina Premyslova Departments of Physiology, Medicine and Obstetrics and Gynecology, University of Toronto, CIHR Group in Fetal Development and Health Toronto, Canada; Liggins Institute, University of Auckland, New Zealand; Department of Obstetrics and Gynecology, Tohoku University, Sendai, Japan; and Program in Development and Fetal Health, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Canada

Parvovirus B19 infection induces apoptosis of erythroid cells in vitro and in vivo

OBJECTIVE intrauterine parvovirus B19 infection is related to non-immune hydrops fetalis, the pathogenesis of which is based on the strict tropism of B19 for erythroid precursor cells and the massive destruction of the infected erythroid cells, although the mechanism of beta19-induced cytotoxicity has not been studied in detail. The purpose of this study is to provide empirical evidence that beta19 induces apoptosis of erythroid cells both in vitro and ill vivo. METHODS we analysed culture cells infected in vitro by B19 and tissues of nine cases of hydrops fetalis caused by B19 intrauterine infection by histological and biological methods. RESULTS cells infected iil vitro by B19 showed nuclear changes characteristic of apoptosis by light microscopic examination and DNA extracted from the infected cells was fragmented. Electron microscopic examination showed the nuclei of infected cells contained crescent-shaped clumps of heterochromatin with increased density and double staining with anti-B1 9 antibody and terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate-digoxigenin nick-end labeling (TUNEL) confirmed apoptosis of individual cells. Tissues of cases of hydrops fetalis caused by B19 contained erythroid cells with nuclear inclusions and characteristic nuclear changes of apoptosis by light microscopy. The double-staining confirmed apoptosis of erythroid cells in the tissues. Immunohistochemical analysis with antibodies against cellular factors involved in apoptosis showed that caspase3, p53 and p21 were positive in infected cells.

Maternal Feeding Controls Fetal Biological Clock

Background It is widely accepted that circadian physiological rhythms of the fetus are affected by oscillators in the maternal brain that are coupled to the environmental light-dark (LD) cycle. Methodology/Principal Findings To study the link between fetal and maternal biological clocks, we investigated the effects of cycles of maternal food availability on the rhythms of Per1 gene expression in the fetal suprachiasmatic nucleus (SCN) and liver using a transgenic rat model whose tissues express luciferase in vitro. Although the maternal SCN remained phase-locked to the LD cycle, maternal restricted feeding phase-advanced the fetal SCN and liver by 5 and 7 hours respectively within the 22-day pregnancy. Conclusions/Significance Our results demonstrate that maternal feeding entrains the fetal SCN and liver independently of both the maternal SCN and the LD cycle. This indicates that maternal-feeding signals can be more influential for the fetal SCN and particular organ oscillators than hormonal signals controlled by the maternal SCN, suggesting the importance of a regular maternal feeding schedule for appropriate fetal molecular clockwork during pregnancy.

Serologic study of human parvovirus B19 infection in pregnancy in Japan

OBJECTIVES To clarify the relationship between hydrops fetalis and parvovirus outbreaks in the community, seroprevalence of B19 antibody among women of childbearing age, and adverse effects of intrauterine B19 infection. METHODS Sera were collected from 168 cases of hydrops fetalis which were diagnosed between 1987 and 1997 in Miyagi prefecture, Japan, from 232 healthy pregnant women in 1987 and 277 healthy pregnant women in 1997 in Miyagi, and from 48 women infected with B19 during pregnancy. The sera were examined for B19 IgG and IgM antibodies by enzyme-linked immunosorbent assay and for B19 DNA by polymerase chain reaction. The number of cases of erythema infectiosum in Miyagi had been monitored each month. RESULTS Thirteen of the 168 cases of hydrops fetalis were found to be caused by intrauterine B19 infection and 12 of the 13 cases clustered in two periods of outbreaks of erythema infectiosum in the community. The positive rates of B19 IgG antibody between 1987 and 1997 were significantly different: 33% in 1987 and 46% in 1997. Nine of the 48 women infected during pregnancy showed adverse effects of the fetus: eight hydrops fetalis and one early abortion with positive B19 DNA. The fetal death rate (>12 weeks of gestation) among them was 15% (7/48), far higher than the calculated 1% among the general population. The nine mothers with adverse fetal outcomes had contact with the infectious source at the 16 weeks of gestation or earlier. CONCLUSIONS These data clearly showed a relationship between hydrops fetalis and parvovirus outbreaks in the community, and it may be important to follow the seroprevalence for an extrapolated period time to predict occurrence of hydrops fetalis caused by B19. Also the data indicated that the gestational week infection occurred is the most important determinant of an adverse effect to the fetus as described previously.

Effect of Pro-inflammatory cytokines on expression and activity of 11β-hydroxysteroid dehydrogenase type 2 in cultured human term placental trophoblast and human choriocarcinoma JEG-3 cells

Objective: 1 β-Hydroxysteroid dehydrogenase type 2 (11β-HSD2) is thought to act as a placental barrier protecting the fetus from high levels of maternal cortisol. On the other hand, intrauterine infection is one of the main causes of preterm birth and adverse fetal outcome, and pro-inflammatory cytokines may contribute to these adverse effects. However, the effect of pro-inflammatory cytokines on 11β-HSD2 is still not clear. Therefore, we have evaluated the effect of tumor necrosisfactor-α (TNF-α) and interleukin-1β (IL-1β) on 11β-HSD2 in cultured human placental trophoblast and in human choriocarcinoma JEG-3 cells. Methods: Placental trophoblast cells were isolatedfrom human term placenta. Placental trophoblast cells and JEG-3 cells were treated with TNF-α (0.1-10 ng/mL) or IL-1β (0.1-10 ng/mL). Real-time reverse transcription polymerase chain reaction and Western blot were used to study the regulation of 1 11β-HSD2 expression. 11β-HSD2 activity was determined by measuring the rate of cortisol to cortisone conversion in the culture medium using thin-layer chromatography (TLC). Results: In placental trophoblast, TNF-α and IL-1β down-regulated 11β-HSD2 mRNA expression and activity (both P <.05). The protein level was decreased only with IL-1β (P <.05). In JEG-3 cells, 11β-HSD2 mRNA was decreased by TNF-α but up-regulated by IL-1β, with no significant change in protein expression and activity. Conclusion: Our results suggest caution in interpreting data using JEG-3 cells. However, our studies with primary trophoblast suggest that TNF-α and IL-1β may increase the amount of cortisol crossing to the placenta and fetal circulation by attenuating 11β-HSD2 activity, potentially contributing to preterm labor and altered fetal outcome in uterine infection.

A transgenic mouse model for non-immune hydrops fetalis induced by the NS1 gene of human parvovirus B19

Human parvovirus B19 (B19) infection during pregnancy is associated with the adverse foetal outcome known as non-immune hydrops fetalis (NIHF). Although B19 is known to infect erythroid-lineage cells in vivo as well as in vitro, the mechanism leading to the occurrence of NIHF is not clear. To investigate the possible involvement of the B19 non-structural protein NS1 in NIHF, three independent lines of transgenic mice were generated that expressed NS1 under the control of the Cre-loxP system and the GATA1 promoter. Two of the three lines expressed NS1 in erythroid-lineage cells. Most of the transgenic mice died at the embryonic stage, some of which developed hydropic changes caused by severe anaemia at embryonic day 15.5 (E15.5). Histological examination of embryos at E15.5 showed significantly fewer erythropoietic islands in the liver parenchyma, whereas their hearts showed no abnormal signs, such as cardiomegaly and apoptotic cells. The NS1-transgenic mouse lines established here provide an animal model for human NIHF and suggest that NS1 plays a crucial role in the adverse outcome associated with intrauterine B19 infection in humans.

Intrapartum management guidelines based on fetal heart rate pattern classification

These guidelines provide the recommended response and treatment for intrapartum fetal management based on the classification of FHR patterns, as determined by the Perinatology Committee of the Japan Society of Obstetrics and Gynecology.

Pulmonary thromboembolism following gynecologic surgery and cesarean section.

Objective: To investigate the incidence and risk factors of symptomatic pulmonary thromboembolism (PTE) in patients following gynecologic surgery or cesarean section in Japan. Method: We examined the clinical records of all patients who have undergone gynecologic surgery or cesarean section at 32 hospitals affiliated with the Tohoku University Hospital between 1990 and 1999. Result: The total number of gynecologic surgical procedures and cesarean sections were 61 648 and 26 502, respectively. Twenty‐five of the 88 150 (0.028%) patients developed clinical PTE. Among the PTE patients who underwent gynecologic surgery, 86% were more than 40 years of age, 36% were obese (BMI>25 kg/m2) and 43% had uterine body cancer. Among the PTE patients who underwent cesarean section, 36% were more than 35 years of age and 55% were obese (BMI>29 kg/m2). Conclusion: Risk factors in Japanese patients undergoing gynecologic surgery may include age and cancer of the uterine body; risk factors for the development of PTE following cesarean section may include age and obesity.