Li-yi Cai

PROP1 coexists with SOX2 and induces PIT1-commitment cells

Prophet of PIT1 (PROP1) is a pituitary-specific factor and responsive gene for the combined pituitary hormone deficiency in Ames dwarf mice and human patients. Our immunohistochemical studies demonstrated that PROP1 is consistently expressed in SOX2-expressing stem/progenitor cells in the rat pituitary from embryonic (E) to postnatal periods. At E13.5, all the cells in Rathkes pouch, the primordium of the pituitary, express PROP1. Afterward, PROP1-positive cells localize along the marginal cell layer, a putative stem cell niche in the pituitary, and stratify in the parenchyma of the anterior pituitary. In the embryonic period, PROP1 coexists transiently with PIT1, which is the anterior pituitary-specific factor and is a target of PROP1, but not any hormones. Thus, the present results imply a regulatory role of PROP1 not only in pituitary organogenesis but also in conversion of PIT1-lineage cells.

Significant quantitative and qualitative transition in pituitary stem / progenitor cells occurs during the postnatal development of the rat anterior pituitary.

We reported recently that a pituitary‐specific transcription factor PROP1 is present in SOX2‐positive cells and disappears at the early stage of the transition from progenitor cell to committed cell during the embryonic development of the rat pituitary. In the present study, we examined the localisation and identification of SOX2‐positive and PROP1/SOX2‐positive cells in the neonatal and postnatal rat pituitaries by immunohistochemistry. Quantitative analysis of immunoreactive cells demonstrated that SOX2‐positive pituitary stem/progenitor cells are not only predominantly localised in the marginal cell layer, but also are scattered in the parenchyma of the adult anterior lobe. In the marginal cell layer, the number of PROP1/SOX2‐positive cells significantly decreased after postnatal day 15, indicating that a significant quantitative transition is triggered in the marginal cell layer during the first postnatal growth wave of the anterior pituitary. By contrast, other phenotypes of SOX2‐positive stem/progenitor cells that express S100β appeared in the postnatal anterior pituitary. These data suggested that quantitative and qualitative transition occurs by acquisition of a novel mechanism in terminal differentiation in the postnatal development of the anterior pituitary.

Lack of an association human dioxin detoxification gene polymorphisms with endometriosis in Japanese women: results of a pilot study

ObjectivesEndometriosis is a chronic disease caused by the presence of endometrial tissue in ectopic locations outside the uterus. Chronic exposure to the environmental pollutant dioxin has been correlated with an increased incidence in the development of endometriosis in non-human primates. We have therefore examined whether there is an association between the polymorphisms of ten dioxin detoxification genes and endometriosis in Japanese women.MethodsThis was a pilot study in which 100 patients with endometriosis and 143 controls were enrolled. The prevalence of five microsatellite and 28 single nucleotide polymorphism markers within ten dioxin detoxification genes (AhR, AHRR, ARNT, CYP1A1, CYP2E1, EPHX1, GSTM1, GSTP1, GSTT1, NAT2) was examined.ResultsTaking into account that this analysis was a preliminary study due to its small sample size and genetic power, the results did not show any statistically significant difference between the cases and controls for any of the allele and genotype frequency distributions examined. In addition, no significant associations between the allele/genotype of all polymorphisms and the stage (I–II or III–IV) of endometriosis were observed.ConclusionBased on the findings of this pilot study, we conclude the polymorphisms of the ten dioxin detoxification genes analyzed did not contribute to the etiology of endometriosis among our patients.

HSV type 1 thymidine kinase protein accumulation in round spermatids induces male infertility by spermatogenesis disruption and apoptotic loss of germ cells

HSV type 1 thymidine kinase (HSV1-TK)-introduced transgenic rodents and HSV-infected humans were reported to suffer male infertility. The present study aimed to find novel clues to clarify the cause of HSV1-TK-induced male infertility using an HSV1-tk transgenic rat line. Two truncated HSV1-TK proteins, 37 and 39kDa, were produced and accumulated in the round spermatids, and their transcription initiation site was identified for the first time at the 65 base downstream of the translation start point of the full-length 43kDa HSV1-TK. Spermatozoa from those young transgenic rats showed malformed heads, looped tails, and missing cell membrane in heads and tails. Furthermore, age-dependent germ cell loss was observed. TUNEL assay suggested that this germ cell loss is caused by increased apoptotic germ cell death. These results suggest that the expression of HSV1-TK in testes brings about not only abnormal spermiogenesis but also a loss of germ cells due to apoptosis. These findings could provide a novel clue to elucidate the molecular mechanism underlying male infertility in transgenic animals and HSV-infected patients.

Dioxins in ascites and serum of women with endometriosis: a pilot study

BACKGROUND Animal studies and laboratory experiments have demonstrated that exposure to dioxins may be involved in the pathophysiology of endometriosis. However, recent epidemiological investigations have shown conflicting results. Although peritoneal fluid is a specific microenvironment playing a pivotal role in the development of endometriosis, to our knowledge, there is no published study evaluating the concentrations of dioxins in serum and peritoneal fluid simultaneously. The present study explores the possible correlation between the local peritoneal fluid levels of dioxins and concurrent endometriosis. METHODS There were 17 infertile women enrolled in the present study. After the diagnostic laparoscopic examination, the women were divided into two groups: endometriosis (n = 10) and controls (n = 7). We measured 29 dioxins simultaneously in serum and peritoneal fluid samples: 7 polychlorinated dibenzo-p-dioxins (PCDDs), 10 polychlorinated dibenzofurans (PCDFs), and 12 polychlorinated biphenyls (dioxin-like PCBs). A dioxin toxic equivalency (TEQ) system was utilized to calculate the dioxin concentration in each sample. RESULTS Serum concentrations of itemized components of 29 dioxins were similar in the endometriosis patients compared with the controls. Higher concentrations of PCDFs and dioxin-like PCBs were observed in peritoneal fluid than in serum, whereas the reverse was shown for PCDDs. Statistical analysis showed that higher levels of dioxin TEQ (PCDDs and PCDFs) in peritoneal fluid were significantly associated with an increased risk of endometriosis (OR: 2.5; 95% CI: 1.17-5.34; P = 0.035). CONCLUSIONS This is the first report suggesting that higher concentrations of dioxins (PCDDs and PCDFs) in peritoneal fluid are linked to endometriosis. More detail and epidemiological research is warranted to further explore this link.

The highly related LIM factors, LMO1, LMO3 and LMO4, play different roles in the regulation of the pituitary glycoprotein hormone α-subunit (αGSU) gene.

LMO1, LMO3 and LMO4 were cloned from the adult porcine pituitary cDNA library. Amino acid sequences of porcine LMO1, LMO3 and LMO4 were highly conserved among mammalian species. Transfection assay of the pituitary-derived cell line L beta T2 was carried out using the pituitary alpha GSU (glycoprotein hormone alpha-subunit) promoter (-1059/+12 b) fused to pSEAP2-Basic vector as a reporter gene. The results demonstrated that, whereas LMO4 showed no apparent effect, alpha GSU promoter activity was markedly repressed by LMO1 but activated by LMO3, indicating the different roles of the three highly homologous proteins, LMO1, LMO3 and LMO4. Knockdown assay by LMO siRNAs (small interfering RNAs) confirmed the above results for LMO1 and LMO3, whereas that by LMO4 siRNA increased the expression, indicating different modes of action. RT-PCR (reverse transcription-PCR) for total RNAs of several cell lines showed that LMO1 and LMO4 mRNAs were present ubiquitously in all cell lines, except for LMO1 in L929 cells. In contrast, LMO3 mRNA was abundant only in L beta T4 and GH3 cells with only small amounts in L beta T2 and MtT/S cells, indicating the cell-type-specific function of this protein. Real-time analyses of porcine pituitary ontogeny revealed that the three LMO genes are expressed during the fetal period and decline immediately afterwards, followed by a remarkably low level of LMO3 and LMO4 after birth. RT-PCR of the porcine tissues examined showed ubiquitous expression of LMO4, whereas LMO1 and LMO3 are expressed tissue specifically. Thus the present study demonstrated that three highly related LIM cofactors, LMO1, LMO3 and LMO4, have different effects on alpha GSU gene expression in the pituitary glands.

Failure to detect significant association between estrogen receptor-alpha gene polymorphisms and endometriosis in Japanese women

ObjectivesThe aim of the study was to test whether estrogen receptor 1 (ESR1) gene polymorphisms are correlated with the risk of the development of endometriosis in Japanese women, as a preliminary study.MethodsTo compare allelic frequencies and genotype distributions, a case-control study of 100 affected women and 143 women with no evidence of disease was performed using 10 microsatellite repeat markers and 66 single-nucleotide polymorphisms (SNPs) in the ESR1 gene region.ResultsAlthough our results might be insufficient to detect genetic susceptibility, owing to the small sample size and low genetic power, statistical analysis of the differences in allelic frequency between the cases and controls at each microsatellite locus demonstrated that no microsatellite locus in the ESR1 gene displayed a significant association with the disease when multiple testing was taken into account. Also, there were no statistically significant differences in the SNP allele frequencies and genotypes between the cases and controls when multiple testing was taken into account.ConclusionThe findings in our pilot study suggest that ESR1 polymorphisms do not contribute to endometriosis susceptibility.

Mapping of susceptibility locus for endometriosis within the HLA region using microsatellite markers in Japanese women

Endometriosis is a female disorder characterized by the presence of uterine endometrial tissue in ectopic loci. Previous studies reported a higher prevalence of particular human leukocyte antigen (HLA) in endometriosis. In order to confirm the association between endometriosis and the HLA region, 15 polymorphic microsatellite markers distributed in the HLA class II to class III region were subjected to association analysis by polymerase chain reaction (PCR)-based DNA typing of 89 patients and 136 healthy controls. Statistical analysis of the allelic frequency at each microsatellite locus showed that there were no statistically significant differences in the allele frequency distributions between the cases and controls. This finding suggests that the etiology of endometriosis does not involve the HLA class II genomic region and a portion of class III genomic region in the Japanese population.

Presence of human herpes virus 1-thymidine kinase in testis of azoospermic infertile herpes-infected patients

Infection with human herpes virus 1 (HHV1) is a suspected cause of human male infertility. However, the correlation between HHV1 infection and infertility is still unclear. We have previously generated transgenic rats that ectopically express the HHV1 thymidine kinase gene (HHV1-TK) in post-meiotic spermatids and found they had aberrant spermatogenesis and infertility. Therefore, we hypothesized that human infertility might be caused by HHV1 infection. Here, we examined whether HHV1-TK is expressed in human testis by analyzing the presence of its transcript and protein. Specimens were collected by biopsy from 30 azoospermic infertile male patients. RT-PCR and immunohistochemistry showed that 23 patients were positive for HHV1-TK expression, while seven patients were negative. Thus, we demonstrated HHV1-TK expression, indicating HHV1 infection, in the testis of human azoospermic infertile males for the first time; our findings represent a great advancement toward the verification of our hypothesis that HHV1-TK expression might cause human infertility.

Future for Elective Single Embryo Transfer: Review on Prognostic Aspect of Multiple Pregnancy

Abstract On the other side of the worldwide advance in assisted reproductive technology (ART), there is increasing concern about the maternal, fetal and post-natal risks of ART. To avoid multiple pregnancies, use of elective single embryo transfer (eSET) has been suggested in north European countries. In this review, we attempted to present an evidence-based discussion of the risks/benefits of ART and eSET. While many perinatal risks might be worse in ART than with natural conception in singleton pregnancies, the case is different in twin pregnancies and mainly depends on their monochorionic nature. Especially for early detection of intrauterine growth restriction, it is important to pay special attention to the location of umbilical cord insertion in the placenta. To evaluate the effectiveness of eSET, more studies are necessary with randomized controls.