Tctex3, related to Drosophila Polycomblike, is expressed in male germ cells and mapped to the mouse t-complex

Tctex3 showing restricted expression in male germ cells has been isolated during the process of chromosome walking in the mouse t-complex region. The total sequence of Tctex3 cDNA predicts a protein of 580 amino acids with two C4HC3 type PHD fingers. The region containing this conserved motif is shared among members of the Polycomblike proteins that include the mouse M96 and Drosophila Polycomblike. A partial cDNA for a human homolog of Tctex3, HUTEX3, has also been isolated. Mouse Tctex3 gene was mapped adjacent to Tsc2 gene on mouse Chromosome (Chr) 17, and HUTEX3 was located closely to HSET gene in the HLA class II region of chromosome 6. Received: 10 April 1998 / Accepted: 22 June 1998     

Mechanisms Underlying Ectopic Persistent Tooth-Pulp Pain following Pulpal Inflammation

In order to clarify the peripheral mechanisms of ectopic persistent pain in a tooth pulp following pulpal inflammation of an adjacent tooth, masseter muscle activity, phosphorylated extracellular signal-regulated protein kinase (pERK) and TRPV1 immunohistochemistries and satellite cell activation using glial fibrillary acidic protein (GFAP) immunohistochemistry in the trigeminal ganglion (TG) were studied in the rats with molar tooth-pulp inflammation. And, Fluorogold (FG) and DiI were also used in a neuronal tracing study to analyze if some TG neurons innervate more than one tooth pulp. Complete Freund’s adjuvant (CFA) or saline was applied into the upper first molar tooth pulp (M1) in pentobarbital-anesthetized rats, and capsaicin was applied into the upper second molar tooth pulp (M2) on day 3 after the CFA or saline application. Mean EMG activity elicited in the masseter muscle by capsaicin application to M2 was significantly larger in M1 CFA-applied rats compared with M1 vehicle-applied rats. The mean number of pERK-immunoreactive (IR) TG cells was significantly larger in M1 CFA-applied rats compared with M1 vehicle-applied rats. Application of the satellite cell inhibitor fluorocitrate (FC) into TG caused a significant depression of capsaicin-induced masseter muscle activity and a significant reduction of satellite cell activation. The number of TRPV1-IR TG cells innervating M2 was significantly larger in M1 CFA-applied rats compared with M1 vehicle-applied rats, and that was decreased following FC injection into TG. Furthermore, 6% of TG neurons innervating M1 and/or M2 innervated both M1 and M2. These findings suggest that satellite cell activation following tooth pulp inflammation and innervation of multiple tooth pulps by single TG neurons may be involved in the enhancement of the activity of TG neurons innervating adjacent non-inflamed teeth that also show enhancement of TRPV1 expression in TG neurons, resulting in the ectopic persistent tooth-pulp pain following pulpal inflammation of adjacent teeth.     

Understanding the X chromosome inactivation cycle in mice: A comprehensive view provided by nuclear transfer

During mouse development, imprinted X chromosome inactivation (XCI) is observed in preimplantation embryos and is inherited to the placental lineage, whereas random XCI is initiated in the embryonic proper. Xist RNA, which triggers XCI, is expressed ectopically in cloned embryos produced by somatic cell nuclear transfer (SCNT). To understand these mechanisms, we undertook a large-scale nuclear transfer study using different donor cells throughout the life cycle. The Xist expression patterns in the reconstructed embryos suggested that the nature of imprinted XCI is the maternal Xist-repressing imprint established at the last stage of oogenesis. Contrary to the prevailing model, this maternal imprint is erased in both the embryonic and extraembryonic lineages. The lack of the Xist-repressing imprint in the postimplantation somatic cells clearly explains how the SCNT embryos undergo ectopic Xist expression. Our data provide a comprehensive view of the XCI cycle in mice, which is essential information for future investigations of XCI mechanisms.     

Prdm9 Polymorphism Unveils Mouse Evolutionary Tracks

PR/SET domain containing 9 (Prdm9) mediates histone modifications such as H3K4me3 and marks hotspots of meiotic recombination. In many mammalian species, the Prdm9 gene is highly polymorphic. Prdm9 polymorphism is assumed to play two critical roles in evolution: to diversify the spectrum of meiotic recombination hotspots and to cause male hybrid sterility, leading to reproductive isolation and speciation. Nevertheless, information about Prdm9 sequences in natural populations is very limited. In this study, we conducted a comprehensive population survey on Prdm9 polymorphism in the house mouse, Mus musculus. Overall M. musculus Prdm9 displays an extraordinarily high level of polymorphism, particularly in regions encoding zinc finger repeats, which recognize recombination hotspots. Prdm9 alleles specific to various M. musculus subspecies dominate in subspecies territories. Moreover, introgression into other subspecies territories was found for highly divergent Prdm9 alleles associated with t-haplotype. The results of our phylogeographical analysis suggest that the requirement for hotspot diversity depends on geographical range and time span in mouse evolution, and that Prdm9 polymorphism has not been maintained by a simple balanced selection in the population of each subspecies.     

The Tumor-Educated-Macrophage Increase of Malignancy of Human Pancreatic Cancer Is Prevented by Zoledronic Acid

We previously defined macrophages harvested from the peritoneal cavity of nude mice with subcutaneous human pancreatic tumors as “tumor-educated-macrophages” (Edu) and macrophages harvested from mice without tumors as “naïve-macrophages” (Naïve), and demonstrated that Edu-macrophages promoted tumor growth and metastasis. In this study, Edu- and Naïve-macrophages were compared for their ability to enhance pancreatic cancer malignancy at the cellular level in vitro and in vivo. The inhibitory efficacy of Zoledronic acid (ZA) on Edu-macrophage-enhanced metastasis was also determined. XPA1 human pancreatic cancer cells in Gelfoam co-cultured with Edu-macrophages proliferated to a greater extent compared to XPA1 cells cultured with Naïve-macrophages (P = 0.014). XPA1 cells exposed to conditioned medium harvested from Edu culture significantly increased proliferation (P = 0.016) and had more migration stimulation capability (P<0.001) compared to cultured cancer cells treated with the conditioned medium from Naïve. The mitotic index of the XPA1 cells, expressing GFP in the nucleus and RFP in the cytoplasm, significantly increased in vivo in the presence of Edu- compared to Naïve-macrophages (P = 0.001). Zoledronic acid (ZA) killed both Edu and Naïve in vitro. Edu promoted tumor growth and metastasis in an orthotopic mouse model of the XPA1 human pancreatic cancer cell line. ZA reduced primary tumor growth (P = 0.006) and prevented metastasis (P = 0.025) promoted by Edu-macrophages. These results indicate that ZA inhibits enhanced primary tumor growth and metastasis of human pancreatic cancer induced by Edu-macrophages.     

New Evidence Confirms That the Mitochondrial Bottleneck Is Generated without Reduction of Mitochondrial DNA Content in Early Primordial Germ Cells of Mice

In mammals, observations of rapid shifts in mitochondrial DNA (mtDNA) variants between generations have led to the creation of the bottleneck theory for the transmission of mtDNA. The bottleneck could be attributed to a marked decline of mtDNA content in germ cells giving rise to the next generation, to a small effective number of mtDNA segregation units resulting from homoplasmic nucleoids rather than the single mtDNA molecule serving as the units of segregation, or to the selective transmission of a subgroup of the mtDNA population to the progeny. We have previously determined mtDNA copy number in single germ cells and shown that the bottleneck occurs without the reduction in germline mtDNA content. Recently one study suggested that the bottleneck is driven by a remarkable decline of mtDNA copies in early primordial germ cells (PGCs), while another study reported that the mtDNA genetic bottleneck results from replication of a subpopulation of the mtDNA genome during postnatal oocyte maturation and not during embryonic oogenesis, despite a detected a reduction in mtDNA content in early PGCs. To clarify these contradictory results, we examined the mtDNA copy number in PGCs isolated from transgenic mice expressing fluorescent proteins specifically in PGCs as in the aforementioned two other studies. We provide clear evidence to confirm that no remarkable reduction in mtDNA content occurs in PGCs and reinforce that the bottleneck is generated without reduction of mtDNA content in germ cells.     

Tetraploid embryos rescue the early defects of tw5/tw5 mouse embryos

tclw5 is a t-complex recessive lethal mutation of the tw5-haplotype. Since tw5/tw5 embryos die soon after implantation, the tclw5 gene is thought to play an important role in early embryogenesis. Previous histological studies have demonstrated that tw5 homozygotes do not survive past the gastrulation stage due to extensive death of the embryonic ectoderm, whereas the extraembryonic tissues were less affected. In the present study, we demonstrate that tw5/tw5 embryos may be distinguished from wildtype littermates at embryonic (E) day 5.5. At this stage, the visceral endoderm of tw5/tw5 embryos appeared to be different, possessing smaller and fewer vacuoles compared to normal littermates. This led us to hypothesize that the visceral endoderm may be affected by tclw5. Confirmation was provided by the rescue of tw5/tw5 embryos following aggregation with tetraploid embryos. However, rescued embryos did not survive past E9.0 and displayed an underdeveloped posterior region. This would indicate that the actions of tclw5 extend beyond the midgestation stage.