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  1. Transcriptional and Translational Heterogeneity among Neonatal Mouse Spermatogonia1

    PubMed Central

    Hermann, Brian P.; Mutoji, Kazadi N.; Velte, Ellen K.; Ko, Daijin; Oatley, Jon M.; Geyer, Christopher B.; McCarrey, John R.

    2015-01-01

    ABSTRACT Spermatogonial stem cells (SSCs) are a subset of undifferentiated spermatogonia responsible for ongoing spermatogenesis in mammalian testes. Spermatogonial stem cells arise from morphologically homogeneous prospermatogonia, but growing evidence suggests that only a subset of prospermatogonia develops into the foundational SSC pool. This predicts that subtypes of undifferentiated spermatogonia with discrete mRNA and protein signatures should be distinguishable in neonatal testes. We used single-cell quantitative RT-PCR to examine mRNA levels of 172 genes in individual spermatogonia from 6-day postnatal (P6) mouse testes. Cells enriched from P6 testes using the StaPut or THY1+ magnetic cell sorting methods exhibited considerable heterogeneity in the abundance of specific germ cell and stem cell mRNAs, segregating into one somatic and three distinct spermatogonial clusters. However, P6 Id4-eGFP+ transgenic spermatogonia, which are known to be enriched for SSCs, were more homogeneous in their mRNA levels, exhibiting uniform levels for the majority of genes examined (122 of 172). Interestingly, these cells displayed nonuniform (50 of 172) expression of a smaller cohort of these genes, suggesting there is substantial heterogeneity even within the Id4-eGFP+ population. Further, although immunofluorescence staining largely demonstrated conformity between mRNA and protein levels, some proteins were observed in patterns that were disparate from those detected for the corresponding mRNAs in Id4-eGFP+ spermatogonia (e.g., Kit, Sohlh2, Stra8), suggesting additional heterogeneity is introduced at the posttranscriptional level. Taken together, these data demonstrate the existence of multiple spermatogonial subtypes in P6 mouse testes and raise the intriguing possibility that these subpopulations may correlate with the development of functionally distinct spermatogenic cell types. PMID:25568304

  2. Distinct purinergic signaling pathways in prepubescent mouse spermatogonia.

    PubMed

    Fleck, David; Mundt, Nadine; Bruentgens, Felicitas; Geilenkirchen, Petra; Machado, Patricia A; Veitinger, Thomas; Veitinger, Sophie; Lipartowski, Susanne M; Engelhardt, Corinna H; Oldiges, Marco; Spehr, Jennifer; Spehr, Marc

    2016-09-01

    Spermatogenesis ranks among the most complex, yet least understood, developmental processes. The physiological principles that control male germ cell development in mammals are notoriously difficult to unravel, given the intricate anatomy and complex endo- and paracrinology of the testis. Accordingly, we lack a conceptual understanding of the basic signaling mechanisms within the testis, which control the seminiferous epithelial cycle and thus govern spermatogenesis. Here, we address paracrine signal transduction in undifferentiated male germ cells from an electrophysiological perspective. We identify distinct purinergic signaling pathways in prepubescent mouse spermatogonia, both in vitro and in situ. ATP-a dynamic, widespread, and evolutionary conserved mediator of cell to cell communication in various developmental contexts-activates at least two different spermatogonial purinoceptor isoforms. Both receptors operate within nonoverlapping stimulus concentration ranges, display distinct response kinetics and, in the juvenile seminiferous cord, are uniquely expressed in spermatogonia. We further find that spermatogonia express Ca(2+)-activated large-conductance K(+) channels that appear to function as a safeguard against prolonged ATP-dependent depolarization. Quantitative purine measurements additionally suggest testicular ATP-induced ATP release, a mechanism that could increase the paracrine radius of initially localized signaling events. Moreover, we establish a novel seminiferous tubule slice preparation that allows targeted electrophysiological recordings from identified testicular cell types in an intact epithelial environment. This unique approach not only confirms our in vitro findings, but also supports the notion of purinergic signaling during the early stages of spermatogenesis. PMID:27574293

  3. Benzo(a)pyrene Is Mutagenic in Mouse Spermatogonial Stem Cells and Dividing Spermatogonia

    PubMed Central

    O’Brien, Jason M.; Beal, Marc A.; Yauk, Carole L.; Marchetti, Francesco

    2016-01-01

    Although many environmental agents are established male germ cell mutagens, few are known to induce mutations in spermatogonial stem cells. Stem cell mutations are of great concern because they result in a permanent increase in the number of mutations carried in sperm. We investigated mutation induction during mouse spermatogenesis following exposure to benzo(a)pyrene (BaP). MutaMouse males were given 0, 12.5, 25, 50, or 100 mg/kg bw/day BaP for 28 days by oral gavage. Germ cells were collected from the cauda epididymis and seminiferous tubules 3 days after exposure and from cauda epididymis 42 and 70 days after exposure. This design enabled targeted investigation of effects on post-spermatogonia, dividing spermatogonia, and spermatogonial stem cells, respectively. BaP increased lacZ mutant frequency (MF) in cauda sperm after exposure of dividing spermatogonia (4.2-fold at highest dose, P < .01) and spermatogonial stem cells (2.1-fold at highest dose, P < .01). No significant increases in MF were detected in cauda sperm or seminiferous tubule cells collected 3 days post-exposure. Dose-response modelling suggested that the mutational response in male germ cells to BaP is sub-linear at low doses. Our results demonstrate that oral exposure to BaP causes spermatogonial stem cell mutations, that different phases of spermatogenesis exhibit varying sensitivities to BaP, with dividing spermatogonia representing a window of peak sensitivity, and that sampling spermatogenic cells from the seminiferous tubules at earlier time-points may underestimate germ cell mutagenicity. This information is critical to optimize the use of the international test guideline for transgenic rodent mutation assays for detecting germ cell mutagens. PMID:27208087

  4. Benzo(a)pyrene Is Mutagenic in Mouse Spermatogonial Stem Cells and Dividing Spermatogonia.

    PubMed

    O'Brien, Jason M; Beal, Marc A; Yauk, Carole L; Marchetti, Francesco

    2016-08-01

    Although many environmental agents are established male germ cell mutagens, few are known to induce mutations in spermatogonial stem cells. Stem cell mutations are of great concern because they result in a permanent increase in the number of mutations carried in sperm. We investigated mutation induction during mouse spermatogenesis following exposure to benzo(a)pyrene (BaP). MutaMouse males were given 0, 12.5, 25, 50, or 100 mg/kg bw/day BaP for 28 days by oral gavage. Germ cells were collected from the cauda epididymis and seminiferous tubules 3 days after exposure and from cauda epididymis 42 and 70 days after exposure. This design enabled targeted investigation of effects on post-spermatogonia, dividing spermatogonia, and spermatogonial stem cells, respectively. BaP increased lacZ mutant frequency (MF) in cauda sperm after exposure of dividing spermatogonia (4.2-fold at highest dose, P < .01) and spermatogonial stem cells (2.1-fold at highest dose, P < .01). No significant increases in MF were detected in cauda sperm or seminiferous tubule cells collected 3 days post-exposure. Dose-response modelling suggested that the mutational response in male germ cells to BaP is sub-linear at low doses. Our results demonstrate that oral exposure to BaP causes spermatogonial stem cell mutations, that different phases of spermatogenesis exhibit varying sensitivities to BaP, with dividing spermatogonia representing a window of peak sensitivity, and that sampling spermatogenic cells from the seminiferous tubules at earlier time-points may underestimate germ cell mutagenicity. This information is critical to optimize the use of the international test guideline for transgenic rodent mutation assays for detecting germ cell mutagens. PMID:27208087

  5. BMP4 Cooperates with Retinoic Acid to Induce the Expression of Differentiation Markers in Cultured Mouse Spermatogonia

    PubMed Central

    Feng, Yanmin; Feng, Xue; Wang, Xiuxia; Gan, Haiyun; Wang, Lixian; Lin, Xiwen

    2016-01-01

    Spermatogenesis is sustained by the proliferation and differentiation of spermatogonial stem cells (SSCs). However, the molecules controlling these processes remain largely unknown. Here, we developed a simplified high concentration serum-containing system for the culture of mouse SSCs. Analysis of SSCs markers and transplantation results revealed that the cultured spermatogonia retained stem cell characteristics after long-term in vitro propagation. Using this culture system, the expression and function of bone morphogenetic protein 4 (BMP4) were explored. Immunostaining showed that BMP4 was predominantly expressed in germ cells and that its level increased as spermatogenesis progresses. BMP4 receptors BMPR1A and BMPRII were present in spermatogonia, spermatocytes, and round spermatids. Moreover, despite the mRNAs of these two genes being present in mouse Sertoli cells, only BMPRII was detected by using Western blotting assays. While exogenous BMP4 by itself did not induce the expression of Stra8 and c-Kit, two marker genes of differentiating spermatogonia, a significant cooperative effect of BMP4 and retinoic acid (RA) was observed. Moreover, pretreatment of cultured spermatogonia with the BMP4 antagonist Noggin could inhibit RA-induced expression of these two marker genes. In conclusion, BMP4 may exert autocrine effects and act cooperatively with RA to induce the differentiation of spermatogonia in vivo. PMID:27795714

  6. Retinoic acid induces multiple hallmarks of the prospermatogonia-to-spermatogonia transition in the neonatal mouse.

    PubMed

    Busada, Jonathan T; Kaye, Evelyn P; Renegar, Randall H; Geyer, Christopher B

    2014-03-01

    In mammals, most neonatal male germ cells (prospermatogonia) are quiescent and located in the center of the testis cords. In response to an unknown signal, prospermatogonia transition into spermatogonia, reenter the cell cycle, divide, and move to the periphery of the testis cords. In mice, these events occur by 3-4 days postpartum (dpp), which temporally coincides with the onset of retinoic acid (RA) signaling in the neonatal testis. RA has a pivotal role in initiating germ cell entry into meiosis in both sexes, yet little is known about the mechanisms and about cellular changes downstream of RA signaling. We examined the role of RA in mediating the prospermatogonia-to-spermatogonia transition in vivo and found 24 h of precocious RA exposure-induced germ cell changes mimicking those that occur during the endogenous transition at 3-4 dpp. These changes included: 1) spermatogonia proliferation; 2) maturation of cellular organelles; and 3), expression of markers characteristic of differentiating spermatogonia. We found that germ cell exposure to RA did not lead to cellular loss from apoptosis but rather resulted in a delay of ∼2 days in their entry into meiosis. Taken together, our results indicate that exogenous RA induces multiple hallmarks of the transition of prospermatogonia to spermatogonia prior to their entry into meiosis.

  7. Paracrine Wnt/β-catenin signaling mediates proliferation of undifferentiated spermatogonia in the adult mouse testis.

    PubMed

    Takase, Hinako M; Nusse, Roeland

    2016-03-15

    Spermatogonial stem cells (SSCs) fuel the production of male germ cells but the mechanisms behind SSC self-renewal, proliferation, and differentiation are still poorly understood. Using the Wnt target gene Axin2 and genetic lineage-tracing experiments, we found that undifferentiated spermatogonia, comprising SSCs and transit amplifying progenitor cells, respond to Wnt/β-catenin signals. Genetic elimination of β-catenin indicates that Wnt/β-catenin signaling promotes the proliferation of these cells. Signaling is likely initiated by Wnt6, which is uniquely expressed by neighboring Sertoli cells, the only somatic cells in the seminiferous tubule that support germ cells and act as a niche for SSCs. Therefore, unlike other stem cell systems where Wnt/β-catenin signaling is implicated in self-renewal, the Wnt pathway in the testis specifically contributes to the proliferation of SSCs and progenitor cells.

  8. Paracrine Wnt/β-catenin signaling mediates proliferation of undifferentiated spermatogonia in the adult mouse testis

    PubMed Central

    Takase, Hinako M.; Nusse, Roeland

    2016-01-01

    Spermatogonial stem cells (SSCs) fuel the production of male germ cells but the mechanisms behind SSC self-renewal, proliferation, and differentiation are still poorly understood. Using the Wnt target gene Axin2 and genetic lineage-tracing experiments, we found that undifferentiated spermatogonia, comprising SSCs and transit amplifying progenitor cells, respond to Wnt/β-catenin signals. Genetic elimination of β-catenin indicates that Wnt/β-catenin signaling promotes the proliferation of these cells. Signaling is likely initiated by Wnt6, which is uniquely expressed by neighboring Sertoli cells, the only somatic cells in the seminiferous tubule that support germ cells and act as a niche for SSCs. Therefore, unlike other stem cell systems where Wnt/β-catenin signaling is implicated in self-renewal, the Wnt pathway in the testis specifically contributes to the proliferation of SSCs and progenitor cells. PMID:26929341

  9. The Effect of Dose Rate on the Frequency of Specific-Locus Mutations Induced in Mouse Spermatogonia is Restricted to Larger Lesions; a Retrospective Analysis of Historical Data

    SciTech Connect

    Russell, Liane B; Hunsicker, Patricia R

    2012-01-01

    A series of 19 large-scale germ-cell mutagenesis experiments conducted several decades ago led to the conclusion that low-LET radiation delivered to mouse spermatogonia at dose rates of 0.8 R/min and below induced only about one-third as many specific-locus mutations as did single, acute exposures at 24 R/min and above. A two-hit origin of the mutations was deemed unlikely in view of the then prevailing evidence for the small size of genetic lesions in spermatogonia. Instead, the dose-rate effect was hypothesized to be the result of a repair system that exists in spermatogonia, but not in more mature male reproductive cells. More recent genetic and molecular studies on the marker genes have identified the phenotypes associated with specific states of the mutant chromosomes, and it is now possible retrospectively to classify individual past mutations as "large lesions" or "other lesions". The mutation-frequency difference between high and low dose rates is restricted to the large lesion mutations, for which the dose-curve slopes differ by a factor exceeding 3.4. For other lesion mutations, there is essentially no difference between the slopes for protracted and acute irradiations; induced other lesions frequencies per unit dose remain similar for dose rates ranging over more than 7 orders of magnitude. For large lesions, these values rise sharply at dose rates >0.8 R/min, though they remain similar within the whole range of protracted doses, failing to provide evidence for a threshold dose rate. The downward bend at high doses that had been noted for X-ray-induced specific-locus mutations as a whole and ascribed to a positive correlation between spermatogonial death and mutation load is now found to be restricted to large lesion mutations. There is a marked difference between the mutation spectra (distributions among the seven loci) for large lesions and other lesions. Within each class, however, the spectra are similar for acute and protracted irradiation.

  10. Distinct requirements for Sin3a in perinatal male gonocytes and differentiating spermatogonia

    PubMed Central

    Gallagher, Shannon J.; Kofman, Amber E.; Huszar, Jessica M.; Dannenberg, Jan-Hermen; DePinho, Ronald A.; Braun, Robert E.; Payne, Christopher J.

    2012-01-01

    Chromatin modifier Swi-independent 3a (SIN3A), together with associated histone deacetylases, influences gene expression during development and differentiation through a variety of transcription factors in a cell-specific manner. Sin3a is essential for the maintenance of inner cell mass cells of mouse blastocysts, embryonic fibroblasts, and myoblasts, but is not required for the survival of trophectoderm or Sertoli cells. To better understand how this transcriptional regulator modulates cells at different developmental stages within a single lineage, we used conditional gene targeting in mice to ablate Sin3a from perinatal quiescent male gonocytes and from postnatal differentiating spermatogonia. Mitotic germ cells expressing stimulated by retinoic acid gene 8 (Stra8) that lacked Sin3a exhibited increased DNA damage and apoptosis, yet collectively progressed through meiosis and spermiogenesis and generated epididymal sperm at approximately 50% of control levels, sufficient for normal fertility. In contrast, perinatal gonocytes lacking Sin3a underwent rapid depletion that coincided with cell cycle reentry, exhibiting 2.5-fold increased histone H3 phosphorylation upon cycling that suggested a prophase/metaphase block; germ cells were almost entirely absent two weeks after birth, resulting in sterility. Gene expression profiling of neonatal testes containing Sin3a-deleted gonocytes identified upregulated transcripts highly associated with developmental processes and pattern formation, and downregulated transcripts involved in nuclear receptor activity, including Nr4a1 (Nur77). Interestingly, Nr4a1 levels were elevated in testes containing Stra8-expressing, Sin3a-deleted spermatogonia. SIN3A directly binds to the Nr4a1 promoter, and Nr4a1 expression is diminished upon spermatogonial differentiation in vitro. We conclude that within the male germline, Sin3a is required for the mitotic reentry of gonocytes, but is dispensable for the maintenance of differentiating

  11. Mammalian gonocyte and spermatogonia differentiation: recent advances and remaining challenges.

    PubMed

    Manku, Gurpreet; Culty, Martine

    2015-03-01

    The production of spermatozoa relies on a pool of spermatogonial stem cells (SSCs), formed in infancy from the differentiation of their precursor cells, the gonocytes. Throughout adult life, SSCs will either self-renew or differentiate, in order to maintain a stem cell reserve while providing cells to the spermatogenic cycle. By contrast, gonocytes represent a transient and finite phase of development leading to the formation of SSCs or spermatogonia of the first spermatogenic wave. Gonocyte development involves phases of quiescence, cell proliferation, migration, and differentiation. Spermatogonia, on the other hand, remain located at the basement membrane of the seminiferous tubules throughout their successive phases of proliferation and differentiation. Apoptosis is an integral part of both developmental phases, allowing for the removal of defective cells and the maintenance of proper germ-Sertoli cell ratios. While gonocytes and spermatogonia mitosis are regulated by distinct factors, they both undergo differentiation in response to retinoic acid. In contrast to postpubertal spermatogenesis, the early steps of germ cell development have only recently attracted attention, unveiling genes and pathways regulating SSC self-renewal and proliferation. Yet, less is known on the mechanisms regulating differentiation. The processes leading from gonocytes to spermatogonia have been seldom investigated. While the formation of abnormal gonocytes or SSCs could lead to infertility, defective gonocyte differentiation might be at the origin of testicular germ cell tumors. Thus, it is important to better understand the molecular mechanisms regulating these processes. This review summarizes and compares the present knowledge on the mechanisms regulating mammalian gonocyte and spermatogonial differentiation. PMID:25670871

  12. Production of donor-derived offspring by allogeneic transplantation of spermatogonia in the yellowtail (Seriola quinqueradiata).

    PubMed

    Morita, Tetsuro; Kumakura, Naoki; Morishima, Kagayaki; Mitsuboshi, Toru; Ishida, Masashi; Hara, Takashi; Kudo, Satomi; Miwa, Misako; Ihara, Shoko; Higuchi, Kentaro; Takeuchi, Yutaka; Yoshizaki, Goro

    2012-06-01

    Although the yellowtail (Seriola quinqueradiata) is the fish most commonly farmed in Japan, breeding of this species has not yet started. This is primarily due to the lack of sufficiently sophisticated methods for manipulating gametogenesis, which makes it difficult to collect gametes from specific dams and sires. If it were possible to produce large numbers of surrogate fish by transplanting germ cells isolated from donor individuals harboring desirable genetic traits, then the probability of acquiring gametes carrying the donor-derived haplotype would increase, and breeding programs involving this species might increase as a result. As a first step, we established a method for the allogeneic transplantation of yellowtail spermatogonia and the production of donor-derived offspring. Donor cells were collected from immature (10-month-old) yellowtail males with testes containing abundant type A spermatogonia, labeled with PKH26 fluorescent dye, and transferred into the peritoneal cavities of 8-day-old larvae. Fluorescence observation at 28 days post-transplantation revealed that PKH26-labeled cells were incorporated into recipients' gonads. To assess whether donor-derived spermatogonia could differentiate into functional gametes in the allogeneic recipient gonads, gametes collected from nine male and four female adult recipients were fertilized with wild-type eggs and milt. Analysis of microsatellite DNA markers confirmed that some of the first filial (F(1)) offspring were derived from donor fish, with the average contribution of donor-derived F(1) offspring being 66% and the maximum reaching 99%. These findings confirmed that our method was effective for transplanting yellowtail spermatogonia into allogeneic larvae to produce donor-derived offspring. PMID:22460666

  13. Depletion of the spermatogonia from the seminiferous epithelium of the rhesus monkey after X irradiation

    SciTech Connect

    van Alphen, M.M.; van de Kant, H.J.; de Rooij, D.G.

    1988-03-01

    In unirradiated testes large differences were found in the total number of spermatogonia among different monkeys, but the number of spermatogonia in the right and the left testes of the same monkey appeared to be rather similar. During the first 11 days after irradiation with 0.5 to 4.0 Gy of X rays the number of Apale spermatogonia (Ap) decreased to about 13% of the control level, while the number of Adark spermatogonia (Ad) did not change significantly. A significant decrease in the number of Ad spermatogonia was seen at Day 14 together with a significant increase in the number of Ap spermatogonia. It was concluded that the resting Ad spermatogonia are activated into proliferating Ap spermatogonia. After Day 16 the number of both Ap and Ad spermatogonia decreased to low levels. Apparently the new Ap spermatogonia were formed by lethally irradiated Ad spermatogonia and degenerated while attempting to divide. The activation of the Ad spermatogonia was found to take place throughout the cycle of the seminiferous epithelium. Serum FSH, LH, and testosterone levels were measured before and after irradiation. Serum FSH levels already had increased during the first week after irradiation to 160% of the control level. Serum LH levels increased between 18 and 25 days after irradiation. Serum testosterone levels did not change at all. The results found in the rhesus monkey are in line with those found in humans, but due to the presence of Ad spermatogonia they differ from those obtained in non-primates.

  14. Genetic mapping of tumor susceptibility genes involved in mouse plasmacytomagenesis

    SciTech Connect

    Mock, B.A.; Krall, M.M.; Dosik, J.K. )

    1993-10-15

    Plasmacytomas (PCTs) were induced in 47% of BALB/cAnPt mice by the intraperitoneal injection of pristane, in 2% of (BALB/c [times] DBA/2N)F[sub 1], and in 11% of 773 BALB/cAnPt [times] (BALB/cAnPt [times] DBA/2N)F[sub 1]N[sub 2] backcross mice. This result indicates a multigenic mode of inheritance for PCT susceptibility. To locate genes controlling this complex genetic trait, tumor susceptibility in backcross progeny generated from BALB/c and DBA/2N (resistant) mice was correlated with alleles of 83 marker loci. The genotypes of the PCT-susceptible progeny displayed an excess homozygosity for BALB/c alleles with a 32-centimorgan stretch of mouse chromosome 4 (>95% probability of linkage) with minimal recombination (12%) near Gt10. Another susceptibility gene on mouse chromosome 1 may be linked to Fcgr2 (90% probability of linkage); there were excess heterozygotes for Fcgr2 among the susceptible progeny and excess homozygotes among the resistant progeny. Regions of mouse chromosomes 4 and 1 that are correlated with PCT susceptibility share extensive linkage homology with regions of human chromosome 1 that have been associated with cytogenetic abnormalities in multiple myeloma and lymphoid, breast, and endocrine tumors. 68 refs., 2 figs., 1 tab.

  15. Sleeping Beauty mouse models identify candidate genes involved in gliomagenesis.

    PubMed

    Vyazunova, Irina; Maklakova, Vilena I; Berman, Samuel; De, Ishani; Steffen, Megan D; Hong, Won; Lincoln, Hayley; Morrissy, A Sorana; Taylor, Michael D; Akagi, Keiko; Brennan, Cameron W; Rodriguez, Fausto J; Collier, Lara S

    2014-01-01

    Genomic studies of human high-grade gliomas have discovered known and candidate tumor drivers. Studies in both cell culture and mouse models have complemented these approaches and have identified additional genes and processes important for gliomagenesis. Previously, we found that mobilization of Sleeping Beauty transposons in mice ubiquitously throughout the body from the Rosa26 locus led to gliomagenesis with low penetrance. Here we report the characterization of mice in which transposons are mobilized in the Glial Fibrillary Acidic Protein (GFAP) compartment. Glioma formation in these mice did not occur on an otherwise wild-type genetic background, but rare gliomas were observed when mobilization occurred in a p19Arf heterozygous background. Through cloning insertions from additional gliomas generated by transposon mobilization in the Rosa26 compartment, several candidate glioma genes were identified. Comparisons to genetic, epigenetic and mRNA expression data from human gliomas implicates several of these genes as tumor suppressor genes and oncogenes in human glioblastoma.

  16. Mouse models for genes involved in impaired spermatogenesis.

    PubMed

    O'Bryan, M K; de Kretser, D

    2006-02-01

    Since the introduction of molecular biology and gene ablation technologies there have been substantial advances in our understanding of how sperm are made and fertilization occurs. There have been at least 150 different models of specifically altered gene function produced that have resulted in male infertility spanning virtually all aspects of the spermatogenic, sperm maturation and fertilization processes. While each has, or potentially will reveal, novel aspects of these processes, there is still much of which we have little knowledge. The current review is by no means a comprehensive list of these mouse models, rather it gives an overview of the potential for such models which up to this point have generally been 'knockouts'; it presents alternative strategies for the production of new models and emphasizes the importance of thorough phenotypic analysis in order to extract a maximum amount of information from each model.

  17. Involvement of mouse and porcine PLCζ-induced calcium oscillations in preimplantation development of mouse embryos

    SciTech Connect

    Yoneda, Akihiro; Watanabe, Tomomasa

    2015-05-01

    In mammals, phospholipase Cζ (PLCζ) has the ability to trigger calcium (Ca{sup 2+}) oscillations in oocytes, leading to oocyte activation. Although there is a species-specific difference in the PLCζ-induced Ca{sup 2+} oscillatory pattern, whether PLCζ-induced Ca{sup 2+} oscillations affect preimplantation embryonic development remains unclear. Here, we show that Ca{sup 2+} oscillations in mouse PLCζ cRNA-injected oocytes stopped just before pronuclear formation, while that in porcine PLCζ cRNA-injected oocytes continued for several hours after pronuclei had been formed. This difference of Ca{sup 2+} oscillations in oocytes after pronuclear formation was dependent on the difference in the nuclear localization signal (NLS) sequence of PLCζ between the mouse and pig. However, mouse and porcine PLCζ cRNA-injected oocytes parthenogenetically developed to blastocysts regardless of the absence or presence of Ca{sup 2+} oscillations after pronuclear formation. Furthermore, the developmental rate of mouse or porcine PLCζ-activated oocytes injected with round spermatids to the blastocyst stage was not significantly different from that of strontium-activated oocytes injected with round spermatids. These results suggest that the PLCζ-induced Ca{sup 2+} oscillatory pattern in mouse oocytes is dependent on the NLS sequence of PLCζ and injection of PLCζ may be a useful method for activation of round spermatid-injected and somatic nuclear transferred oocytes. - Highlights: • Porcine PLCζ-induced Ca{sup 2+} oscillations continued after pronuclear formation. • The Ca{sup 2+} oscillatory pattern was dependent on the difference in the NLS sequence of PLCζ. • PLCζ-activated oocytes parthenogenetically developed to blastocysts. • PLCζ-activated oocytes injected with round spermatids developed to blastocysts.

  18. Sleeping Beauty Mouse Models Identify Candidate Genes Involved in Gliomagenesis

    PubMed Central

    Vyazunova, Irina; Maklakova, Vilena I.; Berman, Samuel; De, Ishani; Steffen, Megan D.; Hong, Won; Lincoln, Hayley; Morrissy, A. Sorana; Taylor, Michael D.; Akagi, Keiko; Brennan, Cameron W.; Rodriguez, Fausto J.; Collier, Lara S.

    2014-01-01

    Genomic studies of human high-grade gliomas have discovered known and candidate tumor drivers. Studies in both cell culture and mouse models have complemented these approaches and have identified additional genes and processes important for gliomagenesis. Previously, we found that mobilization of Sleeping Beauty transposons in mice ubiquitously throughout the body from the Rosa26 locus led to gliomagenesis with low penetrance. Here we report the characterization of mice in which transposons are mobilized in the Glial Fibrillary Acidic Protein (GFAP) compartment. Glioma formation in these mice did not occur on an otherwise wild-type genetic background, but rare gliomas were observed when mobilization occurred in a p19Arf heterozygous background. Through cloning insertions from additional gliomas generated by transposon mobilization in the Rosa26 compartment, several candidate glioma genes were identified. Comparisons to genetic, epigenetic and mRNA expression data from human gliomas implicates several of these genes as tumor suppressor genes and oncogenes in human glioblastoma. PMID:25423036

  19. Lentiviral vector-mediated transduction of goat undifferentiated spermatogonia.

    PubMed

    Abbasi, Hassan; Hosseini, Sayyed Morteza; Hajian, Mahdi; Nasiri, Zahra; Bahadorani, Mehrnoosh; Tahmoorespur, Mojtaba; Nasiri, Mohammad Reza; Nasr-Esfahani, Mohammad Hossein

    2015-12-01

    Recent studies show that spermatogonial stem cells (SSCs) are able to colonize and form mature spermatozoa following transplantation into germ cell depleted testes of recipient males. Therefore, efficient ways for enrichment and gene transfer into SSCs provides a powerful tool for production of transgenic animals. In order to adapt the technique to goats, three issues were addressed: (i) enrichment of the undifferentiated spermatogonia including SSCs using magnetic activated cell sorting (MACS), (ii) lentiviral vector-mediated transduction of an enhanced green fluorescent protein (EGFP) transgene into enriched cells, and (iii) transplantation of transduced undifferentiated spermatogonia into the germ cell depleted testes of immune-suppressed mice to assess for migration and colony formation ability. Enriched cells were transduced by lentiviral vectors and subsequently analyzed for expression of THY1, PLZF, VASA, UCHL1 and BCL6B genes. Cells were also analyzed for GFP and PLZF by flow cytometry. Enriched transduced cells were transplanted into germ cell depleted mice testis. Quantitative analysis of transcripts revealed that MACS-enrichment significantly increased the expression of SSC-characteristic genes THY1, PLZF, VASA, UCHL1 and BCL6B compared to non-enriched population (P≤0.05). EGFP transduction did not affect the expression levels of SSC-characteristic genes. Flow cytometry revealed that 72% of transduced-enriched cells were positive for EGFP. Finally, transduced-enriched goat SSCs could colonize within the cells into the seminiferous tubules of germ cell depleted recipient mice at higher frequency than non-enriched cells. The results indicated that enrichment of goat undifferentiated spermatogonia by magnetic-activated cell sorting for THY1 antibody combined with lentiviral vector-mediated transduction has the potential to be used for production of transgenic goats.

  20. Isolation of undifferentiated and early differentiating Type A spermatogonia from Pou5f1-GFP reporter mice

    PubMed Central

    Garcia, Thomas; Hofmann, Marie-Claude

    2012-01-01

    Limited understanding of the mechanisms underlying self-renewal and differentiation of spermatogonial stem cells hampers our ability to develop new therapeutic and contraceptive approaches. Mouse models of spermatogonial stem cell development are key to developing new insights into the biology of both the normal and diseased testis. Advances in transgenic reporter mice have enabled the isolation, molecular characterization, and functional analysis of mouse Type A spermatogonia subpopulations from the normal testis, including populations enriched for spermatogonial stem cells. Application of these reporters both to the normal testis and to gene-deficient and over-expression models will promote a better understanding of the earliest steps of spermatogenesis, and the role of spermatogonial stem cells in germ cell tumor. PMID:22144234

  1. Involvement of Transient Receptor Potential Vanilloid (TRPV) 4 in mouse sperm thermotaxis

    PubMed Central

    HAMANO, Koh-ichi; KAWANISHI, Tae; MIZUNO, Atsuko; SUZUKI, Makoto; TAKAGI, Yuji

    2016-01-01

    Transient Receptor Potential Vanilloid (TRPV) 4 is one of the temperature-sensitive ion channels involved in temperature receptors, and it is known to be activated from 35 to 40ºC. Here we analyzed sperm motility function of Trpv4 knockout (KO) mouse in temperature-gradient conditions to elucidate the thermotaxis of mouse sperm and the involvement of TRPV4 in thermotaxis. The sperm were introduced at the vertical column end of a T-shaped chamber filled with medium in a plastic dish, and we measured the number of sperm that arrived at both ends of the wide column where we had established a temperature gradient of approx. 2ºC, and we evaluated the sperm’s thermotaxis. Large numbers of wild-type (WT) mouse sperm migrated into the high level of the temperature gradient that was set in the wide column, and thermotaxis was confirmed. The ratio of migrated sperm at the high temperature level of the T-shaped chamber was decreased in the KO sperm and Ruthenium red (a TRPV antagonist) treated sperm compared with the WT sperm. The thermotaxis of the mouse sperm was confirmed, and the involvement of TRPV4 in this thermotaxis was suggested. PMID:27180924

  2. Involvement of Transient Receptor Potential Vanilloid (TRPV) 4 in mouse sperm thermotaxis.

    PubMed

    Hamano, Koh-Ichi; Kawanishi, Tae; Mizuno, Atsuko; Suzuki, Makoto; Takagi, Yuji

    2016-08-25

    Transient Receptor Potential Vanilloid (TRPV) 4 is one of the temperature-sensitive ion channels involved in temperature receptors, and it is known to be activated from 35 to 40ºC. Here we analyzed sperm motility function of Trpv4 knockout (KO) mouse in temperature-gradient conditions to elucidate the thermotaxis of mouse sperm and the involvement of TRPV4 in thermotaxis. The sperm were introduced at the vertical column end of a T-shaped chamber filled with medium in a plastic dish, and we measured the number of sperm that arrived at both ends of the wide column where we had established a temperature gradient of approx. 2ºC, and we evaluated the sperm's thermotaxis. Large numbers of wild-type (WT) mouse sperm migrated into the high level of the temperature gradient that was set in the wide column, and thermotaxis was confirmed. The ratio of migrated sperm at the high temperature level of the T-shaped chamber was decreased in the KO sperm and Ruthenium red (a TRPV antagonist) treated sperm compared with the WT sperm. The thermotaxis of the mouse sperm was confirmed, and the involvement of TRPV4 in this thermotaxis was suggested. PMID:27180924

  3. Evidences for the agmatine involvement in antidepressant like effect of bupropion in mouse forced swim test.

    PubMed

    Kotagale, Nandkishor R; Tripathi, Sunil J; Aglawe, Manish M; Chopde, Chandrabhan T; Umekar, Milind J; Taksande, Brijesh G

    2013-06-01

    Although bupropion has been widely used in the treatment of depression, the precise mechanism of its therapeutic actions is not fully understood. The present study investigated the role of agmatine in an antidepressant like effect of bupropion in mouse forced swim test. The antidepressant like effect of bupropion was potentiated by pretreatment with agmatine (10-20mg/kg, ip) and by the drugs known to increase endogenous agmatine levels in brain viz., l-arginine (40 μg/mouse, icv), an agmatine biosynthetic precursor, ornithine decarboxylase inhibitor, dl-α-difluoromethyl ornithine hydrochloride, DFMO (12.5 μg/mouse, icv), diamine oxidase inhibitor, aminoguanidine (6.5 μg/mouse, icv) and agmatinase inhibitor, arcaine (50 μg/mouse, icv) as well as imidazoline I1 receptor agonists, moxonidine (0.25mg/kg, ip) and clonidine (0.015 mg/kg, ip) and imidazoline I2 receptor agonist, 2-(2-benzofuranyl)-2-imidazoline hydrochloride, 2-BFI (5mg/kg, ip). Conversely, prior administration of I1 receptor antagonist, efaroxan (1mg/kg, ip) and I2 receptor antagonist, idazoxan (0.25mg/kg, ip) blocked the antidepressant like effect of bupropion and its synergistic combination with agmatine. These results demonstrate involvement of agmatine in the antidepressant like effect of bupropion and suggest agmatine and imidazoline receptors as a potential therapeutic target for the treatment of depressive disorders.

  4. Mouse neuron navigator 1, a novel microtubule-associated protein involved in neuronal migration.

    PubMed

    Martínez-López, María José; Alcántara, Soledad; Mascaró, Cristina; Pérez-Brangulí, Francesc; Ruiz-Lozano, Pilar; Maes, Tamara; Soriano, Eduardo; Buesa, Carlos

    2005-04-01

    The development of the nervous system (NS) requires the coordinated migration of multiple waves of neurons and subsequent processes of neurite maturation, both involving selective guidance mechanisms. In Caenorhabditis elegans, unc-53 codes for a new multidomain protein involved in the directional migration of a subset of cells. We describe here the first functional characterization of the mouse homologue, mouse Neuron navigator 1 (mNAV1), whose expression is largely restricted to the NS during development. EGFP-mNAV1 associates with microtubules (MTs) plus ends present in the growth cone through a new microtubule-binding (MTB) domain. Moreover, its overexpression in transfected cells leads to MT bundling. The abolition of mNAV1 causes loss of directionality in the leading processes of pontine-migrating cells, providing evidence for a role of mNAV1 in mediating Netrin-1-induced directional migration. PMID:15797708

  5. Axial elongation in mouse embryos involves mediolateral cell intercalation behavior in the paraxial mesoderm

    NASA Astrophysics Data System (ADS)

    Yen, WeiWei; Burdsal, Carol; Periasamy, Ammasi; Sutherland, Ann E.

    2006-02-01

    The cell mechanical and signaling pathways involved in gastrulation have been studied extensively in invertebrates and amphibians, such as Xenopus, and more recently in non-mammalian vertebrates such as zebrafish and chick. However, because culturing mouse embryos extra-utero is very difficult, this fundamental process has been least characterized in the mouse. As the primary mammalian model for genetics, biochemistry, and the study of human disease and birth defects, it is important to investigate how gastrulation proceeds in murine embryos. We have developed a method of using 4D multiphoton excitation microscopy and extra-utero culture to visualize and characterize the morphogenetic movements in mouse embryos dissected at 8.5 days of gestation. Cells are labeled by expression of an X chromosome-linked enhanced green fluorescent protein (EGFP) transgene. This method has provided a unique approach, where, for the first time, patterns of cell behavior in the notochord and surrounding paraxial mesoderm can be visualized and traced quantitatively. Our observations of mouse embryos reveal both distinct differences as well as striking similarities in patterned cell motility relative to other vertebrate models such as Xenopus, where axial extension is driven primarily by mediolateral oriented cell behaviors in the notochord and paraxial somitic mesoderm. Unlike Xenopus, the width of the mouse notochord remains the same between 4-somite stage and 8-somite stage embryos. This implies the mouse notochord plays a lesser role in driving axial extension compared to Xenopus, although intercalation may occur where the anterior region of the node becomes notochordal plate. In contrast, the width of mouse paraxial mesoderm narrows significantly during this period and cells within the paraxial mesoderm are both elongated and aligned perpendicular to the midline. In addition, these cells are observed to intercalate, consistent with a role for paraxial mesoderm in driving convergence

  6. Molecular Mechanisms Elicited by d-Aspartate in Leydig Cells and Spermatogonia

    PubMed Central

    Di Fiore, Maria Maddalena; Santillo, Alessandra; Falvo, Sara; Longobardi, Salvatore; Chieffi Baccari, Gabriella

    2016-01-01

    A bulk of evidence suggests that d-aspartate (d-Asp) regulates steroidogenesis and spermatogenesis in vertebrate testes. This review article focuses on intracellular signaling mechanisms elicited by d-Asp possibly via binding to the N-methyl-d-aspartate receptor (NMDAR) in both Leydig cells, and spermatogonia. In Leydig cells, the amino acid upregulates androgen production by eliciting the adenylate cyclase-cAMP and/or mitogen-activated protein kinase (MAPK) pathways. d-Asp treatment enhances gene and protein expression of enzymes involved in the steroidogenic cascade. d-Asp also directly affects spermatogonial mitotic activity. In spermatogonial GC-1 cells, d-Asp induces phosphorylation of MAPK and AKT serine-threonine kinase proteins, and stimulates expression of proliferating cell nuclear antigen (PCNA) and aurora kinase B (AURKB). Further stimulation of spermatogonial GC-1 cell proliferation might come from estradiol/estrogen receptor β (ESR2) interaction. d-Asp modulates androgen and estrogen levels as well as the expression of their receptors in the rat epididymis by acting on mRNA levels of Srd5a1 and Cyp19a1 enzymes, hence suggesting involvement in spermatozoa maturation. PMID:27428949

  7. The spatiotemporal expression and localization implicates a potential role for SerpinB11 in the process of mouse spermatogenesis and apoptosis.

    PubMed

    Yang, Yanzhou; Ma, Wenzhi; Ma, Huiming; Sun, Miao; Chang, Qing; Pei, Xiuying; Wang, Yanrong

    2015-01-01

    In this study, the spatiotemporal expression of SerpinB11 in the mouse testis from postnatal 1-60 d was checked, the SerpinB11 protein strongly localized in the intermediate spermatogonia, B-type spermatogonium, preleptotene spermatocyte, leptonema spermatocyte, zygotene spermatocyte, but weakly localized in the pachytene spermatocyte, diplotene spermatocyte, sphere sperm, and the apoptotic sperm was positive stained of SerpinB11 protein, the localization of cell cycle marker CDK4 and meiosis marker SCP3 were investigated, and the SCP3 and SerpinB11 colocalized in the intermediate spermatogonia, B-type spermatogonium, preleptotene spermatocyte. Taken together, these results suggested that SerpinB11 might involved in spermatogenesis and apoptosis. PMID:24785531

  8. A-Single Spermatogonia Heterogeneity and Cell Cycles Synchronize with Rat Seminiferous Epithelium Stages VIII–IX1

    PubMed Central

    Abid, Shadaan N.; Richardson, Timothy E.; Powell, Heather M.; Jaichander, Priscilla; Chaudhary, Jaideep; Chapman, Karen M.; Hamra, F. Kent

    2014-01-01

    ABSTRACT In mammalian testes, “A-single” spermatogonia function as stem cells that sustain sperm production for fertilizing eggs. Yet, it is not understood how cellular niches regulate the developmental fate of A-single spermatogonia. Here, immunolabeling studies in rat testes define a novel population of ERBB3+ germ cells as approximately 5% of total SNAP91+ A-single spermatogonia along a spermatogenic wave. As a function of time, ERBB3+ A-single spermatogonia are detected during a 1- to 2-day period each 12.9-day sperm cycle, representing 35%–40% of SNAP91+ A-single spermatogonia in stages VIII–IX of the seminiferous epithelium. Local concentrations of ERBB3+ A-single spermatogonia are maintained under the mean density measured for neighboring SNAP91+ A-single spermatogonia, potentially indicative of niche saturation. ERBB3+ spermatogonia also synchronize their cell cycles with epithelium stages VIII–IX, where they form physical associations with preleptotene spermatocytes transiting the blood-testis barrier and Sertoli cells undergoing sperm release. Thus, A-single spermatogonia heterogeneity within this short-lived and reoccurring microenvironment invokes novel theories on how cellular niches integrate with testicular physiology to orchestrate sperm development in mammals. PMID:24389876

  9. AZFc deletions do not affect the function of human spermatogonia in vitro

    PubMed Central

    Nickkholgh, B.; Korver, C.M.; van Daalen, S.K.M.; van Pelt, A.M.M.; Repping, S.

    2015-01-01

    Azoospermic factor c (AZFc) deletions are the underlying cause in 10% of azoo- or severe oligozoospermia. Through extensive molecular analysis the precise genetic content of the AZFc region and the origin of its deletion have been determined. However, little is known about the effect of AZFc deletions on the functionality of germ cells at various developmental steps. The presence of normal, fertilization-competent sperm in the ejaculate and/or testis of the majority of men with AZFc deletions suggests that the process of differentiation from spermatogonial stem cells (SSCs) to mature spermatozoa can take place in the absence of the AZFc region. To determine the functionality of AZFc-deleted spermatogonia, we compared in vitro propagated spermatogonia from six men with complete AZFc deletions with spermatogonia from three normozoospermic controls. We found that spermatogonia of AZFc-deleted men behave similar to controls during culture. Short-term (18 days) and long-term (48 days) culture of AZFc-deleted spermatogonia showed the same characteristics as non-deleted spermatogonia. This similarity was revealed by the same number of passages, the same germ cell clusters formation and similar level of genes expression of spermatogonial markers including ubiquitin carboxyl-terminal esterase L1 (UCHL1), zinc finger and BTB domain containing 16 (ZBTB16) and glial cell line-derived neurotrophic factor family receptor alpha 1 (GFRA1), as well as germ cell differentiation markers including signal transducer and activator of transcription 3 (STAT3), spermatogenesis and oogenesis specific basic helix-loophelix 2 (SOHLH2), v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT) and synaptonemal complex protein 3 (SYCP3). The only exception was melanoma antigen family A4 (MAGEA4) which showed significantly lower expression in AZFc-deleted samples than controls in short-term culture while in long-term culture it was hardly detected in both AZFc-deleted and control

  10. Extraordinary sequence divergence at Tsga8, an X-linked gene involved in mouse spermiogenesis.

    PubMed

    Good, Jeffrey M; Vanderpool, Dan; Smith, Kimberly L; Nachman, Michael W

    2011-05-01

    The X chromosome plays an important role in both adaptive evolution and speciation. We used a molecular evolutionary screen of X-linked genes potentially involved in reproductive isolation in mice to identify putative targets of recurrent positive selection. We then sequenced five very rapidly evolving genes within and between several closely related species of mice in the genus Mus. All five genes were involved in male reproduction and four of the genes showed evidence of recurrent positive selection. The most remarkable evolutionary patterns were found at Testis-specific gene a8 (Tsga8), a spermatogenesis-specific gene expressed during postmeiotic chromatin condensation and nuclear transformation. Tsga8 was characterized by extremely high levels of insertion-deletion variation of an alanine-rich repetitive motif in natural populations of Mus domesticus and M. musculus, differing in length from the reference mouse genome by up to 89 amino acids (27% of the total protein length). This population-level variation was coupled with striking divergence in protein sequence and length between closely related mouse species. Although no clear orthologs had previously been described for Tsga8 in other mammalian species, we have identified a highly divergent hypothetical gene on the rat X chromosome that shares clear orthology with the 5' and 3' ends of Tsga8. Further inspection of this ortholog verified that it is expressed in rat testis and shares remarkable similarity with mouse Tsga8 across several general features of the protein sequence despite no conservation of nucleotide sequence across over 60% of the rat-coding domain. Overall, Tsga8 appears to be one of the most rapidly evolving genes to have been described in rodents. We discuss the potential evolutionary causes and functional implications of this extraordinary divergence and the possible contribution of Tsga8 and the other four genes we examined to reproductive isolation in mice. PMID:21186189

  11. Differential gene expression profiling of enriched human spermatogonia after short- and long-term culture.

    PubMed

    Conrad, Sabine; Azizi, Hossein; Hatami, Maryam; Kubista, Mikael; Bonin, Michael; Hennenlotter, Jörg; Renninger, Markus; Skutella, Thomas

    2014-01-01

    This study aimed to provide a molecular signature for enriched adult human stem/progenitor spermatogonia during short-term (<2 weeks) and long-term culture (up to more than 14 months) in comparison to human testicular fibroblasts and human embryonic stem cells. Human spermatogonia were isolated by CD49f magnetic activated cell sorting and collagen(-)/laminin(+) matrix binding from primary testis cultures obtained from ten adult men. For transcriptomic analysis, single spermatogonia-like cells were collected based on their morphology and dimensions using a micromanipulation system from the enriched germ cell cultures. Immunocytochemical, RT-PCR and microarray analyses revealed that the analyzed populations of cells were distinct at the molecular level. The germ- and pluripotency-associated genes and genes of differentiation/spermatogenesis pathway were highly expressed in enriched short-term cultured spermatogonia. After long-term culture, a proportion of cells retained and aggravated the "spermatogonial" gene expression profile with the expression of germ and pluripotency-associated genes, while in the majority of long-term cultured cells this molecular profile, typical for the differentiation pathway, was reduced and more genes related to the extracellular matrix production and attachment were expressed. The approach we provide here to study the molecular status of in vitro cultured spermatogonia may be important to optimize the culture conditions and to evaluate the germ cell plasticity in the future.

  12. Involvement of Autophagic Pathway in the Progression of Retinal Degeneration in a Mouse Model of Diabetes.

    PubMed

    Piano, Ilaria; Novelli, Elena; Della Santina, Luca; Strettoi, Enrica; Cervetto, Luigi; Gargini, Claudia

    2016-01-01

    The notion that diabetic retinopathy (DR) is essentially a micro-vascular disease has been recently challenged by studies reporting that vascular changes are preceded by signs of damage and loss of retinal neurons. As to the mode by which neuronal death occurs, the evidence that apoptosis is the main cause of neuronal loss is far from compelling. The objective of this study was to investigate these controversies in a mouse model of streptozotocin (STZ) induced diabetes. Starting from 8 weeks after diabetes induction there was loss of rod but not of cone photoreceptors, together with reduced thickness of the outer and inner synaptic layers. Correspondingly, rhodopsin expression was downregulated and the scotopic electroretinogram (ERG) is suppressed. In contrast, cone opsin expression and photopic ERG response were not affected. Suppression of the scotopic ERG preceded morphological changes as well as any detectable sign of vascular alteration. Only sparse apoptotic figures were detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and glia was not activated. The physiological autophagy flow was altered instead, as seen by increased LC3 immunostaining at the level of outer plexiform layer (OPL) and upregulation of the autophagic proteins Beclin-1 and Atg5. Collectively, our results show that the streptozotocin induced DR in mouse initiates with a functional loss of the rod visual pathway. The pathogenic pathways leading to cell death develop with the initial dysregulation of autophagy well before the appearance of signs of vascular damage and without strong involvement of apoptosis.

  13. Proteins involved in motility and sperm-egg interaction evolve more rapidly in mouse spermatozoa.

    PubMed

    Vicens, Alberto; Lüke, Lena; Roldan, Eduardo R S

    2014-01-01

    Proteomic studies of spermatozoa have identified a large catalog of integral sperm proteins. Rapid evolution of these proteins may underlie adaptive changes of sperm traits involved in different events leading to fertilization, although the selective forces underlying such rapid evolution are not well understood. A variety of selective forces may differentially affect several steps ending in fertilization, thus resulting in a compartmentalized adaptation of sperm proteins. Here we analyzed the evolution of genes associated to various events in the sperm's life, from sperm formation to sperm-egg interaction. Evolutionary analyses were performed on gene sequences from 17 mouse strains whose genomes have been sequenced. Four of these are derived from wild Mus musculus, M. domesticus, M. castaneus and M. spretus. We found a higher proportion of genes exhibiting a signature of positive selection among those related to sperm motility and sperm-egg interaction. Furthermore, sperm proteins involved in sperm-egg interaction exhibited accelerated evolution in comparison to those involved in other events. Thus, we identified a large set of candidate proteins for future comparative analyses of genotype-phenotype associations in spermatozoa of species subjected to different sexual selection pressures. Adaptive evolution of proteins involved in motility could be driven by sperm competition, since this selective force is known to increase the proportion of motile sperm and their swimming velocity. On the other hand, sperm proteins involved in gamete interaction could be coevolving with their egg partners through episodes of sexual selection or sexual conflict resulting in species-specific sperm-egg interactions and barriers preventing interspecies fertilization.

  14. On the disentangling modes of ortho- and para-braids in mitosing mammalian spermatogonia.

    PubMed

    Matioli, G T

    1994-10-01

    This note complements a previous one (1), regarding in particular, mitotic separation of replica homologs from their templates, in mammalian spermatogonia. Topological differences between Ortho- and Para-braids determine different modes of antipodal segregation of paired homologs prior to onset of meiosis.

  15. High telomerase is a hallmark of undifferentiated spermatogonia and is required for maintenance of male germline stem cells

    PubMed Central

    Pech, Matthew F.; Garbuzov, Alina; Hasegawa, Kazuteru; Sukhwani, Meena; Zhang, Ruixuan J.; Benayoun, Bérénice A.; Brockman, Stephanie A.; Lin, Shengda; Brunet, Anne; Orwig, Kyle E.; Artandi, Steven E.

    2015-01-01

    Telomerase inactivation causes loss of the male germline in worms, fish, and mice, indicating a conserved dependence on telomere maintenance in this cell lineage. Here, using telomerase reverse transcriptase (Tert) reporter mice, we found that very high telomerase expression is a hallmark of undifferentiated spermatogonia, the mitotic population where germline stem cells reside. We exploited these high telomerase levels as a basis for purifying undifferentiated spermatogonia using fluorescence-activated cell sorting. Telomerase levels in undifferentiated spermatogonia and embryonic stem cells are comparable and much greater than in somatic progenitor compartments. Within the germline, we uncovered an unanticipated gradient of telomerase activity that also enables isolation of more mature populations. Transcriptomic comparisons of TertHigh undifferentiated spermatogonia and TertLow differentiated spermatogonia by RNA sequencing reveals marked differences in cell cycle and key molecular features of each compartment. Transplantation studies show that germline stem cell activity is confined to the TertHigh cKit− population. Telomere shortening in telomerase knockout strains causes depletion of undifferentiated spermatogonia and eventual loss of all germ cells after undifferentiated spermatogonia drop below a critical threshold. These data reveal that high telomerase expression is a fundamental characteristic of germline stem cells, thus explaining the broad dependence on telomerase for germline immortality in metazoans. PMID:26584619

  16. Mechanisms involved in spinal cord central synapse loss in a mouse model of spinal muscular atrophy.

    PubMed

    Tarabal, Olga; Caraballo-Miralles, Víctor; Cardona-Rossinyol, Andrea; Correa, Francisco J; Olmos, Gabriel; Lladó, Jerònia; Esquerda, Josep E; Calderó, Jordi

    2014-06-01

    Motoneuron (MN) cell death is the histopathologic hallmark of spinal muscular atrophy (SMA), although MN loss seems to be a late event. Conversely, disruption of afferent synapses on MNs has been shown to occur early in SMA. Using a mouse model of severe SMA (SMNΔ7), we examined the mechanisms involved in impairment of central synapses. We found that MNs underwent progressive degeneration in the course of SMA, with MN loss still occurring at late stages. Loss of afferent inputs to SMA MNs was detected at embryonic stages, long before MN death. Reactive microgliosis and astrogliosis were present in the spinal cord of diseased animals after the onset of MN loss. Ultrastructural observations indicate that dendrites and microglia phagocytose adjacent degenerating presynaptic terminals. Neuronal nitric oxide synthase was upregulated in SMNΔ7 MNs, and there was an increase in phosphorylated myosin light chain expression in synaptic afferents on MNs; these observations implicate nitric oxide in MN deafferentation and suggest that the RhoA/ROCK pathway is activated. Together, our observations suggest that the earliest change occurring in SMNΔ7 mice is the loss of excitatory glutamatergic synaptic inputs to MNs; reduced excitability may enhance their vulnerability to degeneration and death.

  17. Prion Infection of Mouse Brain Reveals Multiple New Upregulated Genes Involved in Neuroinflammation or Signal Transduction

    PubMed Central

    Striebel, James F.; Race, Brent; Phillips, Katie; Chesebro, Bruce

    2014-01-01

    ABSTRACT Gliosis is often a preclinical pathological finding in neurodegenerative diseases, including prion diseases, but the mechanisms facilitating gliosis and neuronal damage in these diseases are not understood. To expand our knowledge of the neuroinflammatory response in prion diseases, we assessed the expression of key genes and proteins involved in the inflammatory response and signal transduction in mouse brain at various times after scrapie infection. In brains of scrapie-infected mice at pre- and postclinical stages, we identified 15 previously unreported differentially expressed genes related to inflammation or activation of the STAT signal transduction pathway. Levels for the majority of differentially expressed genes increased with time postinfection. In quantitative immunoblotting experiments of STAT proteins, STAT1α, phosphorylated-STAT1α (pSTAT1α), and pSTAT3 were increased between 94 and 131 days postinfection (p.i.) in brains of mice infected with strain 22L. Furthermore, a select group of STAT-associated genes was increased preclinically during scrapie infection, suggesting early activation of the STAT signal transduction pathway. Comparison of inflammatory markers between mice infected with scrapie strains 22L and RML indicated that the inflammatory responses and gene expression profiles in the brains were strikingly similar, even though these scrapie strains infect different brain regions. The endogenous interleukin-1 receptor antagonist (IL-1Ra), an inflammatory marker, was newly identified as increasing preclinically in our model and therefore might influence scrapie pathogenesis in vivo. However, in IL-1Ra-deficient or overexpressor transgenic mice inoculated with scrapie, neither loss nor overexpression of IL-1Ra demonstrated any observable effect on gliosis, protease-resistant prion protein (PrPres) formation, disease tempo, pathology, or expression of the inflammatory genes analyzed. IMPORTANCE Prion infection leads to Pr

  18. L1 adhesion molecule on mouse leukocytes: regulation and involvement in endothelial cell binding.

    PubMed

    Hubbe, M; Kowitz, A; Schirrmacher, V; Schachner, M; Altevogt, P

    1993-11-01

    L1 is a cell surface glycoprotein of the immunoglobulin superfamily which was initially shown to mediate adhesion between neural cells. Recently we have reported that L1 is expressed by bone marrow cells and the majority of mature lymphocytes (Kowitz et al., Eur. J. Immunol. 1992. 22: 1199-1205). To analyze the function of L1 on leukocytes we studied its regulation following cell activation. In vitro activation of B lymphocytes with lipopolysaccharide or T lymphocytes with phorbol 12-myristate 13-acetate/Ca2+ ionophore, concanavalin A or anti-CD3 monoclonal antibody as well as in vivo activation of V beta 8+ T cells with staphylococcal enterotoxin B (SEB) revealed a down-regulation of L1 within 48 h. A rapid loss of L1 expression was seen when mouse neutrophils were activated with PMA alone. This rapid loss paralleled the shedding of L-selectin. We also studied a possible role of L1 in the binding of leukocytes to endothelial cells. ESb-MP lymphoma cells with a high expression of L1 (L1hi) could bind to bend3 endothelioma cells without prior activation with inflammatory cytokines. The interaction was inhibited by anti-L1 antibodies. In contrast, ESb-MP cells with low L1 expression (L1lo) were only marginally bound. Latex beads coated with affinity-isolated L1 antigen were also able to bind to the endothelioma cells in a specific fashion. The binding of ESb-MP lymphoma cells required Ca2+ and Mg2+ ions and was sensitive to cold temperature. Since the endothelioma cells did not express L1 the binding mechanism studied here is distinct from the established L1-L1 homotypic interaction. It is possible that the novel L1-mediated adhesion pathway involves an unidentified ligand and could play a role in leukocyte migration.

  19. L1 adhesion molecule on mouse leukocytes: regulation and involvement in endothelial cell binding.

    PubMed

    Hubbe, M; Kowitz, A; Schirrmacher, V; Schachner, M; Altevogt, P

    1993-11-01

    L1 is a cell surface glycoprotein of the immunoglobulin superfamily which was initially shown to mediate adhesion between neural cells. Recently we have reported that L1 is expressed by bone marrow cells and the majority of mature lymphocytes (Kowitz et al., Eur. J. Immunol. 1992. 22: 1199-1205). To analyze the function of L1 on leukocytes we studied its regulation following cell activation. In vitro activation of B lymphocytes with lipopolysaccharide or T lymphocytes with phorbol 12-myristate 13-acetate/Ca2+ ionophore, concanavalin A or anti-CD3 monoclonal antibody as well as in vivo activation of V beta 8+ T cells with staphylococcal enterotoxin B (SEB) revealed a down-regulation of L1 within 48 h. A rapid loss of L1 expression was seen when mouse neutrophils were activated with PMA alone. This rapid loss paralleled the shedding of L-selectin. We also studied a possible role of L1 in the binding of leukocytes to endothelial cells. ESb-MP lymphoma cells with a high expression of L1 (L1hi) could bind to bend3 endothelioma cells without prior activation with inflammatory cytokines. The interaction was inhibited by anti-L1 antibodies. In contrast, ESb-MP cells with low L1 expression (L1lo) were only marginally bound. Latex beads coated with affinity-isolated L1 antigen were also able to bind to the endothelioma cells in a specific fashion. The binding of ESb-MP lymphoma cells required Ca2+ and Mg2+ ions and was sensitive to cold temperature. Since the endothelioma cells did not express L1 the binding mechanism studied here is distinct from the established L1-L1 homotypic interaction. It is possible that the novel L1-mediated adhesion pathway involves an unidentified ligand and could play a role in leukocyte migration. PMID:8223869

  20. atRA-induced apoptosis of mouse embryonic palate mesenchymal cells involves activation of MAPK pathway

    SciTech Connect

    Yu Zengli . E-mail: yuzengli@263.net; Xing Ying . E-mail: xingy@zzu.edu.cn

    2006-08-15

    Our previous studies have shown that atRA treatment resulted in cell-cycle block and growth inhibition in mouse embryonic palatal mesenchymal (MEPM). In the current study, gestation day (GD) 13 MEPM cells were used to test the hypothesis that the growth inhibition by atRA is due to apoptosis. The effects of atRA on apoptosis were assessed by performing MTT assay, Cell Death Detection ELISA and flow cytometry, respectively. Data analysis confirmed that atRA treatment induced apoptosis-like cell death, as shown by decreased cell viability and increased fragmented DNA and sub-G1 fraction. atRA-induced apoptosis was associated with upregulation of bcl-2, translocation of bax protein to the mitochondria from the cytosol, activation of caspase-3 and cytochrome c release into cytosol. atRA-induced apoptosis was abrogated by z-DEVD-fmk, a caspase-3 specific inhibitor, and z-VAD-fmk, a general caspase inhibitor, suggesting that the atRA-induced cell death of MEPM cells occurs through the cytochrome c- and caspase-3-dependent pathways. In addition, atRA treatment caused a strong and sustained activation of c-Jun N-terminal kinase (JNK) and p38 kinase (p38), as well as an early but transient activation of extracellular signal-regulated kinase (ERK). Importantly, atRA-induced DNA fragmentation and capase-3 activation were prevented by pretreatment with the JNK inhibitor (SP600125) and the p38 MAPK inhibitor (SB202190), but not by pretreatment with MEK inhibitor (U0126). From these results, we suggest that mitogen-activated protein kinase-dependent pathways is involved in the atRA-induced apoptosis of MEPM cells.

  1. Involvement of catecholaminergic neurons in motor innervation of striated muscle in the mouse esophagus.

    PubMed

    van der Keylen, Piet; Garreis, Fabian; Steigleder, Ruth; Sommer, Daniel; Neuhuber, Winfried L; Wörl, Jürgen

    2016-05-01

    Enteric co-innervation is a peculiar innervation pattern of striated esophageal musculature. Both anatomical and functional data on enteric co-innervation related to various transmitters have been collected in different species, although its function remains enigmatic. However, it is unclear whether catecholaminergic components are involved in such a co-innervation. Thus, we examined to identify catecholaminergic neuronal elements and clarify their relationship to other innervation components in the esophagus, using immunohistochemistry with antibodies against tyrosine hydroxylase (TH), vesicular acetylcholine transporter (VAChT), choline acetyltransferase (ChAT) and protein gene product 9.5 (PGP 9.5), α-bungarotoxin (α-BT) and PCR with primers for amplification of cDNA encoding TH and dopamine-β-hydroxylase (DBH). TH-positive nerve fibers were abundant throughout the myenteric plexus and localized on about 14% of α-BT-labelled motor endplates differing from VAChT-positive vagal nerve terminals. TH-positive perikarya represented a subpopulation of only about 2.8% of all PGP 9.5-positive myenteric neurons. Analysis of mRNA showed both TH and DBH transcripts in the mouse esophagus. As ChAT-positive neurons in the compact formation of the nucleus ambiguus were negative for TH, the TH-positive nerve varicosities on motor endplates are presumably of enteric origin, although a sympathetic origin cannot be excluded. In the medulla oblongata, the cholinergic ambiguus neurons were densely supplied with TH-positive varicosities. Thus, catecholamines may modulate vagal motor innervation of esophageal-striated muscles not only at the peripheral level via enteric co-innervation but also at the central level via projections to the nucleus ambiguus. As Parkinson's disease, with a loss of central dopaminergic neurons, also affects the enteric nervous system and dysphagia is prevalent in patients with this disease, investigation of intrinsic catecholamines in the esophagus may

  2. Retinoic Acid Receptors Control Spermatogonia Cell-Fate and Induce Expression of the SALL4A Transcription Factor

    PubMed Central

    Gely-Pernot, Aurore; Raverdeau, Mathilde; Teletin, Marius; Vernet, Nadège; Féret, Betty; Klopfenstein, Muriel; Dennefeld, Christine; Davidson, Irwin; Benoit, Gérard; Mark, Manuel; Ghyselinck, Norbert B.

    2015-01-01

    All-trans retinoic acid (ATRA) is instrumental to male germ cell differentiation, but its mechanism of action remains elusive. To address this question, we have analyzed the phenotypes of mice lacking, in spermatogonia, all rexinoid receptors (RXRA, RXRB and RXRG) or all ATRA receptors (RARA, RARB and RARG). We demonstrate that the combined ablation of RXRA and RXRB in spermatogonia recapitulates the set of defects observed both upon ablation of RAR in spermatogonia. We also show that ATRA activates RAR and RXR bound to a conserved regulatory region to increase expression of the SALL4A transcription factor in spermatogonia. Our results reveal that this major pluripotency gene is a target of ATRA signaling and that RAR/RXR heterodimers are the functional units driving its expression in spermatogonia. They add to the mechanisms through which ATRA promote expression of the KIT tyrosine kinase receptor to trigger a critical step in spermatogonia differentiation. Importantly, they indicate also that meiosis eventually occurs in the absence of a RAR/RXR pathway within germ cells and suggest that instructing this process is either ATRA-independent or requires an ATRA signal originating from Sertoli cells. PMID:26427057

  3. Cytotoxic effects of WR-2721 on mouse testicular cells

    SciTech Connect

    Meistrich, M.L.; Finch, M.V.; Hunter, N.; Milas, L.

    1984-09-01

    WR-2721 (S-2-(3-aminopropylamino)ethylphosphorothioic acid) has been demonstrated to be cytotoxic to stem spermatogonia in the mouse. Five and 10 injections of 300 mg/kg killed sufficient numbers of stem cells to reduce sperm production 56 days after treatment by 16 and 43%, respectively. Single injections of 300 or 400 mg/kg of WR-2721 given 15 min after irradiation produced negligible toxicity to stem cells as measured by counts of repopulated tubules; 600 mg/kg reduced stem cell survival by 47%. Four daily injections of 300 mg/kg given 4, 3, 2, and 1 days prior to irradiation reduced stem cell survival by about 60%. The cytotoxic effects of WR-2721 on testicular stem cells at least partially explains the reduced protection factors observed in the testis with low doses of radiation and during fractionated treatments involving multiple injections of drug.

  4. Comparison of the involvement protein kinase C in agonist-induced contractions in mouse aorta and corpus cavernosum

    PubMed Central

    Jin, Liming; Teixeira, Cleber E; Webb, R. Clinton; Leite, Romulo

    2008-01-01

    Protein kinase C (PKC) is involved in the regulation of vascular smooth muscle contraction. However, the role of PKC in erectile function is poorly understood. This study investigated whether PKC mediates agonist-induced contractions in mouse penile tissue (corpora cavernosa). We also compared the effects of PKC activators and inhibitors on contractile responses in mouse corpus cavernosum with those in mouse aorta. Aortic rings and corpus cavernosal strips from C57BL/6J mice was isolated, mounted in the organ bath for isometric tension recording. Our data showed that a PKCα/β selective inhibitor, Gö6976 (10 µM), inhibited phenylephrine and 9,11-dideoxy-11α,9α-epoxymethanoprostaglandin F2α (U46619, a thromboxane mimetic)-induced contractions in mouse aorta, reducing the maximum contraction from 123 ± 2% of KCl-induced maximum contraction to 7 ± 2% and 13 ± 1%, respectively. A non-selective PKC inhibitor, chelerythrine (30 µM), also significantly reduced phenylephrine-and U46619-induced maximum contractions in mouse aorta. However, Gö6976 and chelerythrine had no significant effects on phenylephrine-and U46619-induced contractions in corpus cavernosum. Furthermore, a PKC activator, phorbol-12,13-dibutyrate (0.1 µM), significantly increased contractions in aorta (208 ± 14% of KCl-induced maximum contraction) but failed to cause contractions in corpus cavernosum at 1 and 10 µM. Western blot analysis data suggested that protein expression of PKC was similar in aorta and corpus cavernosum. Taken together, our data indicate that PKC does not have a significant role in agonist-induced contractions in mouse corpus cavernosum, whereas it mediates the contractile response to agonists in the aorta. PMID:18614166

  5. Metabolic activation of chrysene in mouse skin: evidence for the involvement of a triol-epoxide.

    PubMed

    Hodgson, R M; Weston, A; Grover, P L

    1983-12-01

    All three possible dihydrodiols of chrysene and a chrysene triol, formed from the further metabolism of the chrysene-1,2-diol, were detected when ether extracts of mouse skin that had been treated with 3H-labelled chrysene were examined by h.p.l.c. The major deoxyribonucleoside-hydrocarbon adducts present in hydrolysates of DNA isolated from the mouse skin were examined by chromatography on Sephadex LH20 and by h.p.l.c. on Zorbax ODS. One adduct had chromatographic properties identical to those of the major adduct formed when r-1,t-2-dihydroxy-t-3,4-oxy-1,2,3,4-tetrahydrochrysene reacts with DNA. A second major adduct was present that had chromatographic properties that were indistinguishable from those of an adduct that was formed when either chrysene-1,2-diol or 3-hydroxychrysene were incubated with DNA in a rat liver microsomal metabolising system. The results provide evidence that this new adduct is formed via the reaction of a 'triol-epoxide', that appears to be 9-hydroxy-chrysene-1,2-diol 3,4-oxide, with DNA in mouse skin.

  6. Disruption of PPARγ signaling results in mouse prostatic intraepithelial neoplasia involving active autophagy

    PubMed Central

    Jiang, Ming; Fernandez, Suzanne; Jerome, W. Gray; He, Yue; Yu, Xiuping; Cai, Hui; Boone, Braden; Yi, Yajun; Magnuson, Mark A.; Roy-Burman, Pradip; Matusik, Robert J.; Shappell, Scott B.; Hayward, Simon W.

    2009-01-01

    Peroxisome proliferator-activated receptor-gamma (PPARγ) regulates the interface between cellular lipid metabolism, redox status and organelle differentiation. Conditional prostatic epithelial knockout of PPARγ in mice resulted in focal hyperplasia which developed into mouse prostatic intraepithelial neoplasia (mPIN). The grade of PIN became more severe with time. Electron microscopy (EM) showed accumulated secondary lysosomes containing cellular organelles and debris suggestive of autophagy. Consistent with this analysis the autophagy marker LC3 was found to be upregulated in areas of PIN in PPARγ KO tissues. We selectively knocked down PPARγ2 isoform in wild-type mouse prostatic epithelial cells and examined the consequences of this in a tissue recombination model. Histopathologically grafted tissues resembled the conditional PPARγ KO mouse prostates. EM studies of PPARγ- and PPARγ2-deficient epithelial cells in vitro were suggestive of autophagy, consistent with the prostatic tissue analysis. This was confirmed by examining expression of beclin-1 and LC3. Gene expression profiling in PPARγ-/γ2-deficient cells indicated a major dysregulation of cell cycle control and metabolic signaling networks related to peroxisomal and lysosomal maturation, lipid oxidation and degradation. The putative autophagic phenotypes of PPARγ-deficient cells could be rescued by re-expression of either γ1 or γ2 isoform. We conclude that disruption of PPARγ signaling results in autophagy and oxidative stress during mPIN pathogenesis. PMID:19834493

  7. Isolation, identification and enrichment of type a spermatogonia from the testis of chinese cross-bred buffaloes (swamp × river).

    PubMed

    Ahmad, S; Xiao, Y; Han, L; Hua, H; Riaz, H; Liang, A; Yang, L-G

    2013-06-01

    The proportion of type A spermatogonia in the isolated testis cells is a prerequisite for conducting experiments and the manipulation of these germ cells. Thus, this study was designed to examine the wide range of strategies for the isolation, identification and enrichment of type A spermatogonia in pre-pubertal buffalo calves (3-6 months). Histological findings revealed the presence of maximum number of type A spermatogonia at 5 months, which was further confirmed by DBA immunohistochemistry. In a newly modified strategy for the isolation of testis tissues, mincing followed by trituration and two rounds of digestion with collagenase, hyaluronidase and DNase yielded more than 95% testis cell population. Differential plating with laminin, poly-l-lysine and gelatin significantly (p < 0.05) affected the purity of type A spermatogonia. Among these extracellular matrix (ECMs) molecules, laminin and gelatin performed well and reached at a purity of 39.38 ± 1.21% and 32.15 ± 1.60%, respectively. In addition, combination of laminin and gelatin followed by Percoll centrifugation performed the best and yielded >90% type A spermatogonial purity. Moreover, viability of the cells was not affected (p > 0.05) irrespective of different enrichment methods. In conclusion, type A spermatogonia isolation and enrichment system was developed using different ECM molecules in buffaloes, which will aid in solving wide range of problems especially fertility-related problems and transgenic animal production in buffaloes.

  8. Cerebellar and Extracerebellar Involvement in Mouse Eyeblink Conditioning: the ACDC Model

    PubMed Central

    Boele, Henk-Jan; Koekkoek, Sebastiaan K. E.; De Zeeuw, Chris I.

    2009-01-01

    Over the past decade the advent of mouse transgenics has generated new perspectives on the study of cerebellar molecular mechanisms that are essential for eyeblink conditioning. However, it also appears that results from eyeblink conditioning experiments done in mice differ in some aspects from results previously obtained in other mammals. In this review article we will, based on studies using (cell-specific) mouse mutants and region-specific lesions, re-examine the general eyeblink behavior in mice and the neuro-anatomical circuits that might contribute to the different peaks in the conditioned eyeblink trace. We conclude that the learning process in mice has at least two stages: An early stage, which includes short-latency responses that are at least partly controlled by extracerebellar structures such as the amygdala, and a later stage, which is represented by well-timed conditioned responses that are mainly controlled by the pontocerebellar and olivocerebellar systems. We refer to this overall concept as the Amygdala-Cerebellum-Dynamic-Conditioning Model (ACDC model). PMID:20126519

  9. In vitro differentiation of rat spermatogonia into round spermatids in tissue culture

    PubMed Central

    Reda, A.; Hou, M.; Winton, T.R.; Chapin, R.E.; Söder, O.; Stukenborg, J.-B.

    2016-01-01

    STUDY QUESTION Do the organ culture conditions, previously defined for in vitro murine male germ cell differentiation, also result in differentiation of rat spermatogonia into post-meiotic germ cells exhibiting specific markers for haploid germ cells? SUMMARY ANSWER We demonstrated the differentiation of rat spermatogonia into post-meiotic cells in vitro, with emphasis on exhibiting, protein markers described for round spermatids. WHAT IS KNOWN ALREADY Full spermatogenesis in vitro from immature germ cells using an organ culture technique in mice was first reported 5 years ago. However, no studies reporting the differentiation of rat spermatogonia into post-meiotic germ cells exhibiting the characteristic protein expression profile or into functional sperm have been reported. STUDY DESIGN, SAMPLES/MATERIALS, METHODS Organ culture of testicular fragments of 5 days postpartum (dpp) neonatal rats was performed for up to 52 days. Evaluation of microscopic morphology, testosterone levels, mRNA and protein expression as measured by RT-qPCR and immunostaining were conducted to monitor germ cell differentiation in vitro. Potential effects of melatonin, Glutamax® medium, retinoic acid and the presence of epidydimal fat tissue on the spermatogenic process were evaluated. A minimum of three biological replicates were performed for all experiments presented in this study. One-way ANOVA, ANOVA on ranks and student's t-test were applied to perform the statistical analysis. MAIN RESULTS AND THE ROLE OF CHANCE Male germ cells, present in testicular tissue pieces grown from 5 dpp rats, exhibited positive protein expression for Acrosin and Crem (cAMP (cyclic adenosine mono phosphate) response element modulator) after 52 days of culture in vitro. Intra-testicular testosterone production could be observed after 3 days of culture, while when epididymal fat tissue was added, spontaneous contractility of cultured seminiferous tubules could be observed after 21 days. However, no

  10. Calcium imaging reveals glial involvement in transcranial direct current stimulation-induced plasticity in mouse brain

    PubMed Central

    Monai, Hiromu; Ohkura, Masamichi; Tanaka, Mika; Oe, Yuki; Konno, Ayumu; Hirai, Hirokazu; Mikoshiba, Katsuhiko; Itohara, Shigeyoshi; Nakai, Junichi; Iwai, Youichi; Hirase, Hajime

    2016-01-01

    Transcranical direct current stimulation (tDCS) is a treatment known to ameliorate various neurological conditions and enhance memory and cognition in humans. tDCS has gained traction for its potential therapeutic value; however, little is known about its mechanism of action. Using a transgenic mouse expressing G-CaMP7 in astrocytes and a subpopulation of excitatory neurons, we find that tDCS induces large-amplitude astrocytic Ca2+ surges across the entire cortex with no obvious changes in the local field potential. Moreover, sensory evoked cortical responses are enhanced after tDCS. These enhancements are dependent on the alpha-1 adrenergic receptor and are not observed in IP3R2 (inositol trisphosphate receptor type 2) knockout mice, in which astrocytic Ca2+ surges are absent. Together, we propose that tDCS changes the metaplasticity of the cortex through astrocytic Ca2+/IP3 signalling. PMID:27000523

  11. p75 neurotrophin receptor is involved in proliferation of undifferentiated mouse embryonic stem cells

    SciTech Connect

    Moscatelli, Ilana; Pierantozzi, Enrico; Camaioni, Antonella; Siracusa, Gregorio; Campagnolo, Luisa

    2009-11-01

    Neurotrophins and their receptors are known to play a role in the proliferation and survival of many different cell types of neuronal and non-neuronal lineages. In addition, there is much evidence in the literature showing that the p75 neurotrophin receptor (p75{sup NTR}), alone or in association with members of the family of Trk receptors, is expressed in a wide variety of stem cells, although its role in such cells has not been completely elucidated. In the present work we have investigated the expression of p75{sup NTR} and Trks in totipotent and pluripotent cells, the mouse pre-implantation embryo and embryonic stem and germ cells (ES and EG cells). p75{sup NTR} and TrkA can be first detected in the blastocyst from which ES cell lines are derived. Mouse ES cells retain p75{sup NTR}/TrkA expression. Nerve growth factor is the only neurotrophin able to stimulate ES cell growth in culture, without affecting the expression of stem cell markers, alkaline phosphatase, Oct4 and Nanog. Such proliferation effect was blocked by antagonizing either p75{sup NTR} or TrkA. Interestingly, immunoreactivity to anti-p75{sup NTR} antibodies is lost upon ES cell differentiation. The expression pattern of neurotrophin receptors in murine ES cells differs from human ES cells, that only express TrkB and C, and do not respond to NGF. In this paper we also show that, while primordial germ cells (PGC) do not express p75{sup NTR}, when they are made to revert to an ES-like phenotype, becoming EG cells, expression of p75{sup NTR} is turned on.

  12. TRAIP is involved in chromosome alignment and SAC regulation in mouse oocyte meiosis

    PubMed Central

    Yuan, Yi-Feng; Ren, Yi-Xin; Yuan, Peng; Yan, Li-Ying; Qiao, Jie

    2016-01-01

    Recent whole-exome sequencing (WES) studies demonstrated that TRAIP is associated with primordial dwarfism. Although TRAIP was partially studied in mitosis, its function in oocyte meiosis remained unknown. In this study, we investigated the roles of TRAIP during mouse oocyte meiosis. TRAIP was stably expressed during oocytes meiosis and co-localized with CREST at the centromere region. Knockdown of TRAIP led to DNA damage, as revealed by the appearance of γH2AX. Although oocytes meiotic maturation was not affected, the proportions of misaligned chromosomes and aneuploidy were elevated after TRAIP knockdown, suggesting TRAIP is required for stable kinetochore–microtubule (K-MT) attachment. TRAIP knockdown decreased the accumulation of Mad2 on centromeres, potentially explaining why oocyte maturation was not affected following formation of DNA lesions. Securin, a protein which was prevent from precocious degradation by Mad2, was down-regulated after TRAIP knockdown. Inhibition of TRAIP by microinjection of antibody into pro-metaphase I (pro-MI) stage oocytes resulted in precocious first polar body (PB1) extrusion, and live-cell imaging clearly revealed misaligned chromosomes after TRAIP knockdown. Taken together, these data indicate that TRAIP plays important roles in oocyte meiosis regulation. PMID:27405720

  13. TRAIP is involved in chromosome alignment and SAC regulation in mouse oocyte meiosis.

    PubMed

    Yuan, Yi-Feng; Ren, Yi-Xin; Yuan, Peng; Yan, Li-Ying; Qiao, Jie

    2016-07-11

    Recent whole-exome sequencing (WES) studies demonstrated that TRAIP is associated with primordial dwarfism. Although TRAIP was partially studied in mitosis, its function in oocyte meiosis remained unknown. In this study, we investigated the roles of TRAIP during mouse oocyte meiosis. TRAIP was stably expressed during oocytes meiosis and co-localized with CREST at the centromere region. Knockdown of TRAIP led to DNA damage, as revealed by the appearance of γH2AX. Although oocytes meiotic maturation was not affected, the proportions of misaligned chromosomes and aneuploidy were elevated after TRAIP knockdown, suggesting TRAIP is required for stable kinetochore-microtubule (K-MT) attachment. TRAIP knockdown decreased the accumulation of Mad2 on centromeres, potentially explaining why oocyte maturation was not affected following formation of DNA lesions. Securin, a protein which was prevent from precocious degradation by Mad2, was down-regulated after TRAIP knockdown. Inhibition of TRAIP by microinjection of antibody into pro-metaphase I (pro-MI) stage oocytes resulted in precocious first polar body (PB1) extrusion, and live-cell imaging clearly revealed misaligned chromosomes after TRAIP knockdown. Taken together, these data indicate that TRAIP plays important roles in oocyte meiosis regulation.

  14. TRAIP is involved in chromosome alignment and SAC regulation in mouse oocyte meiosis.

    PubMed

    Yuan, Yi-Feng; Ren, Yi-Xin; Yuan, Peng; Yan, Li-Ying; Qiao, Jie

    2016-01-01

    Recent whole-exome sequencing (WES) studies demonstrated that TRAIP is associated with primordial dwarfism. Although TRAIP was partially studied in mitosis, its function in oocyte meiosis remained unknown. In this study, we investigated the roles of TRAIP during mouse oocyte meiosis. TRAIP was stably expressed during oocytes meiosis and co-localized with CREST at the centromere region. Knockdown of TRAIP led to DNA damage, as revealed by the appearance of γH2AX. Although oocytes meiotic maturation was not affected, the proportions of misaligned chromosomes and aneuploidy were elevated after TRAIP knockdown, suggesting TRAIP is required for stable kinetochore-microtubule (K-MT) attachment. TRAIP knockdown decreased the accumulation of Mad2 on centromeres, potentially explaining why oocyte maturation was not affected following formation of DNA lesions. Securin, a protein which was prevent from precocious degradation by Mad2, was down-regulated after TRAIP knockdown. Inhibition of TRAIP by microinjection of antibody into pro-metaphase I (pro-MI) stage oocytes resulted in precocious first polar body (PB1) extrusion, and live-cell imaging clearly revealed misaligned chromosomes after TRAIP knockdown. Taken together, these data indicate that TRAIP plays important roles in oocyte meiosis regulation. PMID:27405720

  15. An inversion involving the mouse Shh locus results in brachydactyly through dysregulation of Shh expression.

    PubMed

    Niedermaier, Michael; Schwabe, Georg C; Fees, Stephan; Helmrich, Anne; Brieske, Norbert; Seemann, Petra; Hecht, Jochen; Seitz, Volkhard; Stricker, Sigmar; Leschik, Gundula; Schrock, Evelin; Selby, Paul B; Mundlos, Stefan

    2005-04-01

    Short digits (Dsh) is a radiation-induced mouse mutant. Homozygous mice are characterized by multiple defects strongly resembling those resulting from Sonic hedgehog (Shh) inactivation. Heterozygous mice show a limb reduction phenotype with fusion and shortening of the proximal and middle phalanges in all digits, similar to human brachydactyly type A1, a condition caused by mutations in Indian hedgehog (IHH). We mapped Dsh to chromosome 5 in a region containing Shh and were able to demonstrate an inversion comprising 11.7 Mb. The distal breakpoint is 13.298 kb upstream of Shh, separating the coding sequence from several putative regulatory elements identified by interspecies comparison. The inversion results in almost complete downregulation of Shh expression during E9.5-E12.5, explaining the homozygous phenotype. At E13.5 and E14.5, however, Shh is upregulated in the phalangeal anlagen of Dsh/+ mice, at a time point and in a region where WT Shh is never expressed. The dysregulation of Shh expression causes the local upregulation of hedgehog target genes such as Gli1-3, patched, and Pthlh, as well as the downregulation of Ihh and Gdf5. This results in shortening of the digits through an arrest of chondrocyte differentiation and the disruption of joint development.

  16. Cancer stem cells from human breast tumors are involved in spontaneous metastases in orthotopic mouse models

    PubMed Central

    Liu, Huiping; Patel, Manishkumar R.; Prescher, Jennifer A.; Patsialou, Antonia; Qian, Dalong; Lin, Jiahui; Wen, Susanna; Chang, Ya-Fang; Bachmann, Michael H.; Shimono, Yohei; Dalerba, Piero; Adorno, Maddalena; Lobo, Neethan; Bueno, Janet; Dirbas, Frederick M.; Goswami, Sumanta; Somlo, George; Condeelis, John; Contag, Christopher H.; Gambhir, Sanjiv Sam; Clarke, Michael F.

    2010-01-01

    To examine the role of breast cancer stem cells (BCSCs) in metastasis, we generated human-in-mouse breast cancer orthotopic models using patient tumor specimens, labeled with optical reporter fusion genes. These models recapitulate human cancer features not captured with previous models, including spontaneous metastasis in particular, and provide a useful platform for studies of breast tumor initiation and progression. With noninvasive imaging approaches, as few as 10 cells of stably labeled BCSCs could be tracked in vivo, enabling studies of early tumor growth and spontaneous metastasis. These advances in BCSC imaging revealed that CD44+ cells from both primary tumors and lung metastases are highly enriched for tumor-initiating cells. Our metastatic cancer models, combined with noninvasive imaging techniques, constitute an integrated approach that could be applied to dissect the molecular mechanisms underlying the dissemination of metastatic CSCs (MCSCs) and to explore therapeutic strategies targeting MCSCs in general or to evaluate individual patient tumor cells and predict response to therapy. PMID:20921380

  17. Involvement of ceramide in ethanol-induced apoptotic neurodegeneration in the neonatal mouse brain

    PubMed Central

    Saito, Mariko; Chakraborty, Goutam; Hegde, Medha; Ohsie, Jason; Paik, Sun-Mee; Vadasz, Csaba; Saito, Mitsuo

    2010-01-01

    Acute administration of ethanol to 7-day-old mice is known to cause robust apoptotic neurodegeneration in the brain. Our previous studies have shown that such ethanol-induced neurodegeneration is accompanied by increases in lipids including ceramide, triglyceride, cholesterol ester, and N-acyl phosphatidylethanolamine in the brain. In this study, the effects of ethanol on lipid profiles as well as caspase-3 activation were examined in the cortex, hippocampus, cerebellum, and inferior colliculus of the P7 mouse brain. We found that the cortex, hippocampus, and inferior colliculus, which showed substantial caspase-3 activation by ethanol, manifested significant elevations in ceramide, triglyceride, and N-acylphosphatidylethanolamine. In contrast, the cerebellum, with the least caspase-3 activation, failed to show significant changes in ceramide and triglyceride, and exhibits much smaller increases in N-acyl phosphatidylethanolamine than other brain regions. Ethanol-induced increases in cholesterol ester were observed in all brain regions tested. Inhibitors of serine palmitoyltransferase effectively blocked ethanol-induced caspase-3 activation as well as elevations in ceramide, cholesterol ester, and N-acyl phosphatidylethanolamine. Immunohistochemical studies indicated that the expression of serine palmitoyltransferase was mainly localized in neurons and was enhanced in activated caspase-3-positive neurons generated by ethanol. These results indicate that de novo ceramide synthesis has a vital role in ethanol-induced apoptotic neurodegeneration in the developing brain. PMID:20663015

  18. Nrf2 signalling and autophagy are involved in diabetes mellitus-induced defects in the development of mouse placenta.

    PubMed

    He, Mei-Yao; Wang, Guang; Han, Sha-Sha; Jin, Ya; Li, He; Wu, Xia; Ma, Zheng-Lai; Cheng, Xin; Tang, Xiuwen; Yang, Xuesong; Liu, Guo-Sheng

    2016-07-01

    It is widely accepted that diabetes mellitus impairs placental development, but the mechanism by which the disease operates to impair development remains controversial. In this study, we demonstrated that pregestational diabetes mellitus (PGDM)-induced defects in placental development in mice are mainly characterized by the changes of morphological structure of placenta. The alteration of differentiation-related gene expressions in trophoblast cells rather than cell proliferation/apoptosis is responsible for the phenotypes found in mouse placenta. Meanwhile, excess reactive oxygen species (ROS) production and activated nuclear factor erythroid2-related factor 2 (Nrf2) signalling were observed in the placenta of mice suffering from PGDM. Using BeWo cells, we also demonstrated that excess ROS was produced and Nrf2 signalling molecules were activated in settings characterized by a high concentration of glucose. More interestingly, differentiation-related gene expressions in trophoblast cells were altered when endogenous Nrf2 expression is manipulated by transfecting Nrf2-wt or Nrf2-shRNA. In addition, PGDM interferes with autophagy in both mouse placenta and BeWo cells, implying that autophagy is also involved, directly or indirectly, in PGDM-induced placental phenotypes. Therefore, we revealed that dysfunctional oxidative stress-activated Nrf2 signalling and autophagy are probably responsible for PGDM-induced defects in the placental development of mice. The mechanism was through the interference with differentiation-related gene expression in trophoblast cells. PMID:27383629

  19. A transgenic insertion on mouse chromosome 17 inactivates a novel immunoglobulin superfamily gene potentially involved in sperm-egg fusion.

    PubMed

    Lorenzetti, Diego; Poirier, Christophe; Zhao, Ming; Overbeek, Paul A; Harrison, Wilbur; Bishop, Colin E

    2014-04-01

    Fertilization is the process that leads to the formation of a diploid zygote from two haploid gametes. This is achieved through a complex series of cell-to-cell interactions between a sperm and an egg. The final event of fertilization is the fusion of the gametes' membranes, which allows the delivery of the sperm genetic material into the egg cytoplasm. In vivo studies in the laboratory mouse have led to the discovery of membrane proteins that are essential for the fusion process in both the sperm and egg. Specifically, the sperm protein Izumo1 was shown to be necessary for normal fertility. Izumo1-deficient spermatozoa fail to fuse with the egg plasma membrane. Izumo1 is a member of the Immunoglobulin Superfamily of proteins, which are known to be involved in cell adhesion. Here, we describe BART97b, a new mouse line with a recessive mutation that displays a fertilization block associated with a failure of sperm fusion. BART97b mutants carry a deletion that inactivates Spaca6, a previously uncharacterized gene expressed in testis. Similar to Izumo1, Spaca6 encodes an immunoglobulin-like protein. We propose that the Spaca6 gene product may, together with Izumo1, mediate sperm fusion by binding an as yet unidentified egg membrane receptor.

  20. Nrf2 signalling and autophagy are involved in diabetes mellitus-induced defects in the development of mouse placenta

    PubMed Central

    Han, Sha-sha; Jin, Ya; Li, He; Wu, Xia; Ma, Zheng-lai; cheng, Xin; Tang, Xiuwen

    2016-01-01

    It is widely accepted that diabetes mellitus impairs placental development, but the mechanism by which the disease operates to impair development remains controversial. In this study, we demonstrated that pregestational diabetes mellitus (PGDM)-induced defects in placental development in mice are mainly characterized by the changes of morphological structure of placenta. The alteration of differentiation-related gene expressions in trophoblast cells rather than cell proliferation/apoptosis is responsible for the phenotypes found in mouse placenta. Meanwhile, excess reactive oxygen species (ROS) production and activated nuclear factor erythroid2-related factor 2 (Nrf2) signalling were observed in the placenta of mice suffering from PGDM. Using BeWo cells, we also demonstrated that excess ROS was produced and Nrf2 signalling molecules were activated in settings characterized by a high concentration of glucose. More interestingly, differentiation-related gene expressions in trophoblast cells were altered when endogenous Nrf2 expression is manipulated by transfecting Nrf2-wt or Nrf2-shRNA. In addition, PGDM interferes with autophagy in both mouse placenta and BeWo cells, implying that autophagy is also involved, directly or indirectly, in PGDM-induced placental phenotypes. Therefore, we revealed that dysfunctional oxidative stress-activated Nrf2 signalling and autophagy are probably responsible for PGDM-induced defects in the placental development of mice. The mechanism was through the interference with differentiation-related gene expression in trophoblast cells. PMID:27383629

  1. Docosahexaenoic Acid Ameliorates Fructose-Induced Hepatic Steatosis Involving ER Stress Response in Primary Mouse Hepatocytes.

    PubMed

    Zheng, Jinying; Peng, Chuan; Ai, Yanbiao; Wang, Heng; Xiao, Xiaoqiu; Li, Jibin

    2016-01-01

    The increase in fructose consumption is considered to be a risk factor for developing nonalcoholic fatty liver disease (NAFLD). We investigated the effects of docosahexaenoic acid (DHA) on hepatic lipid metabolism in fructose-treated primary mouse hepatocytes, and the changes of Endoplasmic reticulum (ER) stress pathways in response to DHA treatment. The hepatocytes were treated with fructose, DHA, fructose plus DHA, tunicamycin (TM) or fructose plus 4-phenylbutyric acid (PBA) for 24 h. Intracellular triglyceride (TG) accumulation was assessed by Oil Red O staining. The mRNA expression levels and protein levels related to lipid metabolism and ER stress response were determined by real-time PCR and Western blot. Fructose treatment led to obvious TG accumulation in primary hepatocytes through increasing expression of fatty acid synthase (FAS) and acetyl-CoA carboxylase (ACC), two key enzymes in hepatic de novo lipogenesis. DHA ameliorates fructose-induced TG accumulation by upregulating the expression of carnitine palmitoyltransferase 1A (CPT-1α) and acyl-CoA oxidase 1 (ACOX1). DHA treatment or pretreatment with the ER stress inhibitor PBA significantly decreased TG accumulation and reduced the expression of glucose-regulated protein 78 (GRP78), total inositol-requiring kinase 1 (IRE1α) and p-IRE1α. The present results suggest that DHA protects against high fructose-induced hepatocellular lipid accumulation. The current findings also suggest that alleviating the ER stress response seems to play a role in the prevention of fructose-induced hepatic steatosis by DHA. PMID:26805874

  2. Transcriptional coactivator undifferentiated embryonic cell transcription factor 1 expressed in spermatogonial stem cells: a putative marker of boar spermatogonia.

    PubMed

    Lee, Won-Young; Lee, Kyung-Hoon; Heo, Young-Tae; Kim, Nam-Hyung; Kim, Jin-Hoi; Kim, Jae-Hwan; Moon, Sung-Hwan; Chung, Hak-Jae; Yoon, Min-Jung; Song, Hyuk

    2014-11-30

    Spermatogenesis is initiated from spermatogonial stem cells (SSCs), which are derived from gonocytes. Although some rodent SSC markers have been investigated, other species- and developmental stage-specific markers of spermatogonia have not been identified. The objective of this study was to characterize the expression of undifferentiated embryonic cell transcription factor 1 (UTF1) gene as a potential marker for spermatogonia and SSCs in the boar testis. In boar testis tissue at pre-pubertal stages (tissues collected at 5, 30, and 60 days of age), UTF1 gene expression was detected in almost all spermatogonia cells that expressed a protein gene product 9.5 (PGP9.5), and immunocytochemical analysis of isolated total testicular cells showed that 91.14% of cells staining for PGP9.5 also stained for UTF1. However, in boar testis tissue at pubertal and post-pubertal stages (tissues collected at 90, 120, 150, and 180 days of age), UTF1 was not detected in all PGP9.5-positive cells in the basement membrane. While some PGP9.5-positive cells stained for UTF1, other cells stained only for PGP9.5 or UTF1. PGP9.5, UTF1, and NANOG was assessed in in vitro cultures of pig SSCs (pSSCs) from testes collected at 5 days of age. The relative amounts of PGP9.5, NANOG, and UTF1 mRNA were greater in pSSC colonies than in testis and muscle tissue. Thus, the UTF1 gene is expressed in PGP9.5-positive spermatogonia cells of pigs at 5 days of age, and its expression is maintained in cultured pSSC colonies, suggesting that UTF1 is a putative marker for early-stage spermatogonia in the pre-pubertal pig testis. These findings will facilitate the study of spermatogenesis and applications in germ cell research.

  3. Generation of slow wave type action potentials in the mouse small intestine involves a non-L-type calcium channel.

    PubMed

    Malysz, J; Richardson, D; Farraway, L; Christen, M O; Huizinga, J D

    1995-10-01

    Intrinsic electrical activities in various isolated segments of the mouse small intestine were recorded (i) to characterize action potential generation and (ii) to obtain a profile on the ion channels involved in initiating the slow wave type action potentials (slow waves). Gradients in slow wave frequency, resting membrane potential, and occurrence of spiking activity were found, with the proximal intestine exhibiting the highest frequency, the most hyperpolarized cell membrane, and the greatest occurrence of spikes. The slow waves were only partially sensitive to L-type calcium channel blockers. Nifedipine, verapamil, and pinaverium bromide abolished spikes that occurred on the plateau phase of the slow waves in all tissues. The activity that remained in the presence of L-type calcium channel blockers, the upstroke potential, retained a similar amplitude to the original slow wave and was of identical frequency. The upstroke potential was not sensitive to a reduction in extracellular chloride or to the sodium channel blockers tetrodotoxin and mexiletine. Abolishment of the Na+ gradient by removal of 120 mM extracellular Na+ reduced the upstroke potential frequency by 13 - 18% and its amplitude by 50 - 70% in the ileum. The amplitude was similarly reduced by Ni2+ (up to 5 mM), and by flufenamic acid (100 mu M), a nonspecific cation and chloride channel blocker. Gadolinium, a nonspecific blocker of cation and stretch-activated channels, had no effect. Throughout these pharmacological manipulations, a robust oscillation remained at 5 - 10 mV. This oscillation likely reflects pacemaker activity. It was rapidly abolished by removal of extracellular calcium but not affected by L-type calcium channel blockers. In summary, the mouse small intestine has been established as a model for research into slow wave generation and electrical pacemaker activity. The upstroke part of the slow wave has two components, the pacemaker component involves a non-L-type calcium channel

  4. Involvement of MAPK ERK activation in upregulation of water channel protein aquaporin 1 in a mouse model of Bell's palsy.

    PubMed

    Fang, Fan; Liu, Cai-Yue; Zhang, Jie; Zhu, Lie; Qian, Yu-Xin; Yi, Jing; Xiang, Zheng-Hua; Wang, Hui; Jiang, Hua

    2015-05-01

    The aim of this study is to immunolocalize the aquaporin 1 water channel protein (AQP1) in Schwann cells of idiopathic facial nerve and explore its possible role during the development of facial palsy induced by herpes simplex virus type 1 (HSV-1). HSV-1 was inoculated into the surface of posterior auricle of mouse to establish a paralyzed animal model. In HSV-1-induced facial palsy mice, protein levels of AQP1 significantly increased on the 9th to 16th day after inoculation of HSV-1. The upregulation of AQP1 was closely related to the intratemporal facial nerve edema in facial nerve canal, which was also consistent with the symptom of facial palsy in mice. In a hypoxia model of Schwann cells in vitro, we found that U0126, an ERK antagonist, inhibited not only morphological changes of cultures Schwann cells but also upregulation of both AQP1 and phosphorylated ERK. Combined with increased phosphorylated ERK in HSV-1-induced facial palsy mice, we inferred that ERK MAPK pathway might also be involved in increased AQP1 in mouse model of Bell's palsy. Although the precise mechanism needs to be further explored, our findings suggest that AQP1 in Schwann cells of intratemporal facial nerve is involved in the evolution of facial palsy induced by HSV-1 and may play an important role in the pathogenesis of this disease. AQP1 might be a potential target, and the ERK antagonist U0126 could be a new drug for the treatment of HSV-1-induced Bell's palsy in an early stage. PMID:25527444

  5. Hyperhomocysteinemia and bleomycin hydrolase modulate the expression of mouse brain proteins involved in neurodegeneration.

    PubMed

    Suszyńska-Zajczyk, Joanna; Luczak, Magdalena; Marczak, Lukasz; Jakubowski, Hieronim

    2014-01-01

    Homocysteine (Hcy) is a risk factor for Alzheimer's disease (AD). Bleomycin hydrolase (BLMH) participates in Hcy metabolism and is also linked to AD. The inactivation of the Blmh gene in mice causes accumulation of Hcy-thiolactone in the brain and increases susceptibility to Hcy-thiolactone-induced seizures. To gain insight into brain-related Blmh function, we used two-dimensional IEF/SDS-PAGE gel electrophoresis and MALDI-TOF/TOF mass spectrometry to examine brain proteomes of Blmh-/- mice and their Blmh+/+ littermates fed with a hyperhomocysteinemic high-Met or a control diet. We found that: (1) proteins involved in brain-specific function (Ncald, Nrgn, Stmn1, Stmn2), antioxidant defenses (Aop1), cell cycle (RhoGDI1, Ran), and cytoskeleton assembly (Tbcb, CapZa2) were differentially expressed in brains of Blmh-null mice; (2) hyperhomocysteinemia amplified effects of the Blmh-/- genotype on brain protein expression; (3) proteins involved in brain-specific function (Pebp1), antioxidant defenses (Sod1, Prdx2, DJ-1), energy metabolism (Atp5d, Ak1, Pgam-B), and iron metabolism (Fth) showed differential expression in Blmh-null brains only in hyperhomocysteinemic animals; (4) most proteins regulated by the Blmh-/- genotype were also regulated by high-Met diet, albeit in the opposite direction; and (5) the differentially expressed proteins play important roles in neural development, learning, plasticity, and aging and are linked to neurodegenerative diseases, including AD. Taken together, our findings suggest that Blmh interacts with diverse cellular processes from energy metabolism and anti-oxidative defenses to cell cycle, cytoskeleton dynamics, and synaptic plasticity essential for normal brain homeostasis and that modulation of these interactions by hyperhomocysteinemia underlies the involvement of Hcy in AD.

  6. Hypoxic preconditioning involves system Xc- regulation in mouse neural stem cells.

    PubMed

    Sims, Brian; Clarke, Melinda; Francillion, Ludwig; Kindred, Elijah; Hopkins, Elana Shuford; Sontheimer, Harald

    2012-03-01

    In animals, hypoxic preconditioning has been used as a form of neuroprotection. The exact mechanism involved in neuroprotective hypoxic preconditioning has not been described, yet could be valuable for possible neuroprotective strategies. The overexpression of the cystine-glutamate exchanger, system Xc-, has been demonstrated as being neuroprotective (Shih, Erb et al. 2006). Here, using immunohistochemistry, we demonstrate that C57BL/6 mice exposed to hypoxia showed an increase in system Xc- expression, with the highest level of intensity in the hippocampus. Western Blot analysis also showed an almost 2-fold increase in system Xc- protein in hypoxia-exposed versus control mice. The mRNA for the regulatory subunit of system Xc-, xCT, and the xCT/actin ratio were also increased under hypoxic conditions. Experiments using hypoxia-inducible factor (HIF-1α) siRNA showed a statistically significant decrease in HIF-1α and system Xc- expression. Under hypoxic conditions, system Xc- activity, as determined by cystine uptake, increased 2-fold. Importantly, hypoxic preconditioning was attenuated in neural stem cells by pharmacological inhibition of system Xc- activity with S4-carboxyphenylglycine. These data provide the first evidence of hypoxic regulation of the cystine glutamate exchanger system Xc-.

  7. Host and Bacterial Factors Involved in the Innate Ability of Mouse Macrophages To Eliminate Internalized Unopsonized Escherichia coli

    PubMed Central

    Hamrick, Terri S.; Havell, Edward A.; Horton, John R.; Orndorff, Paul E.

    2000-01-01

    In an effort to better understand genetic and cellular factors that influence innate immunity, we examined host and bacterial factors involved in the nonopsonic phagocytosis and killing of Escherichia coli K-12 by mouse macrophages. Unelicited (resident) peritoneal macrophages from five different mouse strains, BALB/c, C57BL/6, CD-1, C3H/HeJ, and C3H/HeN, were employed. Additional macrophage populations were obtained from CD-1 mice (bone marrow-derived macrophages). Also, for BALB/c and C57BL/6 mice, peritoneal macrophages elicited with either thioglycolate or proteose peptone, bone marrow-derived macrophages, and macrophage-like cell lines derived from the two strains were employed. Two E. coli K-12 strains that differed specifically in their abilities to produce type 1 pili containing the adhesive protein FimH were examined. The parameters used to assess macrophage bacteriocidal activity were (i) the killing of internalized (gentamicin-protected) E. coli during the approximately 4-h assay and (ii) the initial rate at which internalized E. coli were eliminated. Data on these parameters allowed the following conclusions: (i) unelicited or proteose peptone-elicited peritoneal macrophages were significantly better at eliminating internalized bacteria than thioglycolate-elicited peritoneal macrophages, bone marrow-derived macrophages, or macrophage cell lines; (ii) the host genetic background had no significant effect upon the ability of unelicited peritoneal macrophages to kill E. coli (even though the mouse strains differ widely in their in vivo susceptibilities to bacterial infection); and (iii) the FimH phenotype had no significant effect upon E. coli survival once the bacterium was inside a macrophage. Additionally, there was no correlation between the bacteriocidal effectiveness of a macrophage population and the number of bacteria bound per macrophage. However, macrophage populations that were the least bacteriocidal tended to bind higher ratios of FimH+ to Fim

  8. Endothelins are involved in regulating the proliferation and differentiation of mouse epidermal melanocytes in serum-free primary culture.

    PubMed

    Hirobe, T

    2001-11-01

    ET-1, -2, and -3 are one member of the keratinocyte-derived factors that are involved in regulating the proliferation and differentiation of mouse epidermal melanocytes in primary culture.

  9. PAK1 is involved in sensing the orientation of collagen stiffness gradients in mouse fibroblasts.

    PubMed

    Pinto, V I; Mohammadi, H; Lee, W S; Cheung, A H; McCulloch, C A

    2015-10-01

    Migrating cells sense variations of stiffness in connective tissue matrices but how cells detect and respond to stiffness orientation is not defined. We examined cell extension formation on collagen with underlying support (vertical stiffness gradient) or on collagen laterally supported by nylon (lateral stiffness gradient). At 6 h after plating, cells plated on laterally-supported collagen exhibited >2-fold more abundant and ~2-fold longer cell extensions than cells plated on collagen with underlying support. We examined whether p21-activated kinase 1 (PAK1) influences extension formation that is dependent on the orientation of support. At 6 h after plating on collagen with underlying support, wild-type cell extensions were 40% shorter than PAK1 knockdown cells. In contrast, on laterally-supported collagen, wild-type cell extensions were 2-fold longer than PAK1 knockdown cells. In cells plated on laterally-supported collagen, there were ~2-fold reductions of collagen fiber alignment and compaction in PAK1 knockdown cells compared with wild-type cells. PAK1 knockdown did not affect collagen fiber alignment or compaction by cells plated on collagen with underlying support. Wild-type cells with lateral support of collagen exhibited 3-fold increases of phospho-myosin staining at 6h, which was 2-fold lower in PAK1 knockdown cells. In contrast, cells on collagen with underlying support showed no increase of phospho-myosin staining at any times. PAK1 knockdown did not affect α2 or β1 integrin expression or function. We conclude that PAK1 is involved in the ability of cells to sense the orientation of stiffness in collagen substrates and generate contractile forces that affect cell extension formation.

  10. Beta-arrestin 1 is involved in the catabolic response stimulated by hyaluronan degradation in mouse chondrocytes.

    PubMed

    Campo, Giuseppe M; Avenoso, Angela; D'Ascola, Angela; Scuruchi, Michele; Calatroni, Alberto; Campo, Salvatore

    2015-08-01

    Beta-arrestin-1 (β-arrestin-1) is an adaptor protein that functions in the termination of G-protein activation and seems to be involved in the mediation of the inflammatory response. Interleukin-1β (IL-1β) elicits the expression of inflammatory mediators through a mechanism involving hyaluronan (HA) degradation, thereby contributing to toll-like receptor 4 (TLR-4) and CD44 activation. Stimulation of both receptors induces nuclear factor kappaB (NF-kB) activation that, through transforming-growth-factor-activated-kinase-1 (TAK-1), in turn stimulates the inflammatory mediators of transcription. As β-arrestin-1 seems to play an inflammatory role in arthritis, we have investigated the involvement of β-arrestin-1 in a model of IL-1β-induced inflammatory response in mouse chondrocytes. IL-1β treatment significantly increases chondrocytes TLR-4, CD44, β-arrestin-1, TAK-1, and serine/threonine kinase (AKT) mRNA expression and related protein levels. NF-kB is also markedly activated with consequent tumor-necrosis-factor-alpha, interleukin-6, and inducible-nitric-oxide-synthase up-regulation. Treatment of IL-1β-stimulated chondrocytes with β-arrestin-1 and/or AKT and/or TAK-1-specific inhibitors significantly reduces all parameters, although the inhibitory effect exerted by TAK-1-mediated pathways is more effective than that of β-arrestin-1. β-Arrestin-1-induced NF-kB activation is mediated by the AKT pathway as shown by IL-1β-stimulated chondrocytes treated with AKT inhibitor. Finally, a specific HA-blocking peptide (Pep-1) has confirmed the inflammatory role of degraded HA as a mediator of the IL-1β-induced activation of β-arrestin-1. PMID:25673209

  11. Beta-arrestin 1 is involved in the catabolic response stimulated by hyaluronan degradation in mouse chondrocytes.

    PubMed

    Campo, Giuseppe M; Avenoso, Angela; D'Ascola, Angela; Scuruchi, Michele; Calatroni, Alberto; Campo, Salvatore

    2015-08-01

    Beta-arrestin-1 (β-arrestin-1) is an adaptor protein that functions in the termination of G-protein activation and seems to be involved in the mediation of the inflammatory response. Interleukin-1β (IL-1β) elicits the expression of inflammatory mediators through a mechanism involving hyaluronan (HA) degradation, thereby contributing to toll-like receptor 4 (TLR-4) and CD44 activation. Stimulation of both receptors induces nuclear factor kappaB (NF-kB) activation that, through transforming-growth-factor-activated-kinase-1 (TAK-1), in turn stimulates the inflammatory mediators of transcription. As β-arrestin-1 seems to play an inflammatory role in arthritis, we have investigated the involvement of β-arrestin-1 in a model of IL-1β-induced inflammatory response in mouse chondrocytes. IL-1β treatment significantly increases chondrocytes TLR-4, CD44, β-arrestin-1, TAK-1, and serine/threonine kinase (AKT) mRNA expression and related protein levels. NF-kB is also markedly activated with consequent tumor-necrosis-factor-alpha, interleukin-6, and inducible-nitric-oxide-synthase up-regulation. Treatment of IL-1β-stimulated chondrocytes with β-arrestin-1 and/or AKT and/or TAK-1-specific inhibitors significantly reduces all parameters, although the inhibitory effect exerted by TAK-1-mediated pathways is more effective than that of β-arrestin-1. β-Arrestin-1-induced NF-kB activation is mediated by the AKT pathway as shown by IL-1β-stimulated chondrocytes treated with AKT inhibitor. Finally, a specific HA-blocking peptide (Pep-1) has confirmed the inflammatory role of degraded HA as a mediator of the IL-1β-induced activation of β-arrestin-1.

  12. THY1 as a reliable marker for enrichment of undifferentiated spermatogonia in the goat.

    PubMed

    Abbasi, Hassan; Tahmoorespur, Mojtaba; Hosseini, Sayyed Morteza; Nasiri, Zahra; Bahadorani, Mehrnoosh; Hajian, Mahdi; Nasiri, Mohammad Reza; Nasr-Esfahani, Mohammad Hossein

    2013-11-01

    Spermatogonial stem cells are unique cells of testes that can restore fertility upon transplantation into recipient testes. However, use of suitable markers for enrichment of these cells have important potential application. THY1, is an established conserved marker of spermatogonial stem cells in bovine, rodents, and primates, but there is no information available in goats. After three rounds of enzymatic digestion of prepubertal goat testicular tissues, undifferentiated spermatogonia positive for THY1 were isolated by magnetic-activated cell sorting and were used for immunocytochemistry, real-time polymerase chain reaction analysis for gene expression, protein expression, and transplantation into recipient mice. Immunocytochemical analyses showed that significantly higher percentage of THY1(+) cells were positive for PLZF and VASA when compared with unselected population. This result for PLZF was further confirmed at the protein level. Real-time polymerase chain reaction analysis revealed that expression of THY1, PLZF, VASA, BCL6B, and UCHL1 as SCCs characteristic genes in THY1(+) cells was significantly higher than in the initial population. Finally, transplantation of PKH26-labeled cells revealed that THY1(+) cells had higher capacity for colony formation when compared with unselected cells. In conclusion, the results provide indications that THY1 surface marker can be reliably used for enrichment of undifferentiated spermatogonial in the goats.

  13. Conservation of spermatogonial stem cell marker expression in undifferentiated felid spermatogonia.

    PubMed

    Vansandt, Lindsey M; Livesay, Janelle L; Dickson, Melissa Joy; Li, Lei; Pukazhenthi, Budhan S; Keefer, Carol L

    2016-09-01

    Spermatogonial stem cells (SSCs) are distinct in their ability to self-renew, transmit genetic information, and persist throughout the life of an individual. These characteristics make SSCs a useful tool for addressing diverse challenges such as efficient transgenic production in nonrodent, biomedical animal models, or preservation of the male genome for species in which survival of frozen-thawed sperm is low. A requisite first step to access this technology in felids is the establishment of molecular markers. This study was designed to evaluate, in the domestic cat (Felis catus), the expression both in situ and following enrichment in vitro of six genes (GFRA1, GPR125, ZBTB16, POU5F1, THY1, and UCHL1) that had been previously identified as SSC markers in other species. Antibodies for surface markers glial cell line-derived neurotrophic factor family receptor alpha 1, G protein-coupled receptor 125, and thymus cell antigen 1 could not be validated, whereas Western blot analysis of prepubertal, peripubertal, and adult cat testis confirmed protein expression for the intracellular markers ubiquitin carboxy-terminal hydrolase 1, zinc finger and BTB domain-containing protein 16, and POU domain, class 5, transcription factor 1. Colocalization of the markers by immunohistochemistry revealed that several cells within the subpopulation adjacent to the basement membrane of the seminiferous tubules and identified morphologically as spermatogonia, expressed all three intracellular markers. Studies performed on cheetah (Acinonyx jubatus) and Amur leopard (Panthera pardus orientalis) testis exhibited a conserved expression pattern in protein molecular weights, relative abundance, and localization of positive cells within the testis. The expression of the three intracellular SSC marker proteins in domestic and wild cat testes confirms conservation of these markers in felids. Enrichment of marker transcripts after differential plating was also observed. These markers will

  14. Involvement of insulin-degrading enzyme in insulin- and atrial natriuretic peptide-sensitive internalization of amyloid-β peptide in mouse brain capillary endothelial cells.

    PubMed

    Ito, Shingo; Ohtsuki, Sumio; Murata, Sho; Katsukura, Yuki; Suzuki, Hiroya; Funaki, Miho; Tachikawa, Masanori; Terasaki, Tetsuya

    2014-01-01

    Cerebral clearance of amyloid-β peptide (Aβ), which is implicated in Alzheimer's disease, involves elimination across the blood-brain barrier (BBB), and we previously showed that an insulin-sensitive process is involved in the case of Aβ1-40. The purpose of this study was to clarify the molecular mechanism of the insulin-sensitive Aβ1-40 elimination across mouse BBB. An in vivo cerebral microinjection study demonstrated that [125I]hAβ1-40 elimination from mouse brain was inhibited by human natriuretic peptide (hANP), and [125I]hANP elimination was inhibited by hAβ1-40, suggesting that hAβ1-40 and hANP share a common elimination process. Internalization of [125I]hAβ1-40 into cultured mouse brain capillary endothelial cells (TM-BBB4) was significantly inhibited by either insulin, hANP, other natriuretic peptides or insulin-degrading enzyme (IDE) inhibitors, but was not inhibited by phosphoramidon or thiorphan. Although we have reported the involvement of natriuretic peptide receptor C (Npr-C) in hANP internalization, cells stably expressing Npr-C internalized [125I]hANP but not [125I]hAβ1-40, suggesting that there is no direct interaction between Npr-C and hAβ1-40. IDE was detected in plasma membrane of TM-BBB4 cells, and internalization of [125I]hAβ1-40 by TM-BBB4 cells was reduced by IDE-targeted siRNAs. We conclude that elimination of hAβ1-40 from mouse brain across the BBB involves an insulin- and ANP-sensitive process, mediated by IDE expressed in brain capillary endothelial cells.

  15. Conditional Allele Mouse Planner (CAMP): software to facilitate the planning and design of breeding strategies involving mice with conditional alleles.

    PubMed

    Hoffert, Jason D; Pisitkun, Trairak; Miller, R Lance

    2012-06-01

    Transgenic and conditional knockout mouse models play an important role in biomedical research and their use has grown exponentially in the last 5-10 years. Generating conditional knockouts often requires breeding multiple alleles onto the background of a single mouse or group of mice. Breeding these mice depends on parental genotype, litter size, transmission frequency, and the number of breeding rounds. Therefore, a well planned breeding strategy is critical for keeping costs to a minimum. However, designing a viable breeding strategy can be challenging. With so many different variables this would be an ideal task for a computer program. To facilitate this process, we created a Java-based program called Conditional Allele Mouse Planner (CAMP). CAMP is designed to provide an estimate of the number of breeders, amount of time, and costs associated with generating mice of a particular genotype. We provide a description of CAMP, how to use it, and offer it freely as an application.

  16. Nitric oxide synthase expression correlates with death in an experimental mouse model of dengue with CNS involvement

    PubMed Central

    2013-01-01

    Background The clinical presentation of dengue is classified by the World Health Organization into dengue without warning signs, dengue with warning signs and severe dengue. Reports of neurological disease caused by Dengue virus (DENV) are becoming frequent, with symptoms that include reduced consciousness, severe headache, neck stiffness, focal neurological signs, tense fontanelle and convulsions. However, the immune mechanisms involved in neurovirulence remain poorly understood. Here we present a mouse model in which one genotype of DENV is inoculated by the intracranial route and infects C57/BL6 mice and replicates in the brain, causing death of mice. Methods Mice were infected with different serotypes/genotypes of DENV by the intracranial route to evaluate viral replication, host cytokine and nitric oxide synthase 2 (Nos2) expression in the brain via real-time PCR. Histological analysis of the brain tissues was also performed. An analysis of which cells were responsible for the expression of cytokines and Nos2 was performed using flow cytometry. Survival curves of infected animals were also generated Results DENV 3 genotype I infected mice and replicated in the brain, causing death in our murine model. The increased levels of NOS2 could be the cause of the death of infected mice, as viral replication correlates with increased Nos2 and cytokine expression in the brain of C57BL/6 mice. In Nos2−/− mice that were infected with DENV, no clinical signs of infection were observed and cytokines were expressed at low levels, with the exception of interferon gamma (Ifng). Additionally, the Ifng−/− mice infected with DENV exhibited a severe and lethal disease, similar to the disease observed in C57BL/6 mice, while the DENV- infected Nos2−/− mice did not display increased mortality. Analyses of the brains from infected C57BL/6 mice revealed neuronal degeneration and necrosis during histopathologic examination. IFNg and NOS2 were produced in the brains of

  17. Tumor growth affects the metabonomic phenotypes of multiple mouse non-involved organs in an A549 lung cancer xenograft model

    PubMed Central

    Xu, Shan; Tian, Yuan; Hu, Yili; Zhang, Nijia; Hu, Sheng; Song, Dandan; Wu, Zhengshun; Wang, Yulan; Cui, Yanfang; Tang, Huiru

    2016-01-01

    The effects of tumorigenesis and tumor growth on the non-involved organs remain poorly understood although many research efforts have already been made for understanding the metabolic phenotypes of various tumors. To better the situation, we systematically analyzed the metabolic phenotypes of multiple non-involved mouse organ tissues (heart, liver, spleen, lung and kidney) in an A549 lung cancer xenograft model at two different tumor-growth stages using the NMR-based metabonomics approaches. We found that tumor growth caused significant metabonomic changes in multiple non-involved organ tissues involving numerous metabolic pathways, including glycolysis, TCA cycle and metabolisms of amino acids, fatty acids, choline and nucleic acids. Amongst these, the common effects are enhanced glycolysis and nucleoside/nucleotide metabolisms. These findings provided essential biochemistry information about the effects of tumor growth on the non-involved organs. PMID:27329570

  18. Kinetic characteristics of norcocaine N-hydroxylation in mouse and human liver microsomes: involvement of CYP enzymes.

    PubMed

    Pellinen, P; Kulmala, L; Konttila, J; Auriola, S; Pasanen, M; Juvonen, R

    2000-11-01

    The first step in the oxidative metabolism of cocaine is N-demethylation to norcocaine, which is further N-hydroxylated to more toxic N-hydroxynorcocaine. In this study we examined the kinetics of norcocaine N-hydroxylation mediated by cytochrome P450 (CYP) in mouse and human liver microsomes. N-hydroxynorcocaine was identified by analytical HPLC-MS after incubation of norcocaine with mouse liver microsomes in the presence of NADPH. In mouse liver microsomes, there was no apparent difference in Km values for norcocaine N-hydroxylation between male and female microsomes, while the Vmax rate was approximately two times higher in female than in male microsomes (34+/-10 v. 16+/-4 pmol/min per mg protein). The Km value for norcocaine N-hydroxylation in human liver microsomes was approximately three times higher than that observed in comparable incubations using mouse liver microsomes, whereas the Vmax rate was ten times lower. Both cocaine and norcocaine induced type I difference spectra upon interaction with CYP in mouse liver microsomes. In contrast, in human microsomes both type I and type II spectra were recorded. In the 0.01 to 1 mM concentration range, cocaine and norcocaine inhibited mouse microsomal testosterone 6alpha-, 7alpha- and 16alpha-hydroxylation reactions by 20% to 30%. Testosterone 6beta- and 15alpha-hydroxylations were blocked by 60% and 50%, respectively, by 1 mM norcocaine, while only 40% inhibition was obtained with 1 mM cocaine. Coumarin 7-hydroxylation and pentoxyresorufin O-deethylation were inhibited by 50% by 1 and 0.4 mM norcocaine, respectively. In contrast, 10 and 2 mM cocaine, respectively, were needed to obtain the same degrees of inhibition. In human liver microsomes, 1 mM norcocaine and cocaine blocked testosterone 6beta-hydroxylase by 60% and 40%, respectively. Coumarin 7-hydroxylation was inhibited by only 30% by norcocaine (5.4 mM) and cocaine (10 mM). Norcocaine N-hydroxylation in mouse and human liver microsomes was blocked by 30

  19. PCSK9 is not involved in the degradation of LDL receptors and BACE1 in the adult mouse brain

    PubMed Central

    Liu, Mali; Wu, Guoxin; Baysarowich, Jennifer; Kavana, Michael; Addona, George H.; Bierilo, Kathleen K.; Mudgett, John S.; Pavlovic, Guillaume; Sitlani, Ayesha; Renger, John J.; Hubbard, Brian K.; Fisher, Timothy S.; Zerbinatti, Celina V.

    2010-01-01

    Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secreted protein that regulates hepatic low-density lipoprotein receptor (LDLR) levels in humans. PCSK9 has also been shown to regulate the levels of additional membrane-bound proteins in vitro, including the very low-density lipoprotein receptor (VLDLR), apolipoprotein E receptor 2 (ApoER2) and the β-site amyloid precursor protein (APP)-cleaving enzyme 1 (BACE1), which are all highly expressed in the CNS and have been implicated in Alzheimer's disease. To better understand the role of PCSK9 in regulating these additional target proteins in vivo, their steady-state levels were measured in the brain of wild-type, PCSK9-deficient, and human PCSK9 overexpressing transgenic mice. We found that while PCSK9 directly bound to recombinant LDLR, VLDLR, and apoER2 protein in vitro, changes in PCSK9 expression did not alter the level of these receptors in the mouse brain. In addition, we found no evidence that PCSK9 regulates BACE1 levels or APP processing in the mouse brain. In conclusion, our results suggest that while PCSK9 plays an important role in regulating circulating LDL cholesterol levels by reducing the number of hepatic LDLRs, it does not appear to modulate the levels of LDLR and other membrane-bound proteins in the adult mouse brain. PMID:20453200

  20. Involvement of Polycomb Repressive Complex 2 in Maturation of Induced Pluripotent Stem Cells during Reprogramming of Mouse and Human Fibroblasts.

    PubMed

    Khazaie, Niusha; Massumi, Mohammad; Wee, Ping; Salimi, Mahdieh; Mohammadnia, Abdulshakour; Yaqubi, Moein

    2016-01-01

    Induced pluripotent stem cells (iPSCs) provide a reliable source for the study of regenerative medicine, drug discovery, and developmental biology. Despite extensive studies on the reprogramming of mouse and human fibroblasts into iPSCs, the efficiency of reprogramming is still low. Here, we used a bioinformatics and systems biology approach to study the two gene regulatory waves governing the reprogramming of mouse and human fibroblasts into iPSCs. Our results revealed that the maturation phase of reprogramming was regulated by a more complex regulatory network of transcription factors compared to the initiation phase. Interestingly, in addition to pluripotency factors, the polycomb repressive complex 2 (PRC2) members Ezh2, Eed, Jarid2, Mtf2, and Suz12 are crucially recruited during the maturation phase of reprogramming. Moreover, we found that during the maturation phase of reprogramming, pluripotency factors, via the expression and induction of PRC2 complex members, could silence the lineage-specific gene expression program and maintain a ground state of pluripotency in human and mouse naïve iPSCs. The findings obtained here provide us a better understanding of the gene regulatory network (GRN) that governs reprogramming, and the maintenance of the naïve state of iPSCs.

  1. Involvement of Polycomb Repressive Complex 2 in Maturation of Induced Pluripotent Stem Cells during Reprogramming of Mouse and Human Fibroblasts

    PubMed Central

    Khazaie, Niusha; Massumi, Mohammad; Wee, Ping; Salimi, Mahdieh; Mohammadnia, Abdulshakour; Yaqubi, Moein

    2016-01-01

    Induced pluripotent stem cells (iPSCs) provide a reliable source for the study of regenerative medicine, drug discovery, and developmental biology. Despite extensive studies on the reprogramming of mouse and human fibroblasts into iPSCs, the efficiency of reprogramming is still low. Here, we used a bioinformatics and systems biology approach to study the two gene regulatory waves governing the reprogramming of mouse and human fibroblasts into iPSCs. Our results revealed that the maturation phase of reprogramming was regulated by a more complex regulatory network of transcription factors compared to the initiation phase. Interestingly, in addition to pluripotency factors, the polycomb repressive complex 2 (PRC2) members Ezh2, Eed, Jarid2, Mtf2, and Suz12 are crucially recruited during the maturation phase of reprogramming. Moreover, we found that during the maturation phase of reprogramming, pluripotency factors, via the expression and induction of PRC2 complex members, could silence the lineage-specific gene expression program and maintain a ground state of pluripotency in human and mouse naïve iPSCs. The findings obtained here provide us a better understanding of the gene regulatory network (GRN) that governs reprogramming, and the maintenance of the naïve state of iPSCs. PMID:26938987

  2. 5-aza-2'-deoxycytidine impairs mouse spermatogenesis at multiple stages through different usage of DNA methyltransferases.

    PubMed

    Song, Ning; Endo, Daisuke; Song, Bin; Shibata, Yasuaki; Koji, Takehiko

    2016-06-15

    Mammalian spermatogenesis is a progressive process comprising spermatogonial proliferation, spermatocytic meiosis, and later spermiogenesis, which is considered to be under the regulation of epigenetic parameters. To gain insights into the significance of DNA methylation in early spermatogenesis, 5-azadC was used as a molecular biological tool to mimic the level of DNA methylation in vivo. Since the drug is incorporated into DNA during the S-phase, spermatogonia and spermatocytes would be affected primarily in mouse spermatogenesis. Adult male ICR mice were intraperitoneally injected with 5-azadC at a dose of 0.25mg/kg/day for 10 consecutive days, allowing us to examine its maximum effect on the kinetics of spermatogonia and spermatocytes. In this short-term protocol, 5-azadC induced significant histological abnormalities, such as a marked increase in apoptosis of spermatogonia and spermatocytes, followed by severe loss of spermatids, while after termination of 5-azadC treatment, normal histology was restored in the testis within 35days. Quantification of the methylation level of CCGG sites as well as whole DNA showed spermatogonial hypomethylation, which correlated with increased apoptosis of spermatogonia. Interestingly, the hypomethylated cells were simultaneously positive for tri-methylated histone H3 at K4. On the other hand, no changes in methylation level were found in spermatocytes, but PCNA staining clearly showed disordered accumulation of S-phase spermatocytes, which increased their apoptosis in stage XII. In addition, different immunohistochemical staining pattern was found for DNA methyltransferases (DNMTs); DNMT1was expressed in the majority of all germ cells, but DNMT3a and b were only expressed in spermatogonia. Our results indicate that 5-azadC caused DNA hypomethylation in spermatogonia, but induced prolongation of S-phase in spermatocytes, resulting in the induction of apoptosis in both cases. Thus, 5-azadC affects spermatogenesis at more than

  3. Possible involvement of nitric oxide (NO) signaling pathway in the antidepressant-like effect of MK-801(dizocilpine), a NMDA receptor antagonist in mouse forced swim test.

    PubMed

    Dhir, Ashish; Kulkarni, S K

    2008-03-01

    L-arginine-nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) is an important signaling pathway involved in depression. With this information, the present study aimed to study the involvement of this signaling pathway in the antidepressant-like action of MK-801 (dizocilpine; N-methyl-d-aspartate receptor antagonist) in the mouse forced-swim test. Total immobility period was recorded in mouse forced swim test for 6 min. MK-801 (5-25 microg/kg., ip) produced a U-shaped curve in reducing the immobility period. The antidepressant-like effect of MK-801 (10 microg/kg, ip) was prevented by pretreatment with L-arginine (750 mg/kg, ip) [substrate for nitric oxide synthase (NOS)]. Pretreatment of mice with 7-nitroindazole (7-NI) (25 mg/kg, ip) [a specific neuronal nitric oxide synthase inhibitor] produced potentiation of the action of subeffective dose of MK-801 (5 microg/kg, ip). In addition, treatment of mice with methylene blue (10 mg/kg, ip) [direct inhibitor of both nitric oxide synthase and soluble guanylate cyclase] potentiated the effect of MK-801 (5 microg/kg, ip) in the forced-swim test. Further, the reduction in the immobility period elicited by MK-801 (10 microg/kg, ip) was also inhibited by pretreatment with sildenafil (5 mg/kg, ip) [phosphodiesterase 5 inhibitor]. The various modulators used in the study and their combination did not produce any changes in locomotor activity per se and in combination with MK-801. MK-801 however, at higher doses (25 microg/kg, ip) produced hyperlocomotion. The results demonstrated the involvement of nitric oxide signaling pathway in the antidepressant-like effect of MK-801 in mouse forced-swim test.

  4. Involvement of dopamine D1 receptors and alpha1-adrenoceptors in the antidepressant-like effect of chlorpheniramine in the mouse tail suspension test.

    PubMed

    Hirano, Shoko; Miyata, Shigeo; Onodera, Kenji; Kamei, Junzo

    2007-05-01

    It has been reported that chlorpheniramine, a classical antihistamine, has antidepressant-like effects in animal models of depression. In this study, we examined the involvement of dopaminergic (dopamine D(1) and dopamine D(2) receptors), noradrenergic (alpha(1)- and beta-adrenoceptors) and serotonergic (5-HT(1A) and 5-HT(2) receptors) receptors in the antidepressant-like effect of chlorpheniramine in the mouse tail suspension test. We also investigated the involvement of these monoamine receptors in the antidepressant-like effect of imipramine for comparison with the mechanisms of the effect of chlorpheniramine. Both imipramine and chlorpheniramine significantly reduced the duration of immobility in the tail suspension test without affecting spontaneous locomotor activity in mice. The anti-immobility effect of imipramine (30 mg/kg, i.p.) was significantly antagonized by the selective dopamine D(1) receptor antagonist SCH23390 but not by the other receptor antagonists. In contrast, the anti-immobility effect of chlorpheniramine was significantly inhibited by SCH23390 and the selective alpha(1)-adrenoceptor antagonist prazosin, but not by the other receptor antagonists. In conclusion, these results suggest that chlorpheniramine exerts an antidepressant-like effect in the mouse tail suspension test that is mediated by at least the activation of dopamine D(1) receptors and alpha(1)-adrenoceptors. In addition, the antidepressant-like effect of chlorpheniramine may be induced by several mechanisms that are different from those involved in the antidepressant-like effect of imipramine.

  5. Evidence that glucose metabolism is decreased in the cerebrum of aged female senescence-accelerated mouse; possible involvement of a low hexokinase activity.

    PubMed

    Kurokawa, T; Sato, E; Inoue, A; Ishibashi, S

    1996-08-16

    d-Glucose metabolism in cerebral cells prepared from aged senescence-accelerated mouse (SAM), was investigated in consideration of a sex difference. The production of 14CO2 from 6-[14C]D-glucose was reduced in female senescence-accelerated-prone mouse (SAMP) 8, a prone substrain, in comparison with that in female senescence-accelerated-resistant mouse (SAMR) 2, a control substrain, whereas there was no difference in males. The 2-deoxy-D-glucose uptake into cerebral cells from female SAMP8 was also lower than that of control mice. But, the 3-O-methyl-D-glucose uptake in SAMP8 was higher than that of SAMR2, suggesting that the low hexokinase activity was involved in the decreased glucose metabolism in cerebrum of SAMP8 females irrespective of glucose transporter. This possibility was supported by the finding that the contents of glucose 6-phosphate produced from glucose added to cerebral cells from SAMP8 was lower than that in ICR mice. PMID:8873128

  6. Naegleria fowleri glycoconjugates with residues of α-D-mannose are involved in adherence of trophozoites to mouse nasal mucosa.

    PubMed

    Carrasco-Yepez, Maricela; Campos-Rodriguez, Rafael; Godinez-Victoria, Marycarmen; Rodriguez-Monroy, Marco Aurelio; Jarillo-Luna, Adriana; Bonilla-Lemus, Patricia; De Oca, Arturo Contis-Montes; Rojas-Hernandez, Saul

    2013-10-01

    We analyzed the possible role of glycoconjugates containing α-D-mannose and α-D-glucose residues in adherence of trophozoites to mouse nasal epithelium. Trophozoites incubated with 20 μg of one of three different lectins which preferentially recognized these residues were inoculated intranasally in Balb/c mice. Mouse survival was 40% with Pisum sativum and Canavalia ensiformis and 20% with Galanthus nivalis amebic pretreatment, compared with 0% survival for control animals administered trophozoites without pretreatment. Possibly some of the glycoproteins found in Naegleria fowleri represent an adherence factor. Differences in the saccharide sequences of the Naegleria species, even on the same glycoconjugate structure, could explain the different results corresponding to the distinct pretreatments (C. ensiformis, G. nivalis, and P. sativum). We found a higher expression of glycoconjugates recognized by P. sativum in Naegleria lovaniensis than N. fowleri, probably due to the higher number of oligosaccharides containing an α-1,6-linked fucose moiety expressed on the former species. PMID:23922203

  7. Involvement of the T-box transcription factor Brachyury in early-stage embryonic mouse salivary gland.

    PubMed

    Hayashi, Kouhei; Ikari, Tatsuya; Sugiyama, Goro; Sugiura, Tsuyoshi; Ohyama, Yukiko; Kumamaru, Wataru; Shirasuna, Kanemitsu; Mori, Yoshihide

    2016-09-01

    The mouse submandibular gland (SMG) is important organ for embryonic development, and branching morphogenesis is regulated by many molecules containing transcription factors. Real-time reverse transcriptase polymerase chain reaction revealed that the expression of Brachyury increased in the SMG and peaked between E12.5-E13.5, concomitant with the early stage of branching morphogenesis. The expression of Brachyury in SMG rudiments between E12.5-E13.5 was confirmed by western blotting. In addition, fibronectin and Btbd7 (regulated by fibronectin), which are both essential for cleft formation, were expressed strongly during the same period. The Sox2 and Wnt3a, which regulate cell growth, were also expressed strongly during E12.5-E13.5. On the other hand, cleft formation and branching morphogenesis was suppressed by knockdown of Brachyury gene, suggesting that Brachyury plays a central role in regulating cell growth and cleft formation in early-stage embryonic mouse salivary gland development. PMID:27369076

  8. Relocalization of STIM1 in mouse oocytes at fertilization: early involvement of store-operated calcium entry.

    PubMed

    Gómez-Fernández, Carolina; Pozo-Guisado, Eulalia; Gañán-Parra, Miguel; Perianes, Mario J; Alvarez, Ignacio S; Martín-Romero, Francisco Javier

    2009-08-01

    Calcium waves represent one of the most important intracellular signaling events in oocytes at fertilization required for the exit from metaphase arrest and the resumption of the cell cycle. The molecular mechanism ruling this signaling has been described in terms of the contribution of intracellular calcium stores to calcium spikes. In this work, we considered the possible contribution of store-operated calcium entry (SOCE) to this signaling, by studying the localization of the protein STIM1 in oocytes. STIM1 has been suggested to play a key role in the recruitment and activation of plasma membrane calcium channels, and we show here that mature mouse oocytes express this protein distributed in discrete clusters throughout their periphery in resting cells, colocalizing with the endoplasmic reticulum marker calreticulin. However, immunolocalization of the endogenous STIM1 showed considerable redistribution over larger areas or patches covering the entire periphery of the oocyte during Ca(2+) store depletion induced with thapsigargin or ionomycin. Furthermore, pharmacological activation of endogenous phospholipase C induced a similar pattern of redistribution of STIM1 in the oocyte. Finally, fertilization of mouse oocytes revealed a significant and rapid relocalization of STIM1, similar to that found after pharmacological Ca(2+) store depletion. This particular relocalization supports a role for STIM1 and SOCE in the calcium signaling during early stages of fertilization.

  9. Naegleria fowleri glycoconjugates with residues of α-D-mannose are involved in adherence of trophozoites to mouse nasal mucosa.

    PubMed

    Carrasco-Yepez, Maricela; Campos-Rodriguez, Rafael; Godinez-Victoria, Marycarmen; Rodriguez-Monroy, Marco Aurelio; Jarillo-Luna, Adriana; Bonilla-Lemus, Patricia; De Oca, Arturo Contis-Montes; Rojas-Hernandez, Saul

    2013-10-01

    We analyzed the possible role of glycoconjugates containing α-D-mannose and α-D-glucose residues in adherence of trophozoites to mouse nasal epithelium. Trophozoites incubated with 20 μg of one of three different lectins which preferentially recognized these residues were inoculated intranasally in Balb/c mice. Mouse survival was 40% with Pisum sativum and Canavalia ensiformis and 20% with Galanthus nivalis amebic pretreatment, compared with 0% survival for control animals administered trophozoites without pretreatment. Possibly some of the glycoproteins found in Naegleria fowleri represent an adherence factor. Differences in the saccharide sequences of the Naegleria species, even on the same glycoconjugate structure, could explain the different results corresponding to the distinct pretreatments (C. ensiformis, G. nivalis, and P. sativum). We found a higher expression of glycoconjugates recognized by P. sativum in Naegleria lovaniensis than N. fowleri, probably due to the higher number of oligosaccharides containing an α-1,6-linked fucose moiety expressed on the former species.

  10. Relocalization of STIM1 in mouse oocytes at fertilization: early involvement of store-operated calcium entry.

    PubMed

    Gómez-Fernández, Carolina; Pozo-Guisado, Eulalia; Gañán-Parra, Miguel; Perianes, Mario J; Alvarez, Ignacio S; Martín-Romero, Francisco Javier

    2009-08-01

    Calcium waves represent one of the most important intracellular signaling events in oocytes at fertilization required for the exit from metaphase arrest and the resumption of the cell cycle. The molecular mechanism ruling this signaling has been described in terms of the contribution of intracellular calcium stores to calcium spikes. In this work, we considered the possible contribution of store-operated calcium entry (SOCE) to this signaling, by studying the localization of the protein STIM1 in oocytes. STIM1 has been suggested to play a key role in the recruitment and activation of plasma membrane calcium channels, and we show here that mature mouse oocytes express this protein distributed in discrete clusters throughout their periphery in resting cells, colocalizing with the endoplasmic reticulum marker calreticulin. However, immunolocalization of the endogenous STIM1 showed considerable redistribution over larger areas or patches covering the entire periphery of the oocyte during Ca(2+) store depletion induced with thapsigargin or ionomycin. Furthermore, pharmacological activation of endogenous phospholipase C induced a similar pattern of redistribution of STIM1 in the oocyte. Finally, fertilization of mouse oocytes revealed a significant and rapid relocalization of STIM1, similar to that found after pharmacological Ca(2+) store depletion. This particular relocalization supports a role for STIM1 and SOCE in the calcium signaling during early stages of fertilization. PMID:19470709

  11. Temporally controlled site-specific mutagenesis in the germ cell lineage of the mouse testis.

    PubMed

    Weber, Philipp; Schuler, Michael; Gérard, Christelle; Mark, Manuel; Metzger, Daniel; Chambon, Pierre

    2003-02-01

    We have obtained a PrP-Cre-ER(T) transgenic mouse line (28.8) that selectively expresses in testis the tamoxifen-inducible Cre-ER(T) recombinase under the control of a mouse Prion protein (PrP) promoter-containing genomic fragment. Cre-ER(T) is expressed in spermatogonia and spermatocytes, but not in Sertoli and Leydig cells. We also established reporter PrP-L-EGFP-L transgenic mice harboring a LoxP-flanked enhanced green fluorescent protein (EGFP) Cre reporter cassette under the control of the same PrP promoter-containing genomic fragment that exhibits prominent EGFP expression in brain and testis. Using the PrP-L-EGFP-L as well as other Cre-reporter mice, we demonstrate that tamoxifen administration efficiently and selectively induces Cre-mediated recombination in the germ cell lineage. The established PrP-Cre-ER(T) line should provide a valuable tool for studying functions of germ cell-expressed genes involved in spermatogenesis. PMID:12533419

  12. Reconstruction of the gene regulatory network involved in the sonic hedgehog pathway with a potential role in early development of the mouse brain.

    PubMed

    Liu, Jinhua; Wang, Xuelong; Li, Juan; Wang, Haifang; Wei, Gang; Yan, Jun

    2014-10-01

    The Sonic hedgehog (Shh) signaling pathway is crucial for pattern formation in early central nervous system development. By systematically analyzing high-throughput in situ hybridization data of E11.5 mouse brain, we found that Shh and its receptor Ptch1 define two adjacent mutually exclusive gene expression domains: Shh+Ptch1- and Shh-Ptch1+. These two domains are associated respectively with Foxa2 and Gata3, two transcription factors that play key roles in specifying them. Gata3 ChIP-seq experiments and RNA-seq assays on Gata3-knockdown cells revealed that Gata3 up-regulates the genes that are enriched in the Shh-Ptch1+ domain. Important Gata3 targets include Slit2 and Slit3, which are involved in the process of axon guidance, as well as Slc18a1, Th and Qdpr, which are associated with neurotransmitter synthesis and release. By contrast, Foxa2 both up-regulates the genes expressed in the Shh+Ptch1- domain and down-regulates the genes characteristic of the Shh-Ptch1+ domain. From these and other data, we were able to reconstruct a gene regulatory network governing both domains. Our work provides the first genome-wide characterization of the gene regulatory network involved in the Shh pathway that underlies pattern formation in the early mouse brain. PMID:25299227

  13. Involvement of atypical transcription factor E2F8 in the polyploidization during mouse and human decidualization

    PubMed Central

    Qi, Qian-Rong; Zhao, Xu-Yu; Zuo, Ru-Juan; Wang, Tong-Song; Gu, Xiao-Wei; Liu, Ji-Long; Yang, Zeng-Ming

    2015-01-01

    Polyploid decidual cells are specifically differentiated cells during mouse uterine decidualization. However, little is known about the regulatory mechanism and physiological significance of polyploidization in pregnancy. Here we report a novel role of E2F8 in the polyploidization of decidual cells in mice. E2F8 is highly expressed in decidual cells and regulated by progesterone through HB-EGF/EGFR/ERK/STAT3 signaling pathway. E2F8 transcriptionally suppresses CDK1, thus triggering the polyploidization of decidual cells. E2F8-mediated polyploidization is a response to stresses which are accompanied by decidualization. Interestingly, polyploidization is not detected during human decidualization with the down-regulation of E2F8, indicating differential expression of E2F8 may lead to the difference of decidual cell polyploidization between mice and humans. PMID:25892397

  14. Expression patterns of taste receptor type 1 subunit 3 and α-gustducin in the mouse testis during development.

    PubMed

    Gong, Ting; Wei, Quanwei; Mao, Dagan; Shi, Fangxiong

    2016-01-01

    Taste receptor type 1 subunit 3 (T1R3) and its associated heterotrimeric G protein α-gustducin (Gα) are involved in sweet and umami sensing in taste cells. They are also strongly expressed in the testis and sperm, but their expression patterns and potential roles involved were previously unknown. In present study, we investigated the expression patterns of T1R3 and Gα in the mouse testis at critical stages of postnatal life, and throughout the spermatogenic cycle. Our results indicated that T1R3 and Gα exhibited a stage-dependent expression pattern during mouse development, and a cell-specific pattern during the spermatogenic cycle. Their expressions have been increased significantly from prepubertal to pubertal periods (P<005), and decreased significantly in aged mice (P<005). The changes were mainly attributed to the differential expression of T1R3 or Gα in elongated spermatids and Leydig cells at different stages of the spermatogenic cycle. In addition, the expression of T1R3 and Gα were first observed in residual bodies of spermatozoa and endothelial cells of blood vessels at post-pubertal mice, while Gα was located in apoptotic spermatogonia of postnatal mice. These novel expression patterns suggest a role of T1R3 and Gα in the onset of spermatogenesis, pace of spermatogenic cycle, and aging of the testis. PMID:26589384

  15. Cytokinesis failure in RhoA-deficient mouse erythroblasts involves actomyosin and midbody dysregulation and triggers p53 activation

    PubMed Central

    Konstantinidis, Diamantis G.; Giger, Katie M.; Risinger, Mary; Pushkaran, Suvarnamala; Zhou, Ping; Dexheimer, Phillip; Yerneni, Satwica; Andreassen, Paul; Klingmüller, Ursula; Palis, James; Zheng, Yi

    2015-01-01

    RhoA GTPase has been shown in vitro in cell lines and in vivo in nonmammalian organisms to regulate cell division, particularly during cytokinesis and abscission, when 2 daughter cells partition through coordinated actomyosin and microtubule machineries. To investigate the role of this GTPase in the rapidly proliferating mammalian erythroid lineage, we developed a mouse model with erythroid-specific deletion of RhoA. This model was proved embryonic lethal as a result of severe anemia by embryonic day 16.5 (E16.5). The primitive red blood cells were enlarged, poikilocytic, and frequently multinucleated, but were able to sustain life despite experiencing cytokinesis failure. In contrast, definitive erythropoiesis failed and the mice died by E16.5, with profound reduction of maturing erythroblast populations within the fetal liver. RhoA was required to activate myosin-regulatory light chain and localized at the site of the midbody formation in dividing wild-type erythroblasts. Cytokinesis failure caused by RhoA deficiency resulted in p53 activation and p21-transcriptional upregulation with associated cell-cycle arrest, increased DNA damage, and cell death. Our findings demonstrate the role of RhoA as a critical regulator for efficient erythroblast proliferation and the p53 pathway as a powerful quality control mechanism in erythropoiesis. PMID:26228485

  16. Splicing changes in SMA mouse motoneurons and SMN-depleted neuroblastoma cells: Evidence for involvement of splicing regulatory proteins

    PubMed Central

    Huo, Qing; Kayikci, Melis; Odermatt, Philipp; Meyer, Kathrin; Michels, Olivia; Saxena, Smita; Ule, Jernej; Schümperli, Daniel

    2014-01-01

    Spinal Muscular Atrophy (SMA) is caused by deletions or mutations in the Survival Motor Neuron 1 (SMN1) gene. The second gene copy, SMN2, produces some, but not enough, functional SMN protein. SMN is essential to assemble small nuclear ribonucleoproteins (snRNPs) that form the spliceosome. However, it is not clear whether SMA is caused by defects in this function that could lead to splicing changes in all tissues, or by the impairment of an additional, less well characterized, but motoneuron-specific SMN function. We addressed the first possibility by exon junction microarray analysis of motoneurons (MNs) isolated by laser capture microdissection from a severe SMA mouse model. This revealed changes in multiple U2-dependent splicing events. Moreover, splicing appeared to be more strongly affected in MNs than in other cells. By testing mutiple genes in a model of progressive SMN depletion in NB2a neuroblastoma cells, we obtained evidence that U2-dependent splicing changes occur earlier than U12-dependent ones. As several of these changes affect genes coding for splicing regulators, this may acerbate the splicing response induced by low SMN levels and induce secondary waves of splicing alterations. PMID:25692239

  17. Mitofusin 2 expression dominates over mitofusin 1 exclusively in mouse dorsal root ganglia - a possible explanation for peripheral nervous system involvement in Charcot-Marie-Tooth 2A.

    PubMed

    Kawalec, Maria; Zabłocka, Barbara; Kabzińska, Dagmara; Neska, Jacek; Beręsewicz, Małgorzata

    2014-01-01

    Mitofusin 2 (Mfn2), a protein of the mitochondrial outer membrane, is essential for mitochondrial fusion and contributes to the maintenance and operation of the mitochondrial network. Mutations in the mitofusin 2 gene cause axonal Charcot-Marie-Tooth type 2A (CMT2A), an inherited disease affecting peripheral nerve axons. The precise mechanism by which mutations in MFN2 selectively cause the degeneration of long peripheral axons is not known. There is a hypothesis suggesting the involvement of reduced expression of a homologous protein, mitofusin 1 (Mfn1), in the peripheral nervous system, and less effective compensation of defective mitofusin 2 by mitofusin 1. We therefore aimed to perform an analysis of the mitofusin 1 and mitofusin 2 mRNA and protein expression profiles in different mouse tissues, with special attention paid to dorsal root ganglia (DRGs), as parts of the peripheral nervous system. Quantitative measurement relating to mRNA revealed that expression of the Mfn2 gene dominates over Mfn1 mainly in mouse DRG, as opposed to other nervous system samples and other tissues studied. This result was further supported by Western blot evaluation. Both these sets of data confirm the hypothesis that the cellular consequences of mutations in the mitofusin 2 gene can mostly be manifested in the peripheral nervous system. PMID:25574749

  18. Restraint Stress Inhibits Mouse Implantation: Temporal Window and the Involvement of HB-EGF, Estrogen and Progesterone

    PubMed Central

    Yuan, Hong-Jie; Liang, Bo; Zheng, Liang-Liang; Liu, Yu-Xiang; Luo, Ming-Jiu; Tan, Jing-He

    2013-01-01

    It is known that psychological stress affects reproduction in women, but it is unknown whether the effect is by impairing implantation. Although studies suggest that long periods of auditory or restraint stress may inhibit implantation in rats and mice, the exact stage of pregnancy at which stress impairs implantation is unclear. Furthermore, whether stress impairs implantation by decreasing the heparin-binding epidermal growth factor-like growth factor (HB-EGF), estrogen and/or progesterone and whether by acting on embryos or on the uterus need further investigations. In this study, a 24-h restraint stress was initiated at 15:30 of day 3 (regimen 1) or at 07:30 (regimen 2) or 15:30 of day 4 (regimen 3) of pregnancy (vaginal plug  =  day 1) to observe effects of restraint stress applied at different peri-implantation stages on implantation. Among the three regimens, whereas regimens 1 and 3 affected neither term pregnancy nor litter size, regimen 2 reduced both. Further observations indicated that regimen 2 of restraint stress also delayed blastocyst hatching and the attachment reaction, decreased serum concentrations of progesterone and estradiol, and down regulated the expression of HB-EGF in both the endometrium and blastocysts. Taken together, the results suggested that restraint stress inhibited mouse implantation in a temporal window-dependent manner and by impairing blastocyst activation and hatching and uterine receptivity via down-regulating HB-EGF, estrogen and progesterone. Thus, the stress applied within the implantation window impaired implantation by acting on both embryos and the uterus. PMID:24244689

  19. Genetic ablation of caspase-7 promotes solar-simulated light-induced mouse skin carcinogenesis: the involvement of keratin-17.

    PubMed

    Lee, Mee-Hyun; Lim, Do Young; Kim, Myoung Ok; Lee, Sung-Young; Shin, Seung Ho; Kim, Jae Young; Kim, Sung-Hyun; Kim, Dong Joon; Jung, Sung Keun; Yao, Ke; Kundu, Joydeb Kumar; Lee, Hye Suk; Lee, Cheol-Jung; Dickinson, Sally E; Alberts, David; Bowden, G Timothy; Stratton, Steven; Curiel, Clara; Einspahr, Janine; Bode, Ann M; Surh, Young-Joon; Cho, Yong-Yeon; Dong, Zigang

    2015-11-01

    Solar ultraviolet irradiation is an environmental carcinogen that causes skin cancer. Caspase-7 is reportedly expressed at reduced levels in many cancers. The present study was designed to examine the role of caspase-7 in solar-simulated light (SSL)-induced skin cancer and to elucidate its underlying molecular mechanisms. Our study revealed that mice with genetic deficiency of caspase-7 are highly susceptible to SSL-induced skin carcinogenesis. Epidermal hyperplasia, tumor volume and the average number of tumors were significantly increased in caspase-7 knockout (KO) mice compared with SKH1 wild-type mice irradiated with SSL. The expression of cell proliferation markers, such as survivin and Ki-67, was elevated in SSL-irradiated skin of caspase-7 KO mice compared with those observed in SSL-exposed wild-type SKH1 mouse skin. Moreover, SSL-induced apoptosis was abolished in skin from caspase-7 KO mice. Two-dimensional gel electrophoresis, followed by matrix-assisted laser desorption/ionization-time-of-flight analysis of skin tissue lysates from SSL-irradiated SKH1 wild-type and caspase-7 KO mice revealed an aberrant induction of keratin-17 in caspase-7 KO mice. Immunohistochemical analysis of skin tumors also showed an increase of keratin-17 expression in caspase-7 KO mice compared with SKH1 wild-type mice. The expression of keratin-17 was also elevated in SSL-irradiated caspase-7 KO keratinocytes as well as in human basal cell carcinomas. The in vitro caspase activity assay showed keratin-17 as a substrate of caspase-7, but not caspase-3. Overall, our study demonstrates that genetic loss of caspase-7 promotes SSL-induced skin carcinogenesis by blocking caspase-7-mediated cleavage of keratin-17. PMID:26271098

  20. Identification of a novel mouse Dbl proto-oncogene splice variant: evidence that SEC14 domain is involved in GEF activity regulation.

    PubMed

    Ognibene, Marzia; Vanni, Cristina; Blengio, Fabiola; Segalerba, Daniela; Mancini, Patrizia; De Marco, Patrizia; Torrisi, Maria R; Bosco, Maria C; Varesio, Luigi; Eva, Alessandra

    2014-03-10

    The Rho guanine nucleotide exchange factor protoDbl is involved in different biochemical pathways affecting cell proliferation and migration. The N-terminal sequence of protoDbl contains negative regulatory elements that restrict the catalytic activity of the DH-PH module. Here, we report the identification of a new mouse protoDbl splice variant lacking exon 3. We found that the splice variant mRNA is expressed in the spleen and bone marrow lymphocytes, adrenal gland, gonads and brain. The protoDbl variant protein was detectable in the brain. The newly identified variant displays the disruption of the SEC14 domain, positioned on exons 2 and 3 in the protoDbl N-terminal region. We show here that an altered SEC14 sequence leads to enhanced Dbl translocation to the plasma membrane and to augmented transforming and exchange activity.

  1. Curcumin analogue UBS109 prevents bone loss in breast cancer bone metastasis mouse model: involvement in osteoblastogenesis and osteoclastogenesis.

    PubMed

    Yamaguchi, Masayoshi; Zhu, Shijun; Zhang, Shumin; Wu, Daqing; Moore, Terry M; Snyder, James P; Shoji, Mamoru

    2014-07-01

    Bone metastasis of breast cancer typically leads to osteolysis, which causes severe pathological bone fractures and hypercalcemia. Bone homeostasis is skillfully regulated through osteoblasts and osteoclasts. Bone loss with bone metastasis of breast cancer may be due to both activation of osteoclastic bone resorption and suppression of osteoblastic bone formation. This study was undertaken to determine whether the novel curcumin analogue UBS109 has preventive effects on bone loss induced by breast cancer cell bone metastasis. Nude mice were inoculated with breast cancer MDA-MB-231 bone metastatic cells (10(6) cells/mouse) into the head of the right and left tibia. One week after inoculation, the mice were treated with control (vehicle), oral administration (p.o.) of UBS109 (50 or 150 mg/kg body weight), or intraperitoneal administration (i.p.) of UBS109 (10 or 20 mg/kg body weight) once daily for 5 days per week for 7 weeks. After UBS109 administration for 7 weeks, hind limbs were assessed using an X-ray diagnosis system and hematoxylin and eosion staining to determine osteolytic destruction. Bone marrow cells obtained from the femurs and tibias were cultured to estimate osteoblastic mineralization and osteoclastogenesis ex vivo and in vitro. Remarkable bone loss was demonstrated in the tibias of mice inoculated with breast cancer MDA-MB-231 bone metastatic cells. This bone loss was prevented by p.o. administration of UBS109 (50 and 150 mg/kg body weight) and i.p. treatment of UBS109 (10 and 20 mg/kg) in vivo. Culture of bone marrow cells obtained from the bone tissues of mice with breast cancer cell bone metastasis showed suppressed osteoblastic mineralization and stimulated osteoclastogenesis ex vivo. These changes were not seen after culture of the bone marrow cells obtained from mice treated with UBS109. Moreover, UBS109 was found to stimulate osteoblastic mineralization and suppress lipopolysaccharide (LPS)-induced osteoclastogenesis in bone marrow

  2. Interference by 2,3,7,8-tetrachlorodibenzo-p-dioxin with cultured mouse submandibular gland branching morphogenesis involves reduced epidermal growth factor receptor signaling

    SciTech Connect

    Kiukkonen, Anu . E-mail: anummela@mappi.helsinki.fi; Sahlberg, Carin; Partanen, Anna-Maija; Alaluusua, Satu; Pohjanvirta, Raimo; Tuomisto, Jouko; Lukinmaa, Pirjo-Liisa

    2006-05-01

    Toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to mouse embryonic teeth, sharing features of early development with salivary glands in common, involves enhanced apoptosis and depends on the expression of epidermal growth factor (EGF) receptor. EGF receptor signaling, on the other hand, is essential for salivary gland branching morphogenesis. To see if TCDD impairs salivary gland morphogenesis and if the impairment is associated with EGF receptor signaling, we cultured mouse (NMRI) E13 submandibular glands with TCDD or TCDD in combination with EGF or fibronectin (FN), both previously found to enhance branching morphogenesis. Explants were examined stereomicroscopically and processed to paraffin sections. TCDD exposure impaired epithelial branching and cleft formation, resulting in enlarged buds. The glands were smaller than normal. EGF and FN alone concentration-dependently stimulated or inhibited branching morphogenesis but when co-administered with TCDD, failed to compensate for its effect. TCDD induced cytochrome P4501A1 expression in the glandular epithelium, indicating activation of the aryl hydrocarbon receptor. TCDD somewhat increased epithelial apoptosis as observed by terminal deoxynucleotidyl transferase (TdT)-mediated nick end-labeling method but the increase could not be correlated with morphological changes. The frequency of proliferating cells was not altered. Corresponding to the reduced cleft sites in TCDD-exposed explants, FN immunoreactivity in the epithelium was reduced. The results show that TCDD, comparably with EGF and FN at morphogenesis-inhibiting concentrations, impaired salivary gland branching morphogenesis in vitro. Together with the failure of EGF and FN at morphogenesis-stimulating concentrations to compensate for the effect of TCDD this implies that TCDD toxicity to developing salivary gland involves reduced EGF receptor signaling.

  3. Involvement of EZH2, SUV39H1, G9a and associated molecules in pathogenesis of urethane induced mouse lung tumors: Potential targets for cancer control

    SciTech Connect

    Pandey, Manuraj; Sahay, Satya; Tiwari, Prakash; Upadhyay, Daya S.; Sultana, Sarwat; Gupta, Krishna P.

    2014-10-15

    In the present study, we showed the correlation of EZH2, SUV39H1 or G9a expression and histone modifications with the urethane induced mouse lung tumorigenesis in the presence or absence of antitumor agent, inositol hexaphosphate (IP6). Tumorigenesis and the molecular events involved therein were studied at 1, 4, 12 or 36 weeks after the exposure. There were no tumors at 1 or 4 weeks but tumors started appearing at 12 weeks and grew further till 36 weeks after urethane exposure. Among the molecular events, upregulation of EZH2 and SUV39H1 expressions appeared to be time dependent, but G9a expression was altered significantly only at later stages of 12 or 36 weeks. Alteration in miR-138 expression supports the upregulation of its target, EZH2. H3K9me2, H3K27me3 or H4K20me3 was found to be altered at 12 or 36 weeks. However, ChIP analysis of p16 and MLH1 promoters showed their binding with H3K9me2 and H3K27me3 which was maximum at 36 weeks. Thus, histone modification and their interactions with gene promoter resulted in the reduced expression of p16 and MLH1. IP6 prevented the incidence and the size of urethane induced lung tumors. IP6 also prevented the urethane induced alterations in EZH2, SUV39H1, G9a expressions and histone modifications. Our results suggest that the alterations in the histone modification pathways involving EZH2 and SUV39H1 expressions are among the early events in urethane induced mouse lung tumorigenesis and could be exploited for cancer control. - Highlights: • Urethane induces mouse lung tumor in a time dependent manner. • EZH2, SUV39H1, G9a induced by urethane and progress with time • Downregulation of miRNA-138 supports the EZH2 upregulation. • Methylation of histones showed a consequence of upregulated EZH2, SUV39H1 and G9a. • IP6 inhibits urethane induced changes and prevents tumor development.

  4. Regulation of Genes Involved in Carnitine Homeostasis by PPARα across Different Species (Rat, Mouse, Pig, Cattle, Chicken, and Human)

    PubMed Central

    Ringseis, Robert; Wen, Gaiping; Eder, Klaus

    2012-01-01

    Recent studies in rodents convincingly demonstrated that PPARα is a key regulator of genes involved in carnitine homeostasis, which serves as a reasonable explanation for the phenomenon that energy deprivation and fibrate treatment, both of which cause activation of hepatic PPARα, causes a strong increase of hepatic carnitine concentration in rats. The present paper aimed to comprehensively analyse available data from genetic and animal studies with mice, rats, pigs, cows, and laying hens and from human studies in order to compare the regulation of genes involved in carnitine homeostasis by PPARα across different species. Overall, our comparative analysis indicates that the role of PPARα as a regulator of carnitine homeostasis is well conserved across different species. However, despite demonstrating a well-conserved role of PPARα as a key regulator of carnitine homeostasis in general, our comprehensive analysis shows that this assumption particularly applies to the regulation by PPARα of carnitine uptake which is obviously highly conserved across species, whereas regulation by PPARα of carnitine biosynthesis appears less well conserved across species. PMID:23150726

  5. The Pacific bluefin tuna (Thunnus orientalis) dead end gene is suitable as a specific molecular marker of type A spermatogonia.

    PubMed

    Yazawa, Ryosuke; Takeuchi, Yutaka; Morita, Tetsuro; Ishida, Masashi; Yoshizaki, Goro

    2013-10-01

    We developed a spermatogonial transplantation technique to produce donor-derived gametes in surrogate fish. Our ultimate aim is to establish surrogate broodstock that can produce bluefin tuna. We previously determined that only type A spermatogonia (ASG) could colonize recipient gonads in salmonids. Therefore, it is necessary to develop a precise molecular marker that can distinguish ASG in order to develop efficient spermatogonial transplantation methods. In this study, the Pacific bluefin tuna (Thunnus orientalis) dead end (BFTdnd) gene was identified as a specific marker for ASG. In situ hybridization and RT-PCR analysis with various types of spermatogenic cell populations captured by laser microdissection revealed that localization of BFTdnd mRNA was restricted to ASG, and not detected in other differentiated spermatogenic cells. In order to determine if BFTdnd can be used as a molecular marker to identify germ cells with high transplantability, transplantation of dissociated testicular cells isolated from juvenile, immature, and mature Pacific bluefin tuna, which have different proportions of dnd-positive ASG, were performed using chub mackerel as the surrogate recipient species. Colonization of transplanted donor germ cells was only successful with testicular cells from immature Pacific Bluefin tuna, which contained higher proportions of dnd-positive ASG than juvenile and mature fish. Thus, BFTdnd is a useful tool for identifying highly transplantable ASG for spermatogonial transplantation.

  6. Metabotropic glutamate receptors are involved in the detection of IMP and L-amino acids by mouse taste sensory cells.

    PubMed

    Pal Choudhuri, S; Delay, R J; Delay, E R

    2016-03-01

    G-protein-coupled receptors are thought to be involved in the detection of umami and L-amino acid taste. These include the heterodimer taste receptor type 1 member 1 (T1r1)+taste receptor type 1 member 3 (T1r3), taste and brain variants of mGluR4 and mGluR1, and calcium sensors. While several studies suggest T1r1+T1r3 is a broadly tuned lLamino acid receptor, little is known about the function of metabotropic glutamate receptors (mGluRs) in L-amino acid taste transduction. Calcium imaging of isolated taste sensory cells (TSCs) of T1r3-GFP and T1r3 knock-out (T1r3 KO) mice was performed using the ratiometric dye Fura 2 AM to investigate the role of different mGluRs in detecting various L-amino acids and inosine 5' monophosphate (IMP). Using agonists selective for various mGluRs such as (RS)-3,5-dihydroxyphenylglycine (DHPG) (an mGluR1 agonist) and L-(+)-2-amino-4-phosphonobutyric acid (l-AP4) (an mGluR4 agonist), we evaluated TSCs to determine if they might respond to these agonists, IMP, and three L-amino acids (monopotassium L-glutamate, L-serine and L-arginine). Additionally, we used selective antagonists against different mGluRs such as (RS)-L-aminoindan-1,5-dicarboxylic acid (AIDA) (an mGluR1 antagonist), and (RS)-α-methylserine-O-phosphate (MSOP) (an mGluR4 antagonist) to determine if they can block responses elicited by these L-amino acids and IMP. We found that L-amino acid- and IMP-responsive cells also responded to each agonist. Antagonists for mGluR4 and mGluR1 significantly blocked the responses elicited by IMP and each of the L-amino acids. Collectively, these data provide evidence for the involvement of taste and brain variants of mGluR1 and mGluR4 in L-amino acid and IMP taste responses in mice, and support the concept that multiple receptors contribute to IMP and L-amino acid taste. PMID:26701297

  7. Metabotropic glutamate receptors are involved in the detection of IMP and L-amino acids by mouse taste sensory cells.

    PubMed

    Pal Choudhuri, S; Delay, R J; Delay, E R

    2016-03-01

    G-protein-coupled receptors are thought to be involved in the detection of umami and L-amino acid taste. These include the heterodimer taste receptor type 1 member 1 (T1r1)+taste receptor type 1 member 3 (T1r3), taste and brain variants of mGluR4 and mGluR1, and calcium sensors. While several studies suggest T1r1+T1r3 is a broadly tuned lLamino acid receptor, little is known about the function of metabotropic glutamate receptors (mGluRs) in L-amino acid taste transduction. Calcium imaging of isolated taste sensory cells (TSCs) of T1r3-GFP and T1r3 knock-out (T1r3 KO) mice was performed using the ratiometric dye Fura 2 AM to investigate the role of different mGluRs in detecting various L-amino acids and inosine 5' monophosphate (IMP). Using agonists selective for various mGluRs such as (RS)-3,5-dihydroxyphenylglycine (DHPG) (an mGluR1 agonist) and L-(+)-2-amino-4-phosphonobutyric acid (l-AP4) (an mGluR4 agonist), we evaluated TSCs to determine if they might respond to these agonists, IMP, and three L-amino acids (monopotassium L-glutamate, L-serine and L-arginine). Additionally, we used selective antagonists against different mGluRs such as (RS)-L-aminoindan-1,5-dicarboxylic acid (AIDA) (an mGluR1 antagonist), and (RS)-α-methylserine-O-phosphate (MSOP) (an mGluR4 antagonist) to determine if they can block responses elicited by these L-amino acids and IMP. We found that L-amino acid- and IMP-responsive cells also responded to each agonist. Antagonists for mGluR4 and mGluR1 significantly blocked the responses elicited by IMP and each of the L-amino acids. Collectively, these data provide evidence for the involvement of taste and brain variants of mGluR1 and mGluR4 in L-amino acid and IMP taste responses in mice, and support the concept that multiple receptors contribute to IMP and L-amino acid taste.

  8. Novel therapeutic targets in osteoarthritis: Narrative review on knock-out genes involved in disease development in mouse animal models.

    PubMed

    Veronesi, Francesca; Della Bella, Elena; Cepollaro, Simona; Brogini, Silvia; Martini, Lucia; Fini, Milena

    2016-05-01

    Osteoarthritis (OA) can affect every joint, especially the knee. Given the complexity of this pathology, OA is difficult to treat with current therapies, which only relieve pain and inflammation and are not capable of restoring tissues once OA has started. Currently, researchers focus on finding a therapeutic strategy that may help to arrest disease progression. The present narrative review gives an overview of the genes involved in the development and progression of OA, assessing in vivo studies performed in knock-out mice affected by OA, to suggest new therapeutic strategies. The article search was performed on the PubMed database and www.webofknowledge.com website with the following keywords: "knee osteoarthritis" AND "knockout mice". The included studies were in English and published from 2005 to 2015. Additional papers were found within the references of the selected articles. In the 55 analyzed in vivo studies, genes mainly affected chondrocyte homeostasis, inflammatory processes, extracellular matrix and the relationship between obesity and OA. Genes are defined as inducing, preventing and not influencing OA. This review shows that joint homeostasis depends on a variety of genetic factors, and preventing or restoring the loss of a gene encoding for protective proteins, or inhibiting the expression of proteins that induce OA, might be a potential therapeutic approach. However, conclusions cannot be drawn because of the wide variability concerning the technique used for OA induction, the role of the genes, the method for tissue evaluations and the lack of assessments of all joint tissues.

  9. Obesity alters the gustatory perception of lipids in the mouse: plausible involvement of lingual CD36[S

    PubMed Central

    Chevrot, Michael; Bernard, Arnaud; Ancel, Déborah; Buttet, Marjorie; Martin, Céline; Abdoul-Azize, Souleymane; Merlin, Jean-François; Poirier, Hélène; Niot, Isabelle; Khan, Naim Akhtar; Passilly-Degrace, Patricia; Besnard, Philippe

    2013-01-01

    A relationship between orosensory detection of dietary lipids, regulation of fat intake, and body mass index was recently suggested. However, involved mechanisms are poorly understood. Moreover, whether obesity can directly modulate preference for fatty foods remains unknown. To address this question, exploration of the oral lipid sensing system was undertaken in diet-induced obese (DIO) mice. By using a combination of biochemical, physiological, and behavioral approaches, we found that i) the attraction for lipids is decreased in obese mice, ii) this behavioral change has an orosensory origin, iii) it is reversed in calorie-restricted DIO mice, revealing an inverse correlation between fat preference and adipose tissue size, iv) obesity suppresses the lipid-mediated downregulation of the lipid-sensor CD36 in circumvallate papillae, usually found during the refeeding of lean mice, and v) the CD36-dependent signaling cascade controlling the intracellular calcium levels ([Ca2+]i) in taste bud cells is decreased in obese mice. Therefore, obesity alters the lipid-sensing system responsible for the oral perception of dietary lipids. This phenomenon seems to take place through a CD36-mediated mechanism, leading to changes in eating behavior. PMID:23840049

  10. Novel therapeutic targets in osteoarthritis: Narrative review on knock-out genes involved in disease development in mouse animal models.

    PubMed

    Veronesi, Francesca; Della Bella, Elena; Cepollaro, Simona; Brogini, Silvia; Martini, Lucia; Fini, Milena

    2016-05-01

    Osteoarthritis (OA) can affect every joint, especially the knee. Given the complexity of this pathology, OA is difficult to treat with current therapies, which only relieve pain and inflammation and are not capable of restoring tissues once OA has started. Currently, researchers focus on finding a therapeutic strategy that may help to arrest disease progression. The present narrative review gives an overview of the genes involved in the development and progression of OA, assessing in vivo studies performed in knock-out mice affected by OA, to suggest new therapeutic strategies. The article search was performed on the PubMed database and www.webofknowledge.com website with the following keywords: "knee osteoarthritis" AND "knockout mice". The included studies were in English and published from 2005 to 2015. Additional papers were found within the references of the selected articles. In the 55 analyzed in vivo studies, genes mainly affected chondrocyte homeostasis, inflammatory processes, extracellular matrix and the relationship between obesity and OA. Genes are defined as inducing, preventing and not influencing OA. This review shows that joint homeostasis depends on a variety of genetic factors, and preventing or restoring the loss of a gene encoding for protective proteins, or inhibiting the expression of proteins that induce OA, might be a potential therapeutic approach. However, conclusions cannot be drawn because of the wide variability concerning the technique used for OA induction, the role of the genes, the method for tissue evaluations and the lack of assessments of all joint tissues. PMID:27059198

  11. Antinociceptive effects of dehydrocorydaline in mouse models of inflammatory pain involve the opioid receptor and inflammatory cytokines

    PubMed Central

    Yin, Zhi-Yu; Li, Lu; Chu, Shuai-Shuai; Sun, Qing; Ma, Zheng-Liang; Gu, Xiao-Ping

    2016-01-01

    Dehydrocorydaline (DHC) is an alkaloidal component isolated from Rhizoma corydalis. Previous studies have shown that DHC has anti-inflammatory and anti-tumor effects and that it can protect the cardiovascular system. However, there are few studies of the antinociceptive effects of DHC in vivo. This study explored the antinociceptive effects and possible mechanisms of DHC in mice using two inflammatory pain models: the acetic acid-induced writhing test and the formalin paw test. The intraperitoneal administration of DHC (3.6, 6 or 10 mg/kg) showed a dose-dependent antinociceptive effect in the acetic acid-induced writhing test and significantly attenuated the formalin-induced pain responses in mice. The antinociceptive effects of DHC were not associated with changes in the locomotor activity or motor responses of animals, and no obvious acute or chronic toxic effects were observed in the mice. Furthermore, the use of naloxone confirmed the involvement of the opioid receptor in the central antinociceptive effects of DHC. DHC reduced formalin-induced paw edema, which indicated that DHC may produce an anti-inflammatory effect in the periphery. In the formalin test, DHC decreased the expression of caspase 6 (CASP6), TNF-α, IL-1β and IL-6 proteins in the spinal cord. These findings confirm that DHC has antinociceptive effects in mice. PMID:27272194

  12. Biochemical evidence for the tyrosine involvement in cationic intermediate stabilization in mouse β-carotene 15, 15'-monooxygenase

    PubMed Central

    2009-01-01

    Background β-carotene 15,15'-monooxygenase (BCMO1) catalyzes the crucial first step in vitamin A biosynthesis in animals. We wished to explore the possibility that a carbocation intermediate is formed during the cleavage reaction of BCMO1, as is seen for many isoprenoid biosynthesis enzymes, and to determine which residues in the substrate binding cleft are necessary for catalytic and substrate binding activity. To test this hypothesis, we replaced substrate cleft aromatic and acidic residues by site-directed mutagenesis. Enzymatic activity was measured in vitro using His-tag purified proteins and in vivo in a β-carotene-accumulating E. coli system. Results Our assays show that mutation of either Y235 or Y326 to leucine (no cation-π stabilization) significantly impairs the catalytic activity of the enzyme. Moreover, mutation of Y326 to glutamine (predicted to destabilize a putative carbocation) almost eliminates activity (9.3% of wt activity). However, replacement of these same tyrosines with phenylalanine or tryptophan does not significantly impair activity, indicating that aromaticity at these residues is crucial. Mutations of two other aromatic residues in the binding cleft of BCMO1, F51 and W454, to either another aromatic residue or to leucine do not influence the catalytic activity of the enzyme. Our ab initio model of BCMO1 with β-carotene mounted supports a mechanism involving cation-π stabilization by Y235 and Y326. Conclusions Our data are consistent with the formation of a substrate carbocation intermediate and cation-π stabilization of this intermediate by two aromatic residues in the substrate-binding cleft of BCMO1. PMID:20003456

  13. Genetic and molecular analysis of chlorambucil-induced germ-line mutations in the mouse

    SciTech Connect

    Rinchik, E.M.; Bangham, J.W.; Hunsicker, P.R.; Cacheiro, N.L.A.; Russell, L.B. ); Kwon, B.S. ); Jackson, I.J. )

    1990-02-01

    Eighteen variants recovered from specific locus mutation rate experiments involving the mutagen chlorambucil were subjected to several genetic and molecular analyses. Most mutations were found to be homozygous lethal. Because lethality is often presumptive evidence for multilocus-deletion events, 10 mutations were analyzed by Southern blot analysis with probes at, or closely linked to, several of the specific locus test markers, namely, albino (c), brown (b), and dilute (d). All eight mutations (two c; three b; two d; and one dilute-short ear (Df(d se))) that arose in post-spermatogonial germ cells were deleted for DNA sequences. No evidence for deletion of two d-se region probes was obtained for the remaining two d mutations that arose in stem-cell spermatogonia. Six of the primary mutants also produced low litter sizes (semisterility). Karyotypic analysis has, to date, confirmed the presence of reciprocal translocations in four of the six. The high frequency of deletions and translocations among the mutations induced in post-spermatogonial stages by chlorambucil, combined with its overall high efficiency in inducing mutations in these stages, should make chlorambucil mutagenesis useful for generating experimentally valuable germ-line deletions throughout the mouse genome.

  14. The Histone Demethylase FBXL10 Regulates the Proliferation of Spermatogonia and Ensures Long-Term Sustainable Spermatogenesis in Mice.

    PubMed

    Ozawa, Manabu; Fukuda, Tsuyoshi; Sakamoto, Reiko; Honda, Hiroaki; Yoshida, Nobuaki

    2016-04-01

    The F-box and leucine-rich repeat protein 10 (Fbxl10) gene encodes a protein that catalyzes demethylation of H3K4 and H3K36. In this study, we show the important roles of FBXL10 as a histone demethylase in sustainable sperm production using mice in which the JmjC domain of Fbxl10 was deleted (Fbxl10(DeltaJ/DeltaJ)). In histological analysis, testis sections from 10-wk-old Fbxl10(DeltaJ/DeltaJ) mice appeared normal. On the other hand, testes from 7-mo-old Fbxl10(DeltaJ/DeltaJ) mice contained a greater ratio of seminiferous tubules exhibiting degeneration of spermatogenesis. Further analysis using an in vitro spermatogonia culture system, that is, germline stem cells (GSCs), revealed that Fbxl10(DeltaJ/DeltaJ) GSCs expressed a significantly higher level of P21 and P19 mRNA, cyclin-dependent kinase inhibitors and also known as cellular senescence markers, than wild-type (WT) GSCs. Furthermore, the ratio of Fbxl10(DeltaJ/DeltaJ) GSCs in G0/G1 phase was higher and the ratios in S and G2/M phases were lower than the corresponding ratios of WT GSCs, and the doubling speed of Fbxl10(DeltaJ/DeltaJ) GSCs was significantly slower than that of WT GSCs. In addition to these in vitro results, an in vivo study indicated that recovery of spermatogenesis after a transient reduction in the number of testicular germ cells by busulfan treatment was significantly slower in Fbxl10(DeltaJ/DeltaJ) mice than in WT mice. These data suggest that Fbxl10 plays important roles in long-term sustainable spermatogenesis via regulating cell cycle. PMID:26984996

  15. Comparison of the genetic effects of equimolar doses of ENU and MNU: While the chemicals differ dramatically in their mutagenicity in stem-cell spermatogonia, both elicit very high mutation rates in differentiating spermatogonia

    SciTech Connect

    Russell, Liane B; Hunsicker, Patricia R; Russell, William

    2007-03-01

    Mutagenoic, reproductive, and toxicity effects of two closely related chemicals, ethylnitrosourea (ENU) and methylnitrosourea (MNU), were compared at equimolar and near-equimolar doses in the mouse specific-locus test in a screen of all stages of spermatogenesis and spermiogenesis. In stem cell spermatogonial (SG), ENU is more than an order of magnitude more mutagenic than MNU. During post-SG stages, both chemicals exhibit high peaks in mutation yield when differentiating spermatogonial (DG) and preleptotene spermatocytes are exposed. The mutation frequency induced by 75 mg MNU/kg during this peak interval is, to date, the highest induced by any single- xposure mutagenic treatment chemical or radiation that allows survival of the exposed animal and its germ cells, producing an estimated 10 new mutations per genome. There is thus a vast difference between stem cell and differentiating spermatogonial in their sensitivity to MNU, but little difference between these stages in their sensitivity to ENU. During stages following meiotic metaphase, the highest mutation yield is obtained from exposed spermatids, but for both chemicals, that yield is less than one-quarter that obtained from the peak interval. Large-lesion (LL) mutations were induced only in spermatids. Although only a few of the remaining mutations were analyzed molecularly, there is considerable evidence from recent molecular characterizations of the marker genes and their flanking chromosomal regions that most, if not all, mutations induced during the peak-sensitive period did not involve lesions outside the marked loci. Both ENU and MNU treatments of post-SG stages yielded significant numbers of mutants that were recovered as mosaics, with the proportion being higher for ENU than for MNU. Comparing the chemicals for the endpoints studied and additional ones (e.g., chromosome aberrations, toxicity to germ cells and to animals, teratogenicity) revealed that while MNU is generally more effective, the opposite

  16. Human chorionic gonadotropin (hCG) fails to stimulate gonocyte differentiation in newborn mouse testes in organ culture.

    PubMed

    Zhou, B; Hutson, J M

    1995-02-01

    The primitive germ cells within the postnatal testis differentiate through several steps to form primary spermatocytes, which are required for postpubertal spermatogenesis. It is postulated that this germ cell development is controlled by the hypothalamic-pituitary-gonadal axis. To study the effect of human chorionic gonadotropin (hCG) on postnatal testicular germ cell differentiation, especially on gonocytes, newborn mouse testes (n = 75) were cultured for 7 days in vitro. The synthetic serum-free medium contained either hCG or exogenous human recombinant müllerian inhibiting substance (MIS) plus transferrin, insulin and retinoic acid (TIRA). Fetal calf serum 10% (FCS) was used for control medium. Thirty-eight newborn mouse testes were cultured with hCG (0.1 to 2.0 IU/ml). The percentages of differentiated type-A spermatogonia were not significantly increased compared with synthetic medium alone (NS, p > 0.05), but were only 5 to 7% (p < 0.001) of that of serum-containing medium. By contrast, normal transformation from gonocytes to type-A spermatogonia occurred in newborn mouse testes (n = 10) cultured with serum-free medium containing exogenous MIS. The percentage of differentiated type-A spermatogonia was approximately 77% (NS, p > 0.05) of those seen in serum-containing medium. These findings demonstrate that the transformation of gonocytes to type-A spermatogonia is regulated by MIS, rather than hCG. As early germ cell transformation is deficient in boys with cryptorchidism, the role of hCG in the treatment of infertility associated with undescended testes should be reevaluated.

  17. Tissue distribution of 2-(2-nitro-4-trifluoromethylbenzoyl)-cyclohexane-1,3-dione (NTBC) and its effect on enzymes involved in tyrosine catabolism in the mouse.

    PubMed

    Lock, E A; Gaskin, P; Ellis, M K; McLean Provan, W; Robinson, M; Smith, L L

    2000-04-01

    Administration of a single oral dose of 2-(2-nitro-4-trifluoromethyl-benzoyl)-cyclohexane-1,3-dione (NTBC) to mice increases the concentration of tyrosine in the plasma and aqueous humour. The tyrosinaemia is both time and dose-dependent with a single dose of 30 micromol NTBC/kg (10 mg/kg) producing maximal concentrations of tyrosine in plasma of about 1200 nmol/ml and in aqueous humour of about 2200 nmol/ml at 16 h after dosing. Analysis of the key hepatic enzymes involved in tyrosine catabolism, following a single dose of 30 micromol NTBC/kg, showed that 4-hydroxyphenylpyruvate dioxygenase (HPPD) was markedly inhibited soon after dosing and that the activity recovered very slowly. In response to the tyrosinaemia, the activity of hepatic tyrosine aminotransferase (TAT) was induced about two-fold, while the activity of hepatic homogentisic acid oxidase (HGO) was reduced at 4 and 5 days after dosing. Daily oral administration of NTBC at doses up to 480 micromol NTBC/kg (160mg/kg/day) to mice produced a maximal tyrosinaemia of about 600-700nmol/ml plasma, showing some adaptation relative to a single dose. Unlike the rat, no treatment-related corneal lesions of the eye were seen at any dose levels up to 6 weeks. Administration of a single oral dose of [14C]-NTBC at 30 micromol/kg led to selective retention of radiolabel in the liver and to a lesser extent the kidneys. Our studies show that NTBC is a potent inhibitor of mouse liver HPPD, which following repeat exposure produces a marked and persistent tyrosinaemia, which does not result in ocular toxicity.

  18. Amyloid Core Formed of Full-Length Recombinant Mouse Prion Protein Involves Sequence 127–143 but Not Sequence 107–126

    PubMed Central

    Chatterjee, Biswanath; Lee, Chung-Yu; Lin, Chen; Chen, Eric H.-L.; Huang, Chao-Li; Yang, Chien-Chih; Chen, Rita P.-Y.

    2013-01-01

    The principal event underlying the development of prion disease is the conversion of soluble cellular prion protein (PrPC) into its disease-causing isoform, PrPSc. This conversion is associated with a marked change in secondary structure from predominantly α-helical to a high β-sheet content, ultimately leading to the formation of aggregates consisting of ordered fibrillar assemblies referred to as amyloid. In vitro, recombinant prion proteins and short prion peptides from various species have been shown to form amyloid under various conditions and it has been proposed that, theoretically, any protein and peptide could form amyloid under appropriate conditions. To identify the peptide segment involved in the amyloid core formed from recombinant full-length mouse prion protein mPrP(23–230), we carried out seed-induced amyloid formation from recombinant prion protein in the presence of seeds generated from the short prion peptides mPrP(107–143), mPrP(107–126), and mPrP(127–143). Our results showed that the amyloid fibrils formed from mPrP(107–143) and mPrP(127–143), but not those formed from mPrP(107–126), were able to seed the amyloidogenesis of mPrP(23–230), showing that the segment residing in sequence 127–143 was used to form the amyloid core in the fibrillization of mPrP(23–230). PMID:23844138

  19. Expression of genes involved in mouse lung cell differentiation/regulation after acute exposure to photons and protons with or without low-dose preirradiation.

    PubMed

    Tian, Jian; Zhao, WeiLing; Tian, Sisi; Slater, James M; Deng, Zhiyong; Gridley, Daila S

    2011-11-01

    The goal of this study was to compare the effects of acute 2 Gy irradiation with photons (0.8 Gy/min) or protons (0.9 Gy/min), both with and without pre-exposure to low-dose/low-dose-rate γ rays (0.01 Gy at 0.03 cGy/h), on 84 genes involved in stem cell differentiation or regulation in mouse lungs on days 21 and 56. Genes with a ≥1.5-fold difference in expression and P < 0.05 compared to 0 Gy controls are emphasized. Two proteins specific for lung stem cells/progenitors responsible for local tissue repair were also compared. Overall, striking differences were present between protons and photons in modulating the genes. More genes were affected by protons than by photons (22 compared to 2 and 6 compared to 2 on day 21 and day 56, respectively) compared to 0 Gy. Preirradiation with low-dose-rate γ rays enhanced the acute photon-induced gene modulation on day 21 (11 compared to 2), and all 11 genes were significantly downregulated on day 56. On day 21, seven genes (aldh2, bmp2, cdc2a, col1a1, dll1, foxa2 and notch1) were upregulated in response to most of the radiation regimens. Immunoreactivity of Clara cell secretory protein was enhanced by all radiation regimens. The number of alveolar type 2 cells positive for prosurfactant protein C in irradiated groups was higher on day 56 (12.4-14.6 cells/100) than on day 21 (8.5-11.2 cells/100) (P < 0.05). Taken together, these results showed that acute photons and protons induced different gene expression profiles in the lungs and that pre-exposure to low-dose-rate γ rays sometimes had modulatory effects. In addition, proteins associated with lung-specific stem cells/progenitors were highly sensitive to radiation.

  20. Induction of reciprocal translocations in rhesus monkey stem-cell spermatogonia: effects of low doses and low dose rates

    SciTech Connect

    van Buul, P.P.; Richardson, J.F. Jr.; Goudzwaard, J.H.

    1986-01-01

    The induction of reciprocal translocation in rhesus monkey spermatogonial stem cells was studied following exposure to low doses of acute X rays (0.25 Gy, 300 mGy/min) or to low-dose-rate X rays (1 Gy, 2 mGy/min) and gamma rays (1 Gy, 0.2 mGy/min). The results obtained at 0.25 Gy of X rays fitted exactly the linear extrapolation down from the 0.5 and 1.0 Gy points obtained earlier. Extension of X-ray exposure reduced the yield of translocations similar to that in the mouse by about 50%. The reduction to 40% of translocation rate after chronic gamma exposure was clearly less than the value of about 80% reported for the mouse over the same range of dose rates. Differential cell killing with ensuing differential elimination of aberration-carrying cells is the most likely explanation for the differences between mouse and monkey.

  1. Differential involvement of 5-HT(1A) and 5-HT(1B/1D) receptors in human interferon-alpha-induced immobility in the mouse forced swimming test.

    PubMed

    Zhang, Hongmei; Wang, Wei; Jiang, Zhenzhou; Shang, Jing; Zhang, Luyong

    2010-01-01

    Although Interferon-alpha (IFN-alpha, CAS 9008-11-1) is a powerful drug in treating several viral infections and certain tumors, a considerable amount of neuropsychiatric side-effects such as depression and anxiety are an unavoidable consequence. Combination with the selective serotonin (5-HT) reuptake inhibitor (SSRI) fluoxetine (CAS 56296-78-7) significantly improved the situation. However, the potential 5-HT(1A) receptor- and 5-HT(1B) receptor-signals involved in the antidepressant effects are still unclear. The effects of 5-HT(1A) receptor- and 5-HT(1B) receptor signals were analyzed by using the mouse forced swimming test (FST), a predictive test of antidepressant-like action. The present results indicated that (1) fluoxetine (administrated intragastrically, 30 mg/kg; not subactive dose: 15 mg/kg) significantly reduced IFN-alpha-induced increase of the immobility time in the forced swimming test; (2) 5-HT(1A) receptor- and 5-HT(1B) receptor ligands alone or in combination had no effects on IFN-alpha-induced increase of the immobility time in the FST; (3) surprisingly, WAY 100635 (5-HT(1A) receptor antagonist, 634908-75-1) and 8-OH-DPAT(5-HT(1A) receptor agonist, CAS 78950-78-4) markedly enhanced the antidepressant effect of fluoxetine at the subactive dose (15 mg/kg, i. g.) on the IFN-alpha-treated mice in the FST. Further investigations showed that fluoxetine combined with WAY 100635 and 8-OH-DPAT failed to produce antidepressant effects in the FST. (4) Co-application of CGS 12066A (5-HT(1B) receptor agonist, CAS 109028-09-3) or GR 127935 (5-HT(1B/1D) receptor antagonist, CAS 148642-42-6) with fluoxetine had no synergistic effects on the IFN-alpha-induced increase of immobility time in FST. (5) Interestingly, co-administration of GR 127935, WAY 100635 and fluoxetine significantly reduced the IFN-alpha-induced increase in immobility time of FST, being more effective than co-administration of WAY 100635 and fluoxetine. All results suggest that (1) compared to

  2. Behavioral and omics analyses study on potential involvement of dipeptide balenine through supplementation in diet of senescence-accelerated mouse prone 8.

    PubMed

    Wada, Nobuhiro; Yamanaka, Satoru; Shibato, Junko; Rakwal, Randeep; Hirako, Satoshi; Iizuka, Yuzuru; Kim, Hyounju; Matsumoto, Akiyo; Kimura, Ai; Takenoya, Fumiko; Yasunaga, Genta; Shioda, Seiji

    2016-12-01

    This study investigates effects of dipeptide balenine, as a major component of whale meat extract (hereafter, WME), supplementation on senescence-accelerated mouse prone 8 (SAMP8), an Alzheimer's disease (AD) model at level of learning and memory formation and brain expression profiles genome-wide in brain. Mice fed experimental balenine (+ WME) supplemented diet for 26 weeks were subjected to four behavioral tests - open field, Y-maze, new object recognition, and water-filled multiple T-maze - to examine effects on learning and memory. Brain transcriptome of SAMP8 mice-fed the WME diet over control low-safflower oil (LSO) diet-fed mice was delineated on a 4 × 44 K mouse whole genome DNA microarray chip. Results revealed the WME diet not only induced improvements in the learning and memory formation but also positively modulated changes in the brain of the SAMP8 mouse; the gene inventories are publically available for analysis by the scientific community. Interestingly, the SAMP8 mouse model presented many genetic characteristics of AD, and numerous novel molecules (Slc2a5, Treh, Fbp1, Aldob, Ppp1r1a, DNase1, Agxt2l1, Cyp2e1, Acsm1, Acsm2, and Pah) were revealed over the SAMR1 (senescence-accelerated mouse resistant 1) mouse, to be oppositely regulated/recovered under the balenine (+ WME) supplemented diet regime by DNA microarray and bioinformatics analyses. Our present study demonstrates an experimental strategy to understand the effects of dipeptide balenine, prominetly contained in meat diet, on SAMP8, providing new insight into whole brain transcriptome changes genome-wide. The gene expression data has been deposited into the Gene Expression Omnibus (GEO): GSE76459. The data will be a valuable resource in examining the effects of natural products, and which could also serve as a human model for further functional analysis and investigation. PMID:27672559

  3. Toxicogenomic Dissection of the Perfluorooctanoic Acid Transcript Profile in Mouse Liver: Evidence for the Involvement of Nuclear Receptors PPARα and CAR

    EPA Science Inventory

    A number of perfluorinated alkyl acids including perfluorooctanoic acid (PFOA) elicit effects similar to peroxisome proliferator chemicals (PPC) in mouse and rat liver. There is strong evidence that PPC cause many of their effects linked to liver cancer through the nuclear recep...

  4. Toxicogenomic Dissection of the Perfluorooctanoic Acid Transcript Profile in Mouse Liver: Evidence for Involvement of the Nuclear Receptors PPARα and CAR

    EPA Science Inventory

    A number of perfluorinated alkyl acids including perfluorooctanoic acid (PFOA) elicit effects similar to peroxisome proliferator chemicals (PPC) in mouse and rat liver. There is strong evidence that PPC cause many of their effects related to liver carcinogenesis through the nucle...

  5. Mendel: a simple excel workbook to compare the observed and expected distributions of genotypes/phenotypes in transgenic and knockout mouse crosses involving up to three unlinked loci by means of a χ2 test.

    PubMed

    Montoliu, Lluís

    2012-06-01

    The analysis of transgenic and knockout mice always involves the establishment of matings with individuals carrying different loci, segregating independently, whose presence is expected among the progeny, according to a Mendelian distribution. The appearance of distorted inheritance ratios suggests the existence of unexpected lethal or sub-lethal phenotypes associated with some genotypes. These situations are common in a number of cases, including: testing transgenic founder mice for germ-line transmission of their transgenes; setting up heterozygous crosses to obtain homozygous individuals, both for transgenic and knockout mice; establishing matings between floxed mouse lines and suitable cre transgenic mouse lines, etc. The Pearson's χ(2) test can be used to assess the significance of the observed frequencies of genotypes/phenotypes in relation to the expected values, in order to determine whether the observed cases fit the expected distribution. Here, I describe a simple Excel workbook to compare the observed and expected distributions of genotypes/phenotypes in transgenic and knockout mouse crosses involving up to three unlinked loci by means of a χ(2) test. The file is freely available for download from my laboratory's web page at: http://www.cnb.csic.es/~montoliu/Mendel.xls .

  6. A new mouse anti-mouse complement receptor type 2 and 1 (CR2/CR1) monoclonal antibody as a tool to study receptor involvement in chronic models of immune responses and disease

    PubMed Central

    Kulik, Liudmila; Hewitt, Finnegan B.; Willis, Van C.; Rodriguez, Rosa; Tomlinson, Stephen; Holers, V. Michael

    2014-01-01

    Although reagents are available to block mouse complement receptor type 2 and/or type1 (CR2/CR1, CD21/CD35) function in acute or short term models of human disease, a mouse anti-rat antibody response limits their use in chronic models. We have addressed this problem by generating in Cr2−/− mice a mouse monoclonal antibody (mAb 4B2) to mouse CR2/CR1. The binding of murine mAb 4B2 to CR2/CR1 directly blocked C3dg (C3d) ligand binding. In vivo injection of mAb 4B2 induced substantial down regulation of CR2 and CR1 from the B cell surface, an effect that lasted six weeks after a single injection of 2 mg of mAb. The 4B2 mAb was studied in vivo for the capability to affect immunological responses to model antigens. Pre-injection of mAb 4B2 before immunization of C57BL/6 mice reduced the IgG1 antibody response to the T-dependent antigen sheep red blood cells (SRBC) to a level comparable to that found in Cr2−/− mice. We also used the collagen-induced arthritis (CIA) model, a CR2/CR1-dependent autoimmune disease model, and found that mice pre-injected with mAb 4B2 demonstrated substantially reduced levels of pathogenic IgG2a antibodies to both the bovine type II collagen (CII) used to induce arthritis and to endogenous mouse CII. Consistent with this result, mice pre-injected with mAb 4B2 demonstrated only very mild arthritis. This reduction in disease, together with published data in CII-immunized Cr2−/− mice, confirm both that the arthritis development depends on CR2/CR1 receptors and that mAb 4B2 can be used to induce biologically relevant receptor blockade. Thus mAb 4B2 is an excellent candidate for use in chronic murine models to determine how receptor blockage at different points modifies disease activity and autoantibody responses. PMID:25457881

  7. Severe neural tube defects in the loop-tail mouse result from mutation of Lpp1, a novel gene involved in floor plate specification.

    PubMed

    Murdoch, J N; Doudney, K; Paternotte, C; Copp, A J; Stanier, P

    2001-10-15

    Neural tube defects (NTD) are clinically important congenital malformations whose molecular mechanisms are poorly understood. The loop-tail (Lp) mutant mouse provides a model for the most severe NTD, craniorachischisis, in which the brain and spinal cord remain open. During a positional cloning approach, we have identified a mutation in a novel gene, Lpp1, in the Lp mouse, providing a strong candidate for the genetic causation of craniorachischisis in LP: Lpp1 encodes a protein of 521 amino acids, with four transmembrane domains related to the Drosophila protein strabismus/van gogh (vang). The human orthologue, LPP1, shares 89% identity with the mouse gene at the nucleotide level and 99% identity at the amino acid level. Lpp1 is expressed in the ventral part of the developing neural tube, but is excluded from the floor plate where Sonic hedgehog (Shh) is expressed. Embryos lacking Shh express Lpp1 throughout the ventral neural tube, suggesting negative regulation of Lpp1 by SHH: Our findings suggest that the mutual interaction between Lpp1 and Shh may define the lateral boundary of floor plate differentiation. Loss of Lpp1 function disrupts neurulation by permitting more extensive floor plate induction by Shh, thereby inhibiting midline bending of the neural plate during initiation of neurulation.

  8. Functional Comparison of Mx1 from Two Different Mouse Species Reveals the Involvement of Loop L4 in the Antiviral Activity against Influenza A Viruses

    PubMed Central

    Verhelst, Judith; Spitaels, Jan; Nürnberger, Cindy; De Vlieger, Dorien; Ysenbaert, Tine; Staeheli, Peter; Fiers, Walter

    2015-01-01

    ABSTRACT The interferon-induced Mx1 gene is an important part of the mammalian defense against influenza viruses. Mus musculus Mx1 inhibits influenza A virus replication and transcription by suppressing the polymerase activity of viral ribonucleoproteins (vRNPs). Here, we compared the anti-influenza virus activity of Mx1 from Mus musculus A2G with that of its ortholog from Mus spretus. We found that the antiviral activity of M. spretus Mx1 was less potent than that of M. musculus Mx1. Comparison of the M. musculus Mx1 sequence with the M. spretus Mx1 sequence revealed 25 amino acid differences, over half of which were present in the GTPase domain and 2 of which were present in loop L4. However, the in vitro GTPase activity of Mx1 from the two mouse species was similar. Replacement of one of the residues in loop L4 in M. spretus Mx1 by the corresponding residue of A2G Mx1 increased its antiviral activity. We also show that deletion of loop L4 prevented the binding of Mx1 to influenza A virus nucleoprotein and, hence, abolished the antiviral activity of mouse Mx1. These results indicate that loop L4 of mouse Mx1 is a determinant of antiviral activity. Our findings suggest that Mx proteins from different mammals use a common mechanism to inhibit influenza A viruses. IMPORTANCE Mx proteins are evolutionarily conserved in vertebrates and inhibit a wide range of viruses. Still, the exact details of their antiviral mechanisms remain largely unknown. Functional comparison of the Mx genes from two species that diverged relatively recently in evolution can provide novel insights into these mechanisms. We show that both Mus musculus A2G Mx1 and Mus spretus Mx1 target the influenza virus nucleoprotein. We also found that loop L4 in mouse Mx1 is crucial for its antiviral activity, as was recently reported for primate MxA. This indicates that human and mouse Mx proteins, which have diverged by 75 million years of evolution, recognize and inhibit influenza A viruses by a common

  9. Kinetochore Fibers Are Not Involved in the Formation of the First Meiotic Spindle in Mouse Oocytes, but Control the Exit from the First Meiotic M Phase

    PubMed Central

    Brunet, Stéphane; Maria, Angélica Santa; Guillaud, Philippe; Dujardin, Denis; Kubiak, Jacek Z.; Maro, Bernard

    1999-01-01

    During meiosis, two successive divisions occur without any intermediate S phase to produce haploid gametes. The first meiotic division is unique in that homologous chromosomes are segregated while the cohesion between sister chromatids is maintained, resulting in a reductional division. Moreover, the duration of the first meiotic M phase is usually prolonged when compared with mitotic M phases lasting 8 h in mouse oocytes. We investigated the spindle assembly pathway and its role in the progression of the first meiotic M phase in mouse oocytes. During the first 4 h, a bipolar spindle forms and the chromosomes congress near the equatorial plane of the spindle without stable kinetochore– microtubule end interactions. This late prometaphase spindle is then maintained for 4 h with chromosomes oscillating in the central region of the spindle. The kinetochore–microtubule end interactions are set up at the end of the first meiotic M phase (8 h after entry into M phase). This event allows the final alignment of the chromosomes and exit from metaphase. The continuous presence of the prometaphase spindle is not required for progression of the first meiotic M phase. Finally, the ability of kinetochores to interact with microtubules is acquired at the end of the first meiotic M phase and determines the timing of polar body extrusion. PMID:10402455

  10. Early synaptic deficits in the APP/PS1 mouse model of Alzheimer's disease involve neuronal adenosine A2A receptors

    PubMed Central

    Viana da Silva, Silvia; Haberl, Matthias Georg; Zhang, Pei; Bethge, Philipp; Lemos, Cristina; Gonçalves, Nélio; Gorlewicz, Adam; Malezieux, Meryl; Gonçalves, Francisco Q.; Grosjean, Noëlle; Blanchet, Christophe; Frick, Andreas; Nägerl, U Valentin; Cunha, Rodrigo A.; Mulle, Christophe

    2016-01-01

    Synaptic plasticity in the autoassociative network of recurrent connections among hippocampal CA3 pyramidal cells is thought to enable the storage of episodic memory. Impaired episodic memory is an early manifestation of cognitive deficits in Alzheimer's disease (AD). In the APP/PS1 mouse model of AD amyloidosis, we show that associative long-term synaptic potentiation (LTP) is abolished in CA3 pyramidal cells at an early stage. This is caused by activation of upregulated neuronal adenosine A2A receptors (A2AR) rather than by dysregulation of NMDAR signalling or altered dendritic spine morphology. Neutralization of A2AR by acute pharmacological inhibition, or downregulation driven by shRNA interference in a single postsynaptic neuron restore associative CA3 LTP. Accordingly, treatment with A2AR antagonists reverts one-trial memory deficits. These results provide mechanistic support to encourage testing the therapeutic efficacy of A2AR antagonists in early AD patients. PMID:27312972

  11. Synergistic effect of high glucose and ANG II on proliferation of mouse embryonic stem cells: involvement of PKC and MAPKs as well as AT1 receptor.

    PubMed

    Kim, Yun Hee; Han, Ho Jae

    2008-05-01

    This study examined the synergistic effect of high glucose levels and ANG II on proliferation and its related signal pathways using mouse embryonic stem (ES) cells. The combined use of a high glucose concentration (25 mM) and ANG II increased the level of [3H]thymidine/BrdU incorporation, and the number of cells compared with either treatment alone. Each treatment with high glucose or ANG II increased the cell population in the S phase compared with control, and the combined treatment of a high glucose concentration and ANG II significantly increased the number of cells in the S phase according to FACS analysis. Moreover, the high glucose-induced increase in [3H]thymidine incorporation was blocked by inhibiting the ANG II type 1 (AT1) receptor. The combined high glucose and ANG II significantly increased the STAT3 phosphorylation compared with high glucose or ANG II alone. ANG II stimulated the influx of Ca2+ in 25 mM glucose compared with 5 mM glucose. High glucose levels increase the level of PKC alpha, epsilon, and zeta translocation from the cytosol to the membrane fraction. In an examination of other signal pathways, the combined treatment significantly increased the level of p44/42, p38 MAPKs phosphorylation compared with either treatment alone. Indeed, the combined treatment increased the mRNA expression level of the protooncogenes and cell cycle regulatory proteins. In conclusion, the combined treatment of a high glucose concentration and ANG II had a synergistic effect in stimulating mouse ES cell proliferation through the Ca2+/PKC, MAPKs, and the AT1 receptor.

  12. Involvement of Phospholipase Cγ1 in Mouse Egg Activation Induced by a Truncated Form of the C-kit Tyrosine Kinase Present in Spermatozoa

    PubMed Central

    Sette, Claudio; Bevilacqua, Arturo; Geremia, Raffaele; Rossi, Pellegrino

    1998-01-01

    Microinjection of a truncated form of the c-kit tyrosine kinase present in mouse spermatozoa (tr-kit) activates mouse eggs parthenogenetically, and tr-kit– induced egg activation is inhibited by preincubation with an inhibitor of phospholipase C (PLC) (Sette, C., A. Bevilacqua, A. Bianchini, F. Mangia, R. Geremia, and P. Rossi. 1997. Development [Camb.]. 124:2267–2274). Co-injection of glutathione-S-transferase (GST) fusion proteins containing the src-homology (SH) domains of the γ1 isoform of PLC (PLCγ1) competitively inhibits tr-kit– induced egg activation. A GST fusion protein containing the SH3 domain of PLCγ1 inhibits egg activation as efficiently as the whole SH region, while a GST fusion protein containing the two SH2 domains is much less effective. A GST fusion protein containing the SH3 domain of the Grb2 adaptor protein does not inhibit tr-kit–induced egg activation, showing that the effect of the SH3 domain of PLCγ1 is specific. Tr-kit–induced egg activation is also suppressed by co-injection of antibodies raised against the PLCγ1 SH domains, but not against the PLCγ1 COOH-terminal region. In transfected COS cells, coexpression of PLCγ1 and tr-kit increases diacylglycerol and inositol phosphate production, and the phosphotyrosine content of PLCγ1 with respect to cells expressing PLCγ1 alone. These data indicate that tr-kit activates PLCγ1, and that the SH3 domain of PLCγ1 is essential for tr-kit–induced egg activation. PMID:9722617

  13. Interleukin 1β Regulation of the System xc- Substrate-specific Subunit, xCT, in Primary Mouse Astrocytes Involves the RNA-binding Protein HuR.

    PubMed

    Shi, Jingxue; He, Yan; Hewett, Sandra J; Hewett, James A

    2016-01-22

    System xc(-) is a heteromeric amino acid cystine/glutamate antiporter that is constitutively expressed by cells of the CNS, where it functions in the maintenance of intracellular glutathione and extracellular glutamate levels. We recently determined that the cytokine, IL-1β, increases the activity of system xc(-) in CNS astrocytes secondary to an up-regulation of its substrate-specific light chain, xCT, and that this occurs, in part, at the level of transcription. However, an in silico analysis of the murine xCT 3'-UTR identified numerous copies of adenine- and uridine-rich elements, raising the possibility that undefined trans-acting factors governing mRNA stability and translation may also contribute to xCT expression. Here we show that IL-1β increases the level of mRNA encoding xCT in primary cultures of astrocytes isolated from mouse cortex in association with an increase in xCT mRNA half-life. Additionally, IL-1β induces HuR translocation from the nucleus to the cytoplasm. RNA immunoprecipitation analysis reveals that HuR binds directly to the 3'-UTR of xCT in an IL-1β-dependent manner. Knockdown of endogenous HuR protein abrogates the IL-1β-mediated increase in xCT mRNA half-life, whereas overexpression of HuR in unstimulated primary mouse astrocytes doubles the half-life of constitutive xCT mRNA. This latter effect is accompanied by an increase in xCT protein levels, as well as a functional increase in system xc(-) activity. Altogether, these data support a critical role for HuR in mediating the IL-1β-induced stabilization of astrocyte xCT mRNA.

  14. Effects of realgar on GSH synthesis in the mouse hippocampus: Involvement of system XAG(-), system XC(-), MRP-1 and Nrf2.

    PubMed

    Wang, Yanlei; Chen, Mo; Zhang, Yinghua; Huo, Taoguang; Fang, Ying; Jiao, Xuexin; Yuan, Mingmei; Jiang, Hong

    2016-10-01

    Realgar is a type of mineral drug that contains arsenic and has neurotoxicity. Glutathione (GSH), which is the main antioxidant in the central nervous system, plays a key role in antioxidant defenses and the detoxification of arsenic. However, whether realgar interferes with the synthesis of GSH in the brain and the molecular mechanisms underlying its effects are largely unknown. Here, we used mouse models of exposure to realgar to show that realgar affects the synthesis of GSH in the hippocampus, leading to ultrastructural changes in hippocampal neurons and synapses and deficiencies in cognitive abilities, and that the mechanisms that cause this effect may be associated with alterations in the expression of system XAG(-), system XC(-), multidrug resistance-associated protein 1(MRP-1), nuclear factor E2-related factor 2 (Nrf2), γ-glutamylcysteine synthetase (γ-GCS), and the levels of glutamate (Glu) and cysteine (Cys) in the extracellular fluid. These findings provide a theoretical basis for preventing the drug-induced chronic arsenic poisoning in the nervous system that is triggered by realgar. PMID:27412851

  15. Effects of realgar on GSH synthesis in the mouse hippocampus: Involvement of system XAG(-), system XC(-), MRP-1 and Nrf2.

    PubMed

    Wang, Yanlei; Chen, Mo; Zhang, Yinghua; Huo, Taoguang; Fang, Ying; Jiao, Xuexin; Yuan, Mingmei; Jiang, Hong

    2016-10-01

    Realgar is a type of mineral drug that contains arsenic and has neurotoxicity. Glutathione (GSH), which is the main antioxidant in the central nervous system, plays a key role in antioxidant defenses and the detoxification of arsenic. However, whether realgar interferes with the synthesis of GSH in the brain and the molecular mechanisms underlying its effects are largely unknown. Here, we used mouse models of exposure to realgar to show that realgar affects the synthesis of GSH in the hippocampus, leading to ultrastructural changes in hippocampal neurons and synapses and deficiencies in cognitive abilities, and that the mechanisms that cause this effect may be associated with alterations in the expression of system XAG(-), system XC(-), multidrug resistance-associated protein 1(MRP-1), nuclear factor E2-related factor 2 (Nrf2), γ-glutamylcysteine synthetase (γ-GCS), and the levels of glutamate (Glu) and cysteine (Cys) in the extracellular fluid. These findings provide a theoretical basis for preventing the drug-induced chronic arsenic poisoning in the nervous system that is triggered by realgar.

  16. The laminin binding protein p40 is involved in inducing limb abnormality of mouse fetuses as the effects of methoxyacetic acid treatment.

    PubMed

    Ruyani, Aceng; Sudarwati, Sri; Sutasurya, Lien A; Sumarsono, Sony H; Gloe, Torsten

    2003-09-01

    This study is intended to characterize a protein that is linked with mouse limb teratogenicity as the effects of methoxyacetic acid (MAA) treatment. A single dose of MAA (10 mmol/kg body weight) was given by gavage on gestation day (GD) 11, whereas the control group were administered vehicle only. The pregnant mice were killed at 4 h after MAA treatment, and forelimb buds were isolated from both the control and treated group embryos. Proteins from forelimb buds GD 11 + 4 h, which were precipitated out using 40-60% ammonium sulfate, then were analyzed by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (2-D SDS-PAGE) technique. The 2-D gels reveal one protein with 41.6 kDa and pI 6.4, which expression was downregulated after MAA treatment. Tentative protein identification via peptide mass database search and definitive protein identification via a primary sequence database search indicate that the protein matches exactly to 34/67 kDa laminin binding protein (LBP; P14206, SwissProt), which is encoded by p40 gene (MGI:105381). The identity was further verified by Western blotting with an antibody against the 67 kDa LBP. The results suggest that MAA treatment to pregnant mice downregulates the LBP-p40 in the forelimb buds.

  17. Neuroprotective Effects of Salidroside in the MPTP Mouse Model of Parkinson's Disease: Involvement of the PI3K/Akt/GSK3β Pathway

    PubMed Central

    He, Hong; Song, Hujie; Zhao, Junjie; Li, Tao; Wu, Leitao

    2016-01-01

    The degenerative loss through apoptosis of dopaminergic neurons in the substantia nigra pars compacta plays a primary role in the progression of Parkinson's disease (PD). Our in vitro experiments suggested that salidroside (Sal) could protect against 1-methyl-4-phenylpyridine-induced cell apoptosis in part by regulating the PI3K/Akt/GSK3β pathway. The current study aims to increase our understanding of the protective mechanisms of Sal in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropypridine- (MPTP-) induced PD mouse model. We found that pretreatment with Sal could protect against MPTP-induced increase of the time of turning downwards and climbing down to the floor. Sal also prevented MPTP-induced decrease of locomotion frequency and the increase of the immobile time. Sal provided a protection of in MPTP-induced loss of tyrosine hydroxylase-positive neurons in SNpc and the level of DA, DOPAC, and HVA in the striatum. Furthermore, Sal could increase the phosphorylation level of Akt and GSK3β, upregulate the ratio of Bcl-2/Bax, and inhibit the activation of caspase-3, caspase-6, and caspase-9. These results show that Sal prevents the loss of dopaminergic neurons and the PI3K/Akt/GSK3β pathway signaling pathway may have mediated the protection of Sal against MPTP, suggesting that Sal may be a potential candidate in neuroprotective treatment for PD. PMID:27738547

  18. BNC1 is required for maintaining mouse spermatogenesis

    PubMed Central

    Zhang, Xiaohong; Chou, Weichin; Haig-Ladewig, Lisa; Zeng, Wenxian; Cao, Wenlei; Gerton, George; Dobrinski, Ina; Tseng, Hung

    2012-01-01

    Basonuclin (BNC1) is a zinc finger protein expressed primarily in gametogenic cells and proliferative keratinocytes. Our previous work suggested that BNC1 is present in spermatogonia, spermatocytes and spermatids, but absent in the Sertoli cells. BNC1's role in spermatogenesis is unknown. Here we show that BNC1 is required for the maintenance of spermatogenesis. Bnc1-null male mice were sub-fertile, losing germ cells progressively with age. The Bnc1-null seminiferous epithelia began to degenerate before 8 weeks of age and eventually became Sertoli cell-only. Sperm count and motility also declined with age. Furthermore, Bnc1 heterozygotes, although fertile, showed a significant drop in sperm count and in testis weight by 24 weeks of age, suggesting a dosage effect of Bnc1 on testis development. In conclusion, our data demonstrate for the first time BNC1's essential role in maintaining mouse spermatogenesis. PMID:22266914

  19. NRG1 and KITL signal downstream of retinoic acid in the germline to support soma-free syncytial growth of differentiating spermatogonia

    PubMed Central

    Chapman, KM; Medrano, GA; Chaudhary, J; Hamra, FK

    2015-01-01

    Defined culture systems supporting spermatogonial differentiation will provide experimental platforms to study spermatogenesis. However, germline-intrinsic signaling mechanisms sufficient to support spermatogonial differentiation without somatic cells remain largely undefined. Here we analyzed EGF superfamily receptor and ligand diversity in rat testis cells and delineated germline-intrinsic signaling via an ERBB3 co-transducer, ERBB2, as essential for retinoic acid-induced syncytial growth by differentiating spermatogonia. Similar to the ERBB2/3 agonist NRG1, we found that KIT Ligand (KITL) robustly supported spermatogonial differentiation without serum or somatic cells. ERBB2 inhibitors failed to disrupt KITL-dependent spermatogonial development, and KITL prevented ERBB3-deficient spermatogonial degeneration upon differentiation. Thus we report that NRG1 and KITL activate alternative pathways downstream of retinoic acid signaling in the germline that are essential for stem cells to undergo premeiotic steps of spermatogenesis in culture. Robust serum/soma-free spermatogonial differentiation opens new doors to study mammalian germ cell biology in culture and to discover factors that can drive meiotic progression in vitro. PMID:26500786

  20. Extracellular Ca2+-induced force restoration in K+-depressed skeletal muscle of the mouse involves an elevation of [K+]i: implications for fatigue

    PubMed Central

    Leader, John P.; Loiselle, Denis S.; Higgins, Amanda; Lin, Wei; Renaud, Jean-Marc

    2015-01-01

    We examined whether a Ca2+-K+ interaction was a potential mechanism operating during fatigue with repeated tetani in isolated mouse muscles. Raising the extracellular Ca2+ concentration ([Ca2+]o) from 1.3 to 10 mM in K+-depressed slow-twitch soleus and/or fast-twitch extensor digitorum longus muscles caused the following: 1) increase of intracellular K+ activity by 20–60 mM (raised intracellular K+ content, unchanged intracellular fluid volume), so that the K+-equilibrium potential increased by ∼10 mV and resting membrane potential repolarized by 5–10 mV; 2) large restoration of action potential amplitude (16–54 mV); 3) considerable recovery of excitable fibers (∼50% total); and 4) restoration of peak force with the peak tetanic force-extracellular K+ concentration ([K+]o) relationship shifting rightward toward higher [K+]o. Double-sigmoid curve-fitting to fatigue profiles (125 Hz for 500 ms, every second for 100 s) showed that prior exposure to raised [K+]o (7 mM) increased, whereas lowered [K+]o (2 mM) decreased, the rate and extent of force loss during the late phase of fatigue (second sigmoid) in soleus, hence implying a K+ dependence for late fatigue. Prior exposure to 10 mM [Ca2+]o slowed late fatigue in both muscle types, but was without effect on the extent of fatigue. These combined findings support our notion that a Ca2+-K+ interaction is plausible during severe fatigue in both muscle types. We speculate that a diminished transsarcolemmal K+ gradient and lowered [Ca2+]o contribute to late fatigue through reduced action potential amplitude and excitability. The raised [Ca2+]o-induced slowing of fatigue is likely to be mediated by a higher intracellular K+ activity, which prolongs the time before stimulation-induced K+ efflux depolarizes the sarcolemma sufficiently to interfere with action potentials. PMID:25571990

  1. Involvement of calyculin A inhibitable protein phosphatases in the cyclic AMP signal transduction pathway of mouse corticotroph tumour (AtT20) cells

    PubMed Central

    Antaraki, A; Ang, K L; Antoni, F A

    1997-01-01

    The role of non-calcineurin protein phosphatases in the cyclic AMP signal transduction pathway was examined in mouse pituitary corticotroph tumour (AtT20) cells. Blockers of protein phosphatases, calyculin A and okadaic acid, were applied in AtT20 cells depleted of rapidly mobilizable pools of intracellular calcium and activated by various cyclic AMP generating agonists. Inhibitors of cyclic nucleotide phosphodiesterases were present throughout. The accumulation of cyclic AMP was monitored by radioimmunoassay, phosphodiesterase activity in cell homogenates was measured by radiometric assay. Neither calyculin A nor okadaic acid altered basal cyclic AMP levels but cyclic AMP formation induced by 41 amino acid residue corticotrophin releasing-factor (CRF) was strongly inhibited (up to 80%). 1-Norokadaone was inactive. Similar data were also obtained when isoprenaline or pituitary adenylate cyclase activating peptide1–38 were used as agonists. Pertussis toxin did not modify the inhibition of CRF-induced cyclic AMP production by calyculin A. Pretreatment with calyculin A completely prevented the stimulation of cyclic AMP formation by cholera toxin even in the presence of 0.5 mM isobutylmethylxanthine (IBMX) and 0.1 mM rolipram. Cholera toxin mediated ADP-ribosylation of the 45K and 52K molecular weight Gsα isoforms in membranes from calyculin A-pretreated cells was enhanced to 150–200% when compared with controls. Cholera toxin-induced cyclic AMP was reduced by calyculin A within 10 min when calyculin A was applied after a 90 min pretreatment with cholera toxin. Under these conditions the effect of calyculin A could be blocked by the combination of 0.5 mM IBMX and 0.1 mM rolipram, but not by 0.5 mM IBMX alone. Phosphodiesterase activity in AtT20 cell homogenates showed a significant, 2.7 fold increase after treatment with calyculin A. In control cells phosphodiesterase activity was blocked by 80% in the presence of IBMX (0.5 mM), or IBMX plus

  2. Identification of a locus on mouse chromosome 3 involved in differential susceptibility to Theiler's murine encephalomyelitis virus-induced demyelinating disease.

    PubMed Central

    Melvold, R W; Jokinen, D M; Miller, S D; Dal Canto, M C; Lipton, H L

    1990-01-01

    Theiler's virus-induced demyelinating disease results from a chronic infection in the white matter of the central nervous system and provides an excellent model for human multiple sclerosis. Like multiple sclerosis, there are genetic risk factors in disease development, including genes associated with the major histocompatibility complex and with those encoding the beta chain of the T-cell receptor. Comparisons of the susceptible DBA/2 and resistant C57BL/6 strains have indicated an important role for the H-2D locus and for a non-H-2 gene (not involving the beta chain of the T-cell receptor) in differential susceptibility. In the present report, analysis of recombinant-inbred strains (BXD) between the DBA/2 and C57BL/6 strains indicated that this non-H-2 locus is located at the centromeric end of chromosome 3 near (4 +/- 4 centimorgans) the carbonic anhydrase-2 (Car-2) enzyme locus. PMID:2296080

  3. Protease-Activated Receptor (PAR)2, but Not PAR1, Is Involved in Collateral Formation and Anti-Inflammatory Monocyte Polarization in a Mouse Hind Limb Ischemia Model

    PubMed Central

    Nossent, Anne Yael; van Oeveren-Rietdijk, Annemarie M.; de Vries, Margreet R.; Spek, C. Arnold; van Zonneveld, Anton Jan; Reitsma, Pieter H.; Hamming, Jaap F.; de Boer, Hetty C.; Versteeg, Henri H.; Quax, Paul H. A.

    2013-01-01

    Aims In collateral development (i.e. arteriogenesis), mononuclear cells are important and exist as a heterogeneous population consisting of pro-inflammatory and anti-inflammatory/repair-associated cells. Protease-activated receptor (PAR)1 and PAR2 are G-protein-coupled receptors that are both expressed by mononuclear cells and are involved in pro-inflammatory reactions, while PAR2 also plays a role in repair-associated responses. Here, we investigated the physiological role of PAR1 and PAR2 in arteriogenesis in a murine hind limb ischemia model. Methods and Results PAR1-deficient (PAR1-/-), PAR2-deficient (PAR2-/-) and wild-type (WT) mice underwent femoral artery ligation. Laser Doppler measurements revealed reduced post-ischemic blood flow recovery in PAR2-/- hind limbs when compared to WT, while PAR1-/- mice were not affected. Upon ischemia, reduced numbers of smooth muscle actin (SMA)-positive collaterals and CD31-positive capillaries were found in PAR2-/- mice when compared to WT mice, whereas these parameters in PAR1-/- mice did not differ from WT mice. The pool of circulating repair-associated (Ly6C-low) monocytes and the number of repair-associated (CD206-positive) macrophages surrounding collaterals in the hind limbs were increased in WT and PAR1-/- mice, but unaffected in PAR2-/- mice. The number of repair-associated macrophages in PAR2-/- hind limbs correlated with CD11b- and CD115-expression on the circulating monocytes in these animals, suggesting that monocyte extravasation and M-CSF-dependent differentiation into repair-associated cells are hampered. Conclusion PAR2, but not PAR1, is involved in arteriogenesis and promotes the repair-associated response in ischemic tissues. Therefore, PAR2 potentially forms a new pro-arteriogenic target in coronary artery disease (CAD) patients. PMID:23637930

  4. Relationship between Rgs2 gene expression level and anxiety and depression-like behaviour in a mutant mouse model: serotonergic involvement.

    PubMed

    Lifschytz, Tzuri; Broner, Esther Channah; Zozulinsky, Polina; Slonimsky, Alexandra; Eitan, Renana; Greenbaum, Lior; Lerer, Bernard

    2012-10-01

    RGS2 is a member of a family of proteins that negatively modulate G-protein coupled receptor transmission. Variations in the RGS2 gene were found to be associated in humans with anxious and depressive phenotypes. We sought to study the relationship of Rgs2 expression level to depression and anxiety-like behavioural features, sociability and brain 5-HT1A and 5-HT1B receptor expression. We studied male mice carrying a mutation that causes lower Rgs2 gene expression, employing mice heterozygous (Het) or homozygous (Hom) for this mutation, or wild-type (WT). Mice were subjected to behavioural tests reflecting depressive-like behaviour [forced swim test (FST), novelty suppressed feeding test (NSFT)], elevated plus maze (EPM) for evaluation of anxiety levels and the three-chamber sociability test. The possible involvement of raphe nucleus 5-HT1A receptors in these behavioural features was examined by 8-OH-DPAT-induced hypothermia. Expression levels of 5-HT1A and 5-HT1B receptors in the cortex, raphe nucleus and hypothalamus were compared among mice of the different Rgs2 genotype groups. NSFT results demonstrated that Hom mice showed more depressive-like features than Rgs2 Het and WT mice. A trend for such a relationship was also suggested by the FST results. EPM and sociability test results showed Hom and Het mice to be more anxious and less sociable than WT mice. In addition Hom and Het mice were characterized by lower basal body temperature and demonstrated less 8-OH-DPAT-induced hypothermia than WT mice. Finally, Hom and Het mice had significantly lower 5-HT1A and 5-HT1B receptor expression levels in the raphe than WT mice. Our findings demonstrate a relationship between Rgs2 gene expression level and a propensity for anxious and depressive-like behaviour and reduced social interaction that may involve changes in serotonergic receptor expression.

  5. Generation and characterization of a transgenic mouse with a functional human TSPY.

    PubMed

    Schubert, S; Skawran, B; Dechend, F; Nayernia, K; Meinhardt, A; Nanda, I; Schmid, M; Engel, W; Schmidtke, J

    2003-09-01

    To generate an animal model that is suitable for the analysis of regulation and expression of human testis-specific protein, Y-encoded TSPY, a transgenic mouse line, TgTSPY9, harboring a complete structural human TSPY gene was generated. Fluorescence in situ hybridization and Southern analyses show that approximately 50 copies of the human TSPY transgene are integrated at a single chromosomal site that maps to the distal long arm of the Y chromosome. The transgene is correctly transcribed and spliced according to the human pattern and is mainly expressed in testicular tissue, with spermatogonia and early primary spermatocytes (leptotene and zygotene) as expressing germ cells. TSPY transgenic mice are phenotypically normal, and spermatogenesis is neither impaired nor enhanced by the human transgene. The present study shows that a human TSPY gene integrated into the mouse genome follows the human expression pattern although murine tspy had lost its function in rodent evolution millions of years ago. PMID:12773407

  6. Effect of diet on expression of genes involved in lipid metabolism, oxidative stress, and inflammation in mouse liver-insights into mechanisms of hepatic steatosis.

    PubMed

    Renaud, Helen J; Cui, Julia Y; Lu, Hong; Klaassen, Curtis D

    2014-01-01

    Nutritional intake is a fundamental determinant of health. Many studies have correlated excess caloric intake, as well as a high ratio of n-6:n-3 fatty acids, with detrimental health outcomes, such as the metabolic syndrome. In contrast, low-calorie diets have beneficial health effects. Despite these associations, our understanding of the causal relationship between diet and health remains largely elusive. The present study examined the molecular changes elicited by nine diets with varying fat, sugar, cholesterol, omega-3 fatty acids, omega-6 fatty acids, and calories in C57BL/6 male mice. Microarray analyses were conducted on liver samples from three mice per diet and detected 20,449 genes of which 3,734 were responsive to changes in dietary components. Principal component analysis showed that diet restriction correlated the least with the other diets and also affected more genes than any other diet. Interestingly, Gene Set Enrichment Analysis (GSEA) identified gene sets involved in glutathione metabolism, immune response, fatty acid metabolism, cholesterol metabolism, ABC transporters, and oxidative phosphorylation as being highly responsive to changes in diet composition. On the gene level, this study reveals novel findings such as the induction of the drug efflux pump Abcb1a (p-glycoprotein) by diet restriction and an atherogenic diet, as well as the suppression of the rate limiting step of bile acid synthesis, Cyp7a1, by a high fructose diet. This study provides considerable insight into the molecular changes incurred by a variety of diets and furthers our understanding of the causal relationships between diet and health.

  7. STE20/SPS1-Related Proline/Alanine-Rich Kinase Is Involved in Plasticity of GABA Signaling Function in a Mouse Model of Acquired Epilepsy

    PubMed Central

    Zhou, Jueqian; Chen, Shuda; Chen, Yishu; Chen, Ziyi; Wang, Qian; Fang, Ziyan; Zhou, Liemin

    2013-01-01

    The intracellular concentration of chloride ([Cl-]i) determines the strength and polarity of GABA neurotransmission. STE20/SPS1-related proline/alanine-rich kinase (SPAK) is known as an indirect regulator of [Cl-]i for its activation of Na-K-2 Cl-co-transporters (NKCC) and inhibition of K-Cl-co-transporters (KCC) in many organs. NKCC1 or KCC2 expression changes have been demonstrated previously in the hippocampal neurons of mice with pilocarpine-induced status epilepticus (PISE). However, it remains unclear whether SPAK modulates [Cl-]i via NKCC1 or KCC2 in the brain. Also, there are no data clearly characterizing SPAK expression in cortical or hippocampal neurons or confirming an association between SPAK and epilepsy. In the present study, we examined SPAK expression and co-expression with NKCC1 and KCC2 in the hippocampal neurons of mice with PISE, and we investigated alterations in SPAK expression in the hippocampus of such mice. Significant increases in SPAK mRNA and protein levels were detected during various stages of PISE in the PISE mice in comparison to levels in age-matched sham (control) and blank treatment (control) mice. SPAK and NKCC1 expression increased in vitro, while KCC2 was down-regulated in hippocampal neurons following hypoxic conditioning. However, SPAK overexpression did not influence the expression levels of NKCC1 or KCC2. Using co-immunoprecipitation, we determined that the intensity of interaction between SPAK and NKCC1 and between SPAK and KCC2 increased markedly after oxygen-deprivation, whereas SPAK overexpression strengthened the relationships. The [Cl-]i of hippocampal neurons changed in a corresponding manner under the different conditions. Our data suggests that SPAK is involved in the plasticity of GABA signaling function in acquired epilepsy via adjustment of [Cl-]i in hippocampal neurons. PMID:24058604

  8. Pharmacological evidence for the involvement of the NMDA receptor and nitric oxide pathway in the antidepressant-like effect of lamotrigine in the mouse forced swimming test.

    PubMed

    Ostadhadi, Sattar; Ahangari, Mohammad; Nikoui, Vahid; Norouzi-Javidan, Abbas; Zolfaghari, Samira; Jazaeri, Farahnaz; Chamanara, Mohsen; Akbarian, Reyhaneh; Dehpour, Ahmad-Reza

    2016-08-01

    Lamotrigine is an anticonvulsant agent that shows clinical antidepressant properties. The aim of the present study was to investigate the involvement of N-methyl-d-aspartate (NMDA) receptors and nitric oxide-cyclic guanosine monophosphate (NO-cGMP) synthesis in possible antidepressant-like effect of lamotrigine in forced swimming test (FST) in mice. Intraperitoneal administration of lamotrigine (10mg/kg) decreased the immobility time in the FST (P<0.01) without any effect on locomotor activity in the open-field test (OFT), while higher dose of lamotrigine (30mg/kg) reduced the immobility time in the FST (P<0.001) as well as the number of crossings in the OFT. Pretreatment of animals with NMDA (75mg/kg), l-arginine (750mg/kg, a substrate for nitric oxide synthase [NOS]) or sildenafil (5mg/kg, a phosphodiesterase [PDE] 5 inhibitor) reversed the antidepressant-like effect of lamotrigine (10mg/kg) in the FST. Injection of l-nitroarginine methyl ester (l-NAME, 10mg/kg, a non-specific NOS inhibitor), 7-nitroindazole (30mg/kg, a neuronal NOS inhibitor), methylene blue (20mg/kg, an inhibitor of both NOS and soluble guanylate cyclase [sGC]), or MK-801 (0.05mg/kg), ketamine (1mg/kg), and magnesium sulfate (10mg/kg) as NMDA receptor antagonists in combination with a sub-effective dose of lamotrigine (5mg/kg) diminished the immobility time of animals in the FST compared with either drug alone. None of the drugs produced significant effects on the locomotor activity in the OFT. Based on our findings, it is suggested that the antidepressant-like effect of lamotrigine might mediated through inhibition of either NMDA receptors or NO-cGMP synthesis. PMID:27470415

  9. The involvement of NMDA receptor/NO/cGMP pathway in the antidepressant like effects of baclofen in mouse force swimming test.

    PubMed

    Khan, Muhammad Imran; Ostadhadi, Sattar; Zolfaghari, Samira; Ejtemaei Mehr, Shahram; Hassanzadeh, Gholamreza; Dehpour, Ahmad-Reza

    2016-01-26

    In the current study, the involvement of N-methyl-d-aspartate receptor (NMDAR) and nitric oxide (NO)/cyclic guanosine monophosphate (cGMP) system in the antidepressant-like effects of baclofen was evaluated by using animal model in forced swimming test. Followed by an open field test for the evaluation of locomotor activity, the immobility time for mice in force swimming test was recorded. Only the last four min was analyzed. Administration of Baclofen (0.5 and 1mg/kg, i.p.) reduced the immobility interval in the FST. Prior administration of l-arginine (750mg/kg, i.p.,) a nitric oxide synthase substrate or sildenafil (5mg/kg, i.p.) a phosphodiesterase 5 into mice suppressed the antidepressant-like activity of baclofen (1mg/kg, i.p.).Co-treatment of 7-nitroindazole (50mg/kg, i.p.,) an inhibitor of neuronal nitric oxide synthase, L-NAME (10mg/kg, i.p.,) a non-specific inhibitor of nitric oxide synthase or MK-801 (0.05mg/kg, i.p.) an NMDA receptor antagonist with subeffective dose of baclofen (0.1mg/kg, i.p.), reduced the immobility time in the FST as compared to the drugs when used alone. Co-administrated of lower doses of MK-801 (0.01mg/kg) or l-NAME (1mg/kg) failed to effect immobility time however, simultaneous administration of these two agents in same dose with subeffective dose of baclofen (0.1mg/kg, i.p.), minimized the immobility time in the FST. Thus, our results support the role of NMDA receptors and l-arginine-NO-GMP pathway in the antidepressant-like action of baclofen. PMID:26679225

  10. Delta(9)-tetrahydrocannabinol prolongs the immobility time in the mouse forced swim test: involvement of cannabinoid CB(1) receptor and serotonergic system.

    PubMed

    Egashira, Nobuaki; Matsuda, Tomomi; Koushi, Emi; Higashihara, Fuminori; Mishima, Kenichi; Chidori, Shozo; Hasebe, Nobuyoshi; Iwasaki, Katsunori; Nishimura, Ryoji; Oishi, Ryozo; Fujiwara, Michihiro

    2008-07-28

    In the present study, we investigated the effect of Delta(9)-tetrahydrocannabinol (THC), the principal psychoactive component of marijuana, on immobility time during the forced swim test. THC (2 and 6 mg/kg, i.p.) significantly prolonged the immobility time. In addition, THC at the same doses did not significantly affect locomotor activity in the open-field test. The selective cannabinoid CB(1) receptor antagonist rimonabant (3 mg/kg, i.p.) significantly reduced the enhancement of immobility by THC (6 mg/kg). Similarly, the selective serotonin (5-HT) reuptake inhibitor (SSRI) citalopram (10 mg/kg, i.p.) and 5-HT(1A/7) receptor agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT, 0.3 mg/kg, i.p.) significantly reduced this THC-induced effect. Moreover, the selective 5-HT(1A) receptor antagonist N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl) cyclohexane carboxamide dihydrochloride (WAY100635, 1 mg/kg, i.p.) and the postsynaptic 5-HT(1A) receptor antagonist MM-77 (0.1 mg/kg, i.p.) reversed this reduction effect of 8-OH-DPAT (0.3 mg/kg). In contrast, the selective 5-HT(7) receptor antagonist (R)-3-[2-[2-(4-methylpiperidin-1-yl)ethyl]pyrrolidine-1-sulfonyl]phenol hydrochloride (SB269970) had no effect on this reduction effect of 8-OH-DPAT. WAY100635 (1 mg/kg) also reversed the reduction effect of citalopram (10 mg/kg). These findings suggest that the 5-HT(1A) receptors are involved in THC-induced enhancement of immobility.

  11. The involvement of NMDA receptor/NO/cGMP pathway in the antidepressant like effects of baclofen in mouse force swimming test.

    PubMed

    Khan, Muhammad Imran; Ostadhadi, Sattar; Zolfaghari, Samira; Ejtemaei Mehr, Shahram; Hassanzadeh, Gholamreza; Dehpour, Ahmad-Reza

    2016-01-26

    In the current study, the involvement of N-methyl-d-aspartate receptor (NMDAR) and nitric oxide (NO)/cyclic guanosine monophosphate (cGMP) system in the antidepressant-like effects of baclofen was evaluated by using animal model in forced swimming test. Followed by an open field test for the evaluation of locomotor activity, the immobility time for mice in force swimming test was recorded. Only the last four min was analyzed. Administration of Baclofen (0.5 and 1mg/kg, i.p.) reduced the immobility interval in the FST. Prior administration of l-arginine (750mg/kg, i.p.,) a nitric oxide synthase substrate or sildenafil (5mg/kg, i.p.) a phosphodiesterase 5 into mice suppressed the antidepressant-like activity of baclofen (1mg/kg, i.p.).Co-treatment of 7-nitroindazole (50mg/kg, i.p.,) an inhibitor of neuronal nitric oxide synthase, L-NAME (10mg/kg, i.p.,) a non-specific inhibitor of nitric oxide synthase or MK-801 (0.05mg/kg, i.p.) an NMDA receptor antagonist with subeffective dose of baclofen (0.1mg/kg, i.p.), reduced the immobility time in the FST as compared to the drugs when used alone. Co-administrated of lower doses of MK-801 (0.01mg/kg) or l-NAME (1mg/kg) failed to effect immobility time however, simultaneous administration of these two agents in same dose with subeffective dose of baclofen (0.1mg/kg, i.p.), minimized the immobility time in the FST. Thus, our results support the role of NMDA receptors and l-arginine-NO-GMP pathway in the antidepressant-like action of baclofen.

  12. Id4 Marks Spermatogonial Stem Cells in the Mouse Testis.

    PubMed

    Sun, Feng; Xu, Qing; Zhao, Danfeng; Degui Chen, Charlie

    2015-01-01

    Mammalian spermatogenesis is a classic adult stems cell-dependent process, supported by the self-renewal and differentiation of spermatogonial stem cells (SSCs). However, the identification of SSCs and elucidation of their behaviors in undisturbed testis has long been a big challenge. Here, we generated a knock-in mouse model, Id4-2A-CreERT2-2A-tdTomato, which allowed us to mark Id4-expressing (Id4(+)) cells at different time points in situ and track their behaviors across distinct developmental stages during steady-state and regenerating spermatogenesis. We found that Id4(+) cells continue to produce spermatogonia, spermatocytes and sperm in mouse testis, showing they are capable of self-renewal and have differentiation potential. Consistent with these findings, ablation of Id4(+) cells in mice results in a loss of spermatogenesis. Furthermore, developmental fate mapping reveals that Id4(+) SSCs originate from neonate Id4(+) gonocytes. Therefore, our results indicate that Id4 marks spermatogonial stem cells in the mouse testis.

  13. Loss of Vascular Endothelial Growth Factor A (VEGFA) Isoforms in the Testes of Male Mice Causes Subfertility, Reduces Sperm Numbers, and Alters Expression of Genes That Regulate Undifferentiated Spermatogonia

    PubMed Central

    Lu, Ningxia; Sargent, Kevin M.; Clopton, Debra T.; Pohlmeier, William E.; Brauer, Vanessa M.; McFee, Renee M.; Weber, John S.; Ferrara, Napoleone; Silversides, David W.

    2013-01-01

    Vascular endothelial growth factor A (VEGFA) isoform treatment has been demonstrated to alter spermatogonial stem cell homeostasis. Therefore, we generated pDmrt1-Cre;Vegfa−/− (knockout, KO) mice by crossing pDmrt1-Cre mice to floxed Vegfa mice to test whether loss of all VEGFA isoforms in Sertoli and germ cells would impair spermatogenesis. When first mated, KO males took 14 days longer to get control females pregnant (P < .02) and tended to take longer for all subsequent parturition intervals (9 days; P < .07). Heterozygous males sired fewer pups per litter (P < .03) and after the first litter took 10 days longer (P < .05) to impregnate females, suggesting a more progressive loss of fertility. Reproductive organs were collected from 6-month-old male mice. There were fewer sperm per tubule in the corpus epididymides (P < .001) and fewer ZBTB16-stained undifferentiated spermatogonia (P < .003) in the testes of KO males. Testicular mRNA abundance for Bcl2 (P < .02), Bcl2:Bax (P < .02), Neurog3 (P < .007), and Ret was greater (P = .0005), tended to be greater for Sin3a and tended to be reduced for total Foxo1 (P < .07) in KO males. Immunofluorescence for CD31 and VE-Cadherin showed no differences in testis vasculature; however, CD31-positive staining was evident in undifferentiated spermatogonia only in KO testes. Therefore, loss of VEGFA isoforms in Sertoli and germ cells alters genes necessary for long-term maintenance of undifferentiated spermatogonia, ultimately reducing sperm numbers and resulting in subfertility. PMID:24169552

  14. Obesity in Aging Exacerbates Blood–Brain Barrier Disruption, Neuroinflammation, and Oxidative Stress in the Mouse Hippocampus: Effects on Expression of Genes Involved in Beta-Amyloid Generation and Alzheimer’s Disease

    PubMed Central

    Tucsek, Zsuzsanna; Toth, Peter; Sosnowska, Danuta; Gautam, Tripti; Mitschelen, Matthew; Koller, Akos; Szalai, Gabor; Sonntag, William E.; Csiszar, Anna

    2014-01-01

    There is growing evidence that obesity has deleterious effects on the brain and cognitive function in the elderly population. However, the specific mechanisms through which aging and obesity interact to promote cognitive decline remain unclear. To test the hypothesis that aging exacerbates obesity-induced cerebromicrovascular damage and neuroinflammation, we compared young (7 months) and aged (24 months) high fat diet–fed obese C57BL/6 mice. Aging exacerbated obesity-induced systemic inflammation and blood–brain barrier disruption, as indicated by the increased circulating levels of proinflammatory cytokines and increased presence of extravasated immunoglobulin G in the hippocampus, respectively. Obesity-induced blood–brain barrier damage was associated with microglia activation, upregulation of activating Fc-gamma receptors and proinflammatory cytokines, and increased oxidative stress. Treatment of cultured primary microglia with sera derived from aged obese mice resulted in significantly more pronounced microglia activation and oxidative stress, as compared with treatment with young sera. Serum-induced activation and oxidative stress were also exacerbated in primary microglia derived from aged animals. Hippocampal expression of genes involved in regulation of the cellular amyloid precursor protein–dependent signaling pathways, beta-amyloid generation, and the pathogenesis of tauopathy were largely unaffected by obesity in aged mice. Collectively, obesity in aging is associated with a heightened state of systemic inflammation, which exacerbates blood–brain barrier disruption. The resulting neuroinflammation and oxidative stress in the mouse hippocampus likely contribute to the significant cognitive decline observed in aged obese animals. PMID:24269929

  15. Obesity in aging exacerbates blood-brain barrier disruption, neuroinflammation, and oxidative stress in the mouse hippocampus: effects on expression of genes involved in beta-amyloid generation and Alzheimer's disease.

    PubMed

    Tucsek, Zsuzsanna; Toth, Peter; Sosnowska, Danuta; Gautam, Tripti; Mitschelen, Matthew; Koller, Akos; Szalai, Gabor; Sonntag, William E; Ungvari, Zoltan; Csiszar, Anna

    2014-10-01

    There is growing evidence that obesity has deleterious effects on the brain and cognitive function in the elderly population. However, the specific mechanisms through which aging and obesity interact to promote cognitive decline remain unclear. To test the hypothesis that aging exacerbates obesity-induced cerebromicrovascular damage and neuroinflammation, we compared young (7 months) and aged (24 months) high fat diet-fed obese C57BL/6 mice. Aging exacerbated obesity-induced systemic inflammation and blood-brain barrier disruption, as indicated by the increased circulating levels of proinflammatory cytokines and increased presence of extravasated immunoglobulin G in the hippocampus, respectively. Obesity-induced blood-brain barrier damage was associated with microglia activation, upregulation of activating Fc-gamma receptors and proinflammatory cytokines, and increased oxidative stress. Treatment of cultured primary microglia with sera derived from aged obese mice resulted in significantly more pronounced microglia activation and oxidative stress, as compared with treatment with young sera. Serum-induced activation and oxidative stress were also exacerbated in primary microglia derived from aged animals. Hippocampal expression of genes involved in regulation of the cellular amyloid precursor protein-dependent signaling pathways, beta-amyloid generation, and the pathogenesis of tauopathy were largely unaffected by obesity in aged mice. Collectively, obesity in aging is associated with a heightened state of systemic inflammation, which exacerbates blood-brain barrier disruption. The resulting neuroinflammation and oxidative stress in the mouse hippocampus likely contribute to the significant cognitive decline observed in aged obese animals.

  16. Protective effects of 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside in the MPTP-induced mouse model of Parkinson's disease: Involvement of reactive oxygen species-mediated JNK, P38 and mitochondrial pathways.

    PubMed

    He, Hong; Wang, Songhai; Tian, Jiyu; Chen, Lei; Zhang, Wei; Zhao, Junjie; Tang, Haifeng; Zhang, Xiaojun; Chen, Jianzong

    2015-11-15

    Parkinson's disease (PD) is characterized by the selective death of dopaminergic neurons in the substantia nigra pars compacta. Oxidative stress-induced neuron loss is thought to play a crucial role in the pathogenesis of PD. Previous work from our group suggests that 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside (TSG), an active component extracted from a traditional Chinese herb, Polygonum multiflorum thunb, can attenuate 1-methyl-4-phenyl pyridium-induced apoptosis in the neuronal cell line PC12, by inhibiting reactive oxygen species generation and modulating c-Jun N-terminal kinases (JNK) activation. Here, we investigated the protective effects of TSG against 1-methyl-4-phenyl-1,2,3,6-tetrahydropypridine (MPTP)-induced loss of tyrosine hydroxylase positive cells in mice and the underlying mechanisms. The results showed that MPTP-induced loss of tyrosine hydroxylase positive cells and reactive oxygen species generation were prevented by TSG in a dose-dependent manner. The reactive oxygen species scavenger N-acetylcysteine could also mitigate reactive oxygen species generation. Moreover, JNK and P38 were activated by MPTP, but extracellular signal-regulated protein kinases phosphorylation did not change after MPTP treatment. TSG at different doses blocked the activation of JNK and P38. The protective effect of TSG was also associated with downregulation of the bax/bcl-2 ratio, reversed the release of cytochrome c and smac, and inhibited the activation of caspase-3, -6, and -9 induced by MPTP. In conclusion, our studies demonstrated that the protective effects of TSG in the MPTP-induced mouse model of PD are involved, at least in part, in controlling reactive oxygen species-mediated JNK, P38, and mitochondrial pathways.

  17. Protective effects of 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside in the MPTP-induced mouse model of Parkinson's disease: Involvement of reactive oxygen species-mediated JNK, P38 and mitochondrial pathways.

    PubMed

    He, Hong; Wang, Songhai; Tian, Jiyu; Chen, Lei; Zhang, Wei; Zhao, Junjie; Tang, Haifeng; Zhang, Xiaojun; Chen, Jianzong

    2015-11-15

    Parkinson's disease (PD) is characterized by the selective death of dopaminergic neurons in the substantia nigra pars compacta. Oxidative stress-induced neuron loss is thought to play a crucial role in the pathogenesis of PD. Previous work from our group suggests that 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside (TSG), an active component extracted from a traditional Chinese herb, Polygonum multiflorum thunb, can attenuate 1-methyl-4-phenyl pyridium-induced apoptosis in the neuronal cell line PC12, by inhibiting reactive oxygen species generation and modulating c-Jun N-terminal kinases (JNK) activation. Here, we investigated the protective effects of TSG against 1-methyl-4-phenyl-1,2,3,6-tetrahydropypridine (MPTP)-induced loss of tyrosine hydroxylase positive cells in mice and the underlying mechanisms. The results showed that MPTP-induced loss of tyrosine hydroxylase positive cells and reactive oxygen species generation were prevented by TSG in a dose-dependent manner. The reactive oxygen species scavenger N-acetylcysteine could also mitigate reactive oxygen species generation. Moreover, JNK and P38 were activated by MPTP, but extracellular signal-regulated protein kinases phosphorylation did not change after MPTP treatment. TSG at different doses blocked the activation of JNK and P38. The protective effect of TSG was also associated with downregulation of the bax/bcl-2 ratio, reversed the release of cytochrome c and smac, and inhibited the activation of caspase-3, -6, and -9 induced by MPTP. In conclusion, our studies demonstrated that the protective effects of TSG in the MPTP-induced mouse model of PD are involved, at least in part, in controlling reactive oxygen species-mediated JNK, P38, and mitochondrial pathways. PMID:26477638

  18. Quantitative T2 combined with texture analysis of nuclear magnetic resonance images identify different degrees of muscle involvement in three mouse models of muscle dystrophy: mdx, Largemyd and mdx/Largemyd.

    PubMed

    Martins-Bach, Aurea B; Malheiros, Jackeline; Matot, Béatrice; Martins, Poliana C M; Almeida, Camila F; Caldeira, Waldir; Ribeiro, Alberto F; Loureiro de Sousa, Paulo; Azzabou, Noura; Tannús, Alberto; Carlier, Pierre G; Vainzof, Mariz

    2015-01-01

    Quantitative nuclear magnetic resonance imaging (MRI) has been considered a promising non-invasive tool for monitoring therapeutic essays in small size mouse models of muscular dystrophies. Here, we combined MRI (anatomical images and transverse relaxation time constant-T2-measurements) to texture analyses in the study of four mouse strains covering a wide range of dystrophic phenotypes. Two still unexplored mouse models of muscular dystrophies were analyzed: The severely affected Largemyd mouse and the recently generated and worst double mutant mdx/Largemyd mouse, as compared to the mildly affected mdx and normal mice. The results were compared to histopathological findings. MRI showed increased intermuscular fat and higher muscle T2 in the three dystrophic mouse models when compared to the wild-type mice (T2: mdx/Largemyd: 37.6±2.8 ms; mdx: 35.2±4.5 ms; Largemyd: 36.6±4.0 ms; wild-type: 29.1±1.8 ms, p<0.05), in addition to higher muscle T2 in the mdx/Largemyd mice when compared to mdx (p<0.05). The areas with increased muscle T2 in the MRI correlated spatially with the identified histopathological alterations such as necrosis, inflammation, degeneration and regeneration foci. Nevertheless, muscle T2 values were not correlated with the severity of the phenotype in the 3 dystrophic mouse strains, since the severely affected Largemyd showed similar values than both the mild mdx and worst mdx/Largemyd lineages. On the other hand, all studied mouse strains could be unambiguously identified with texture analysis, which reflected the observed differences in the distribution of signals in muscle MRI. Thus, combined T2 intensity maps and texture analysis is a powerful approach for the characterization and differentiation of dystrophic muscles with diverse genotypes and phenotypes. These new findings provide important noninvasive tools in the evaluation of the efficacy of new therapies, and most importantly, can be directly applied in human translational research.

  19. Quantitative T2 Combined with Texture Analysis of Nuclear Magnetic Resonance Images Identify Different Degrees of Muscle Involvement in Three Mouse Models of Muscle Dystrophy: mdx, Largemyd and mdx/Largemyd

    PubMed Central

    Martins-Bach, Aurea B.; Malheiros, Jackeline; Matot, Béatrice; Martins, Poliana C. M.; Almeida, Camila F.; Caldeira, Waldir; Ribeiro, Alberto F.; Loureiro de Sousa, Paulo; Azzabou, Noura; Tannús, Alberto; Carlier, Pierre G.; Vainzof, Mariz

    2015-01-01

    Quantitative nuclear magnetic resonance imaging (MRI) has been considered a promising non-invasive tool for monitoring therapeutic essays in small size mouse models of muscular dystrophies. Here, we combined MRI (anatomical images and transverse relaxation time constant—T2—measurements) to texture analyses in the study of four mouse strains covering a wide range of dystrophic phenotypes. Two still unexplored mouse models of muscular dystrophies were analyzed: The severely affected Largemyd mouse and the recently generated and worst double mutant mdx/Largemyd mouse, as compared to the mildly affected mdx and normal mice. The results were compared to histopathological findings. MRI showed increased intermuscular fat and higher muscle T2 in the three dystrophic mouse models when compared to the wild-type mice (T2: mdx/Largemyd: 37.6±2.8 ms; mdx: 35.2±4.5 ms; Largemyd: 36.6±4.0 ms; wild-type: 29.1±1.8 ms, p<0.05), in addition to higher muscle T2 in the mdx/Largemyd mice when compared to mdx (p<0.05). The areas with increased muscle T2 in the MRI correlated spatially with the identified histopathological alterations such as necrosis, inflammation, degeneration and regeneration foci. Nevertheless, muscle T2 values were not correlated with the severity of the phenotype in the 3 dystrophic mouse strains, since the severely affected Largemyd showed similar values than both the mild mdx and worst mdx/Largemyd lineages. On the other hand, all studied mouse strains could be unambiguously identified with texture analysis, which reflected the observed differences in the distribution of signals in muscle MRI. Thus, combined T2 intensity maps and texture analysis is a powerful approach for the characterization and differentiation of dystrophic muscles with diverse genotypes and phenotypes. These new findings provide important noninvasive tools in the evaluation of the efficacy of new therapies, and most importantly, can be directly applied in human translational research

  20. Broad-spectrum antimicrobial efficacy of peptide A3-APO in mouse models of multidrug-resistant wound and lung infections cannot be explained by in vitro activity against the pathogens involved.

    PubMed

    Ostorhazi, Eszter; Holub, Marianna Csilla; Rozgonyi, Ferenc; Harmos, Ferenc; Cassone, Marco; Wade, John D; Otvos, Laszlo

    2011-05-01

    Although the designer proline-rich antimicrobial peptide A3-APO has only modest activity against Escherichia coli and Acinetobacter baumannii in vitro, in mouse models of systemic and wound infections it shows superior efficacy compared with conventional antibiotics. In this study, the efficacy of A3-APO in several additional mouse models was investigated, including Staphylococcus aureus wound infection, mixed Klebsiella pneumoniae-A. baumannii-Proteus mirabilis wound infection and K. pneumoniae lung infection, mimicking blast wound infections, foot ulcers and ventilator-induced nosocomial infections, respectively. Whilst the peptide practically did not kill the strains in vitro, when administered intramuscularly or as an aerosol it significantly improved mouse survival and reduced bacterial counts at the infection site and in blood. In the lung infection study, the blood bacterial counts following A3-APO treatment were as low as after treatment with colistin and were lower than after treatment with imipenem or amikacin. The wounds of treated animals, unlike their untreated counterparts, lacked pus and signs of inflammation. In human peripheral blood mononuclear cells, A3-APO upregulated the expression of the anti-inflammatory cytokines interleukin-4 and interleukin-10 by four- to six-fold. One of the mechanisms mediating the in vivo protective effects might be the prevention of inflammation around bacterial infiltration.

  1. Bimodal concentration-response of nicotine involves the nicotinic acetylcholine receptor, transient receptor potential vanilloid type 1, and transient receptor potential ankyrin 1 channels in mouse trachea and sensory neurons.

    PubMed

    Kichko, Tatjana I; Lennerz, Jochen; Eberhardt, Mirjam; Babes, Ramona M; Neuhuber, Winfried; Kobal, Gerd; Reeh, Peter W

    2013-11-01

    High concentrations of nicotine, as in the saliva of oral tobacco consumers or in smoking cessation aids, have been shown to sensitize/activate recombinant transient receptor potential vanilloid type 1 (rTRPV1) and mouse TRPA1 (mTRPA1) channels. By measuring stimulated calcitonin gene-related peptide (CGRP) release from the isolated mouse trachea, we established a bimodal concentration-response relationship with a threshold below 10 µM (-)-nicotine, a maximum at 100 µM, an apparent nadir between 0.5 and 10 mM, and a renewed increase at 20 mM. The first peak was unchanged in TRPV1/A1 double-null mutants as compared with wild-types and was abolished by specific nicotinic acetylcholine receptor (nAChR) inhibitors and by camphor, discovered to act as nicotinic antagonist. The nicotine response at 20 mM was strongly pHe-dependent, - five times greater at pH 9.0 than 7.4, indicating that intracellular permeation of the (uncharged) alkaloid was required to reach the TRPV1/A1 binding sites. The response was strongly reduced in both null mutants, and more so in double-null mutants. Upon measuring calcium transients in nodose/jugular and dorsal root ganglion neurons in response to 100 µM nicotine, 48% of the vagal (but only 14% of the somatic) sensory neurons were activated, the latter very weakly. However, nicotine 20 mM at pH 9.0 repeatedly activated almost every single cultured neuron, partly by releasing intracellular calcium and independent of TRPV1/A1 and nAChRs. In conclusion, in mouse tracheal sensory nerves nAChRs are 200-fold more sensitive to nicotine than TRPV1/A1; they are widely coexpressed with the capsaicin receptor among vagal sensory neurons and twice as abundant as TRPA1. Nicotine is the major stimulant in tobacco, and its sensory impact through nAChRs should not be disregarded.

  2. RBE and genetic susceptibility of mouse and rat spermatogonial stem cells to protons, heavy charged particles and 1.5 MeV neutrons

    NASA Astrophysics Data System (ADS)

    Vaglenov, A.; Fedorenko, B.; Kaltenboeck, B.

    The main purpose of the present study is to provide data on RBE and genetic susceptibility in the mouse and the rat when exposed to protons, HZE particles and neutrons. Genetic damage from exposure to 50 MeV and 9 GeV protons, 4 GeV/nucleon helium ions, 4 GeV/nucleon carbon ions and 1.5 MeV neutrons was studied in adult (CBA × C57Bl/6J) F1 mice. Damage from 9 GeV protons and 4 GeV helium ions was studied in adult Wistar rats. The incidence of reciprocal translocations (RT) induced in the spermatogonial stem cells of each species was recorded. RBE values were derived by comparing linear regression coefficients from dose-responses within the same dose-range for each of the radiation types tested and 60Co γ-rays or by means of a direct nonparametric method. RT yields measured after mouse and rat spermatogonial irradiation with protons, heavy charged particles and neutrons fit the linear model of the dose-response relationship. Relative to 60Co γ-rays, RBE values are as follows for mouse spermatogonia: 0.9 for 50 MeV protons; 1.3 for 9 GeV protons; 0.7 for 4 GeV helium ions; and 1.3 for 4 GeV carbon ions. For rat spermatogonia, values were: 1.7 for 9 GeV protons and 1.3 for helium ions. Compared to mice irradiated using the same experimental design, rats were more susceptible to high-LET radiations, with susceptibility assessed by genetic damage to their spermatogonial stem cells. The RBE of 1.5 MeV neutron is about 6.6.

  3. Mouse models of myasthenia gravis.

    PubMed

    Ban, Joanne; Phillips, William D

    2015-01-01

    Myasthenia gravis is a muscle weakness disease characterized by autoantibodies that target components of the neuromuscular junction, impairing synaptic transmission. The most common form of myasthenia gravis involves antibodies that bind the nicotinic acetylcholine receptors in the postsynaptic membrane. Many of the remaining cases are due to antibodies against muscle specific tyrosine kinase (MuSK). Recently, autoantibodies against LRP4 (another component of the MuSK signaling complex in the postsynaptic membrane) were identified as the likely cause of myasthenia gravis in some patients. Fatiguing weakness is the common symptom in all forms of myasthenia gravis, but muscles of the body are differentially affected, for reasons that are not fully understood. Much of what we have learnt about the immunological and neurobiological aspects of the pathogenesis derives from mouse models. The most widely used mouse models involve either passive transfer of autoantibodies, or active immunization of the mouse with acetylcholine receptors or MuSK protein. These models can provide a robust replication of many of the features of the human disease. Depending upon the protocol, acute fatiguing weakness develops 2 - 14 days after the start of autoantibody injections (passive transfer) or might require repeated immunizations over several weeks (active models). Here we review mouse models of myasthenia gravis, including what they have contributed to current understanding of the pathogenic mechanisms and their current application to the testing of therapeutics.

  4. Unique subcellular distribution of phosphorylated Plk1 (Ser137 and Thr210) in mouse oocytes during meiotic division and pPlk1(Ser137) involvement in spindle formation and REC8 cleavage.

    PubMed

    Du, Juan; Cao, Yan; Wang, Qian; Zhang, Nana; Liu, Xiaoyu; Chen, Dandan; Liu, Xiaoyun; Xu, Qunyuan; Ma, Wei

    2015-01-01

    Polo-like kinase 1 (Plk1) is pivotal for proper mitotic progression, its targeting activity is regulated by precise subcellular positioning and phosphorylation. Here we assessed the protein expression, subcellular localization and possible functions of phosphorylated Plk1 (pPlk1(Ser137) and pPlk1(Thr210)) in mouse oocytes during meiotic division. Western blot analysis revealed a peptide of pPlk1(Ser137) with high and stable expression from germinal vesicle (GV) until metaphase II (MII), while pPlk1(Thr210) was detected as one large single band at GV stage and 2 small bands after germinal vesicle breakdown (GVBD), which maintained stable up to MII. Immunofluorescence analysis showed pPlk1(Ser137) was colocalized with microtubule organizing center (MTOC) proteins, γ-tubulin and pericentrin, on spindle poles, concomitantly with persistent concentration at centromeres and dynamic aggregation between chromosome arms. Differently, pPlk1(Thr210) was persistently distributed across the whole body of chromosomes after meiotic resumption. The specific Plk1 inhibitor, BI2536, repressed pPlk1(Ser137) accumulation at MTOCs and between chromosome arms, consequently disturbed γ-tubulin and pericentrin recruiting to MTOCs, destroyed meiotic spindle formation, and delayed REC8 cleavage, therefore arresting oocytes at metaphase I (MI) with chromosome misalignment. BI2536 completely reversed the premature degradation of REC8 and precocious segregation of chromosomes induced with okadaic acid (OA), an inhibitor to protein phosphatase 2A. Additionally, the protein levels of pPlk1(Ser137) and pPlk1(Thr210), as well as the subcellular distribution of pPlk1(Thr210), were not affected by BI2536. Taken together, our results demonstrate that Plk1 activity is required for meiotic spindle assembly and REC8 cleavage, with pPlk1(Ser137) is the action executor, in mouse oocytes during meiotic division. PMID:26654596

  5. Homologs of the Xenopus developmental gene DG42 are present in zebrafish and mouse and are involved in the synthesis of Nod-like chitin oligosaccharides during early embryogenesis.

    PubMed

    Semino, C E; Specht, C A; Raimondi, A; Robbins, P W

    1996-05-14

    The Xenopus developmental gene DG42 is expressed during early embryonic development, between the midblastula and neurulation stages. The deduced protein sequence of Xenopus DG42 shows similarity to Rhizobium Nod C, Streptococcus Has A, and fungal chitin synthases. Previously, we found that the DG42 protein made in an in vitro transcription/translation system catalyzed synthesis of an array of chitin oligosaccharides. Here we show that cell extracts from early Xenopus and zebrafish embryos also synthesize chitooligosaccharides. cDNA fragments homologous to DG42 from zebrafish and mouse were also cloned and sequenced. Expression of these homologs was similar to that described for Xenopus based on Northern and Western blot analysis. The Xenopus anti-DG42 antibody recognized a 63-kDa protein in extracts from zebrafish embryos that followed a similar developmental expression pattern to that previously described for Xenopus. The chitin oligosaccharide synthase activity found in extracts was inactivated by a specific DG42 antibody; synthesis of hyaluronic acid (HA) was not affected under the conditions tested. Other experiments demonstrate that expression of DG42 under plasmid control in mouse 3T3 cells gives rise to chitooligosaccharide synthase activity without an increase in HA synthase level. A possible relationship between our results and those of other investigators, which show stimulation of HA synthesis by DG42 in mammalian cell culture systems, is provided by structural analyses to be published elsewhere that suggest that chitin oligosaccharides are present at the reducing ends of HA chains. Since in at least one vertebrate system hyaluronic acid formation can be inhibited by a pure chitinase, it seems possible that chitin oligosaccharides serve as primers for hyaluronic acid synthesis.

  6. On Involvement.

    ERIC Educational Resources Information Center

    Greene, Michael B.

    Involvement Ratings In Settings (IRIS), a multi-dimensional non-verbal scale of involvement adaptable to a time-sampling method of data collection, was constructed with the aid of the videotapes of second-grade Follow Through classrooms made by CCEP. Scales were defined through observations of involved and alienated behavior, and the IRIS was…

  7. Ataxia with loss of Purkinje cells in a mouse model for Refsum disease.

    PubMed

    Ferdinandusse, Sacha; Zomer, Anna W M; Komen, Jasper C; van den Brink, Christina E; Thanos, Melissa; Hamers, Frank P T; Wanders, Ronald J A; van der Saag, Paul T; Poll-The, Bwee Tien; Brites, Pedro

    2008-11-18

    Refsum disease is caused by a deficiency of phytanoyl-CoA hydroxylase (PHYH), the first enzyme of the peroxisomal alpha-oxidation system, resulting in the accumulation of the branched-chain fatty acid phytanic acid. The main clinical symptoms are polyneuropathy, cerebellar ataxia, and retinitis pigmentosa. To study the pathogenesis of Refsum disease, we generated and characterized a Phyh knockout mouse. We studied the pathological effects of phytanic acid accumulation in Phyh(-/-) mice fed a diet supplemented with phytol, the precursor of phytanic acid. Phytanic acid accumulation caused a reduction in body weight, hepatic steatosis, and testicular atrophy with loss of spermatogonia. Phenotype assessment using the SHIRPA protocol and subsequent automated gait analysis using the CatWalk system revealed unsteady gait with strongly reduced paw print area for both fore- and hindpaws and reduced base of support for the hindpaws. Histochemical analyses in the CNS showed astrocytosis and up-regulation of calcium-binding proteins. In addition, a loss of Purkinje cells in the cerebellum was observed. No demyelination was present in the CNS. Motor nerve conduction velocity measurements revealed a peripheral neuropathy. Our results show that, in the mouse, high phytanic acid levels cause a peripheral neuropathy and ataxia with loss of Purkinje cells. These findings provide important insights in the pathophysiology of Refsum disease.

  8. Involvement of NADPH oxidase in up-regulation of plasminogen activator inhibitor-1 and heat shock factor-1 in mouse embryo fibroblasts induced by oxidized LDL and in apolipoprotein E-deficient mice.

    PubMed

    Zhao, Ruozhi; Moghadasian, Mohammed H; Shen, Garry X

    2011-09-01

    The present study demonstrated that oxidized LDL (oLDL) increased the generation of superoxide and hydrogen peroxide (H(2)O(2)), the abundances of NADPH oxidase (NOX)4, NOX2, p22-phox and lectin-like oLDL receptor-1 (LOX-1) in wild-type or heat shock factor-1 (HSF1)-deficient mouse embryo fibroblasts (MEF). LOX-1 antibody inhibited LDL or oLDL-induced expression of NOX components in MEF. Abundance of HSF1 or plasminogen activator inhibitor-1 (PAI-1) was increased by oLDL in wild-type, but not in HSF1-deficient MEF. Diphenyleneiodonium or siRNA for NOX or p22-phox inhibited oLDL-induced increases of HSF1, PAI-1 and H(2)O(2) in MEF. Increased NOX4, NOX2, LOX1, HSF1 and PAI-1 were detected in aortae and hearts of apolipoprotein E-knockout (apoE-KO) mice compared to controls, which were associated with increased serum cholesterol or plasma PAI-1. The results suggest that NOX is required for oLDL-induced HSF1 or PAI-1 expression in MEF, which was supported by the up-regulation of NOX, LOX-1, HSF1 and PAI-1 in apoE-KO mice.

  9. Mouse Curve Biometrics

    SciTech Connect

    Schulz, Douglas A.

    2007-10-08

    A biometric system suitable for validating user identity using only mouse movements and no specialized equipment is presented. Mouse curves (mouse movements with little or no pause between them) are individually classied and used to develop classication histograms, which are representative of an individual's typical mouse use. These classication histograms can then be compared to validate identity. This classication approach is suitable for providing continuous identity validation during an entire user session.

  10. Building a Brainier Mouse.

    ERIC Educational Resources Information Center

    Tsien, Joe Z.

    2000-01-01

    Describes a genetic engineering project to build an intelligent mouse. Cites understanding the molecular basis of learning and memory as a very important step. Concludes that while science will never create a genius mouse that plays the stock market, it can turn a mouse into a quick learner with a better memory. (YDS)

  11. Involvement of phosphoinositide 3-kinase class IA (PI3K 110α) and NADPH oxidase 1 (NOX1) in regulation of vascular differentiation induced by vascular endothelial growth factor (VEGF) in mouse embryonic stem cells.

    PubMed

    Bekhite, Mohamed M; Müller, Veronika; Tröger, Sebastian H; Müller, Jörg P; Figulla, Hans-Reiner; Sauer, Heinrich; Wartenberg, Maria

    2016-04-01

    The impact of reactive oxygen species and phosphoinositide 3-kinase (PI3K) in differentiating embryonic stem (ES) cells is largely unknown. Here, we show that the silencing of the PI3K catalytic subunit p110α and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 1 (NOX1) by short hairpin RNA or pharmacological inhibition of NOX and ras-related C3 botulinum toxin substrate 1 (Rac1) abolishes superoxide production by vascular endothelial growth factor (VEGF) in mouse ES cells and in ES-cell-derived fetal liver kinase-1(+) (Flk-1(+)) vascular progenitor cells, whereas the mitochondrial complex I inhibitor rotenone does not have an effect. Silencing p110α or inhibiting Rac1 arrests vasculogenesis at initial stages in embryoid bodies, even under VEGF treatment, as indicated by platelet endothelial cell adhesion molecule-1 (PECAM-1)-positive areas and branching points. In the absence of p110α, tube-like structure formation on matrigel and cell migration of Flk-1(+) cells in scratch migration assays are totally impaired. Silencing NOX1 causes a reduction in PECAM-1-positive areas, branching points, cell migration and tube length upon VEGF treatment, despite the expression of vascular differentiation markers. Interestingly, silencing p110α but not NOX1 inhibits the activation of Rac1, Ras homologue gene family member A (RhoA) and Akt leading to the abrogation of VEGF-induced lamellipodia structure formation. Thus, our data demonstrate that the PI3K p110α-Akt/Rac1 and NOX1 signalling pathways play a pivotal role in VEGF-induced vascular differentiation and cell migration. Rac1, RhoA and Akt phosphorylation occur downstream of PI3K and upstream of NOX1 underscoring a role of PI3K p110α in the regulation of cell polarity and migration. PMID:26553657

  12. Vitamins as radioprotectors in vivo. I. Protection by vitamin C against internal radionuclides in mouse testes: Implications to the mechanism of damage caused by the Auger effect

    SciTech Connect

    Narra, V.R.; Harapanhalli, R.S.; Howell, R.W.; Rao, D.V. ); Sastry, K.S.R. )

    1994-03-01

    The potential of vitamin C, an antioxidant, to protect the radiosensitive spermatogonial cells in mouse testes against the effects of chronic irradiation by radionuclides incorporated into tissue was investigated. Interestingly, when injected intratesticularly, a small and nontoxic amount of vitamin C (1.5 [mu]g in 3 [mu]l saline) protected the spermatogonia against the damage associated with high-LET radiation caused by Auger electrons from similarly administered 5-([sup 125]I)-iodo-2[prime]-deoxyuridine ([sup 125]IdU). A dose modification factor (DMF) of 2.3 was obtained. In contrast, no protection was observed when [sup 210]Po, an [alpha]-particle emitter, was administered similarly. These findings suggest that the mechanism of action of the Auger effect is of an indirect nature, which is in contrast to the direct action generally believed to be responsible for biological damage caused by high-LET radiations. 31 refs., 4 figs., 1 tab.

  13. Mouse Models of Human Phenylketonuria

    PubMed Central

    Shedlovsky, A.; McDonald, J. D.; Symula, D.; Dove, W. F.

    1993-01-01

    Phenylketonuria (PKU) results from a deficiency in phenylalanine hydroxylase, the enzyme catalyzing the conversion of phenylalanine (PHE) to tyrosine. Although this inborn error of metabolism was among the first in humans to be understood biochemically and genetically, little is known of the mechanism(s) involved in the pathology of PKU. We have combined mouse germline mutagenesis with screens for hyperphenylalaninemia to isolate three mutants deficient in phenylalanine hydroxylase (PAH) activity and cross-reactive protein. Two of these have reduced PAH mRNA and display characteristics of untreated human PKU patients. A low PHE diet partially reverses these abnormalities. Our success in using high frequency random germline point mutagenesis to obtain appropriate disease models illustrates how such mutagenesis can complement the emergent power of targeted mutagenesis in the mouse. The mutants now can be used as models in studying both maternal PKU and somatic gene therapy. PMID:8375656

  14. Parent Involvement.

    ERIC Educational Resources Information Center

    LaCrosse, Ed

    The paper discusses the rationale and guidelines for parent involvement in HCEEP (Handicapped Children's Early Education Program) projects. Ways of assessing parents' needs are reviewed, as are four types of services to meet the identified needs: parent education, direct participation, parent counseling, and parent provided programs. Materials and…

  15. An encyclopedia of mouse DNA elements (Mouse ENCODE).

    PubMed

    Stamatoyannopoulos, John A; Snyder, Michael; Hardison, Ross; Ren, Bing; Gingeras, Thomas; Gilbert, David M; Groudine, Mark; Bender, Michael; Kaul, Rajinder; Canfield, Theresa; Giste, Erica; Johnson, Audra; Zhang, Mia; Balasundaram, Gayathri; Byron, Rachel; Roach, Vaughan; Sabo, Peter J; Sandstrom, Richard; Stehling, A Sandra; Thurman, Robert E; Weissman, Sherman M; Cayting, Philip; Hariharan, Manoj; Lian, Jin; Cheng, Yong; Landt, Stephen G; Ma, Zhihai; Wold, Barbara J; Dekker, Job; Crawford, Gregory E; Keller, Cheryl A; Wu, Weisheng; Morrissey, Christopher; Kumar, Swathi A; Mishra, Tejaswini; Jain, Deepti; Byrska-Bishop, Marta; Blankenberg, Daniel; Lajoie, Bryan R; Jain, Gaurav; Sanyal, Amartya; Chen, Kaun-Bei; Denas, Olgert; Taylor, James; Blobel, Gerd A; Weiss, Mitchell J; Pimkin, Max; Deng, Wulan; Marinov, Georgi K; Williams, Brian A; Fisher-Aylor, Katherine I; Desalvo, Gilberto; Kiralusha, Anthony; Trout, Diane; Amrhein, Henry; Mortazavi, Ali; Edsall, Lee; McCleary, David; Kuan, Samantha; Shen, Yin; Yue, Feng; Ye, Zhen; Davis, Carrie A; Zaleski, Chris; Jha, Sonali; Xue, Chenghai; Dobin, Alex; Lin, Wei; Fastuca, Meagan; Wang, Huaien; Guigo, Roderic; Djebali, Sarah; Lagarde, Julien; Ryba, Tyrone; Sasaki, Takayo; Malladi, Venkat S; Cline, Melissa S; Kirkup, Vanessa M; Learned, Katrina; Rosenbloom, Kate R; Kent, W James; Feingold, Elise A; Good, Peter J; Pazin, Michael; Lowdon, Rebecca F; Adams, Leslie B

    2012-08-13

    To complement the human Encyclopedia of DNA Elements (ENCODE) project and to enable a broad range of mouse genomics efforts, the Mouse ENCODE Consortium is applying the same experimental pipelines developed for human ENCODE to annotate the mouse genome.

  16. Involvement of voltage-gated sodium channel Na(v)1.8 in the regulation of the release and synthesis of substance P in adult mouse dorsal root ganglion neurons.

    PubMed

    Tang, He-Bin; Shiba, Eri; Li, Yu-Sang; Morioka, Norimitsu; Zheng, Tai-Xing; Ogata, Nobukuni; Nakata, Yoshihiro

    2008-10-01

    This study was conducted to determine whether Na(v)1.8 contributes to the release and/or synthesis of substance P (SP) in adult mice dorsal root ganglion (DRG) neurons. The SP released from cultured DRG neurons of Na(v)1.8 knock-out mice exposed to either capsaicin or KCl was significantly lower than that from wild-type (C57BL/6) mice based on a radioimmunoassay. The SP level of L6 DRG in Na(v)1.8 knock-out mice was also lower than that in wild-type mice. After chronic constriction injury (CCI) of the sciatic nerve, the level of SP decreased in the L6 ipsilateral DRG of wild-type but not Na(v)1.8 knock-out mice. The preprotachykinin-A (PPT-A) mRNAs in L4 - 6 DRGs of Na(v)1.8 knock-out mice also fell to half their normally abundant levels of expression. There were significant increases in Na(v)1.8 expression of the L6 contralateral DRG from wild-type mice and in the percentage of neurons expressing neurokinin-1 receptor in the cytosol of L6 DRGs from wild-type or Na(v)1.8 knock-out mice. These findings suggest that Na(v)1.8 is involved in the regulation of the release and synthesis of SP in the DRG neurons of wild-type mice. PMID:18845912

  17. Optical mouse acting as biospeckle sensor

    NASA Astrophysics Data System (ADS)

    da Silva, Michel Melo; Nozela, Jose Roberto de Almeida; Chaves, Marcio Jose; Alves Braga, Roberto; Rabal, Hector Jorge

    2011-04-01

    In this work we propose some experiments with the use of optical computer mouse, associated to low cost lasers that can be used to perform several measurements with applications in industry and in human health monitoring. The mouse was used to grab the movements produced by speckle pattern changes and to get information through the adaptation of its structure. We measured displacements in wood samples under strain, variations of the diameter of an artery due to heart beat and, through a hardware simulation, the movement of an eye, an experiment that could be of low cost help for communication to severely handicapped motor patients. Those measurements were done in spite of the fact that the CCD sensor of the mice is monolithically included into an integrated circuit so that the raw image cannot be accessed. If, as was the case with primitive optical mouse, that signal could be accessed, the quality and usefulness of the measurements could be significantly increased. As it was not possible, a webcam sensor was used for measuring the drying of paint, a standard phenomenon for testing biospeckle techniques, in order to prove the usefulness of the mouse design. The results showed that the use of the mouse associated to a laser pointer could be the way to get metrological information from many phenomena involving the whole field spatial displacement, as well as the use of the mouse as in its prime version allowed to get images of the speckle patterns and to analyze them.

  18. Involvement of the 5-HT(1A) receptor in the anti-immobility effects of fluvoxamine in the forced swimming test and mouse strain differences in 5-HT(1A) receptor binding.

    PubMed

    Sugimoto, Yumi; Furutani, Sachiko; Kajiwara, Yoshinobu; Hirano, Kazufumi; Yamada, Shizuo; Tagawa, Noriko; Kobayashi, Yoshiharu; Hotta, Yoshihiro; Yamada, Jun

    2010-03-10

    We previously demonstrated the presence of strain differences in baseline immobility time and sensitivity to the selective serotonin reuptake inhibitor (SSRI) fluvoxamine in five strains of mice (ICR, ddY, C57BL, DBA/2 and BALB/c mice). Furthermore, variations in serotonin (5-HT) transporter binding in the brain were strongly related to strain differences in baseline immobility and sensitivity to fluvoxamine. In the present study, we examined the involvement of the 5-HT(1A) receptor in anti-immobility effects in DBA/2 mice, which show high sensitivity to fluvoxamine. The anti-immobility effects of fluvoxamine in DBA/2 mice were inhibited by the 5-HT(1A) receptor antagonist N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)cyclohexanecarboxamide (WAY 100635). However, the 5-HT(1B) receptor antagonist 3-[3-(dimethylamino)propyl]-4-hydroxy-N-[4-(4-pyridinyl)phenyl]benzamide (GR55562), the 5-HT(2) receptor antagonist 6-methyl-1-(methylethyl)-ergoline-8beta-carboxylic acid 2-hydroxy-1-methylpropyl ester (LY 53857), the 5-HT(3) receptor antagonist ondansetron and the 5-HT(4) receptor antagonist 4-amino-5-chloro-2-methoxy-benzoic acid 2-(diethylamino)ethyl ester (SDZ 205,557) did not influence the anti-immobility effects of fluvoxamine in DBA/2 mice. These results suggest that fluvoxamine-induced antidepressant-like effects in DBA/2 mice are mediated by the 5-HT(1A) receptor. We analyzed 5-HT(1A) receptor binding in the brains of five strains of mice. Strain differences in 5-HT(1A) receptor binding were observed. 5-HT(1A) receptor binding in brain was not correlated with baseline immobility time in the five strains of mice examined. These results suggest that, although the anti-immobility effects of fluvoxamine in DBA/2 mice are mediated by the 5-HT(1A) receptor, strain differences in 5-HT(1A) receptor binding are not related to variation in immobility time and responses to fluvoxamine.

  19. The MOUSE Squad

    ERIC Educational Resources Information Center

    Borja, Rhea R.

    2004-01-01

    This article presents a New York city after-school program started by MOUSE (Making Opportunities for Upgrading Schools and Education), a national nonprofit group that teaches students how to fix computers, and equips them with the communication and problem-solving skills to help them in the working world. The MOUSE program is part of a trend…

  20. Mouse homologues of human hereditary disease.

    PubMed Central

    Searle, A G; Edwards, J H; Hall, J G

    1994-01-01

    Details are given of 214 loci known to be associated with human hereditary disease, which have been mapped on both human and mouse chromosomes. Forty two of these have pathological variants in both species; in general the mouse variants are similar in their effects to the corresponding human ones, but exceptions include the Dmd/DMD and Hprt/HPRT mutations which cause little, if any, harm in mice. Possible reasons for phenotypic differences are discussed. In most pathological variants the gene product seems to be absent or greatly reduced in both species. The extensive data on conserved segments between human and mouse chromosomes are used to predict locations in the mouse of over 50 loci of medical interest which are mapped so far only on human chromosomes. In about 80% of these a fairly confident prediction can be made. Some likely homologies between mapped mouse loci and unmapped human ones are also given. Sixty six human and mouse proto-oncogene and growth factor gene homologies are also listed; those of confirmed location are all in known conserved segments. A survey of 18 mapped human disease loci and chromosome regions in which the manifestation or severity of pathological effects is thought to be the result of genomic imprinting shows that most of the homologous regions in the mouse are also associated with imprinting, especially those with homologues on human chromosomes 11p and 15q. Useful methods of accelerating the production of mouse models of human hereditary disease include (1) use of a supermutagen, such as ethylnitrosourea (ENU), (2) targeted mutagenesis involving ES cells, and (3) use of gene transfer techniques, with production of 'knockout mutations'. PMID:8151633

  1. Pathology of Mouse Models of Accelerated Aging.

    PubMed

    Harkema, L; Youssef, S A; de Bruin, A

    2016-03-01

    Progeroid mouse models display phenotypes in multiple organ systems that suggest premature aging and resemble features of natural aging of both mice and humans. The prospect of a significant increase in the global elderly population within the next decades has led to the emergence of "geroscience," which aims at elucidating the molecular mechanisms involved in aging. Progeroid mouse models are frequently used in geroscience as they provide insight into the molecular mechanisms that are involved in the highly complex process of natural aging. This review provides an overview of the most commonly reported nonneoplastic macroscopic and microscopic pathologic findings in progeroid mouse models (eg, osteoporosis, osteoarthritis, degenerative joint disease, intervertebral disc degeneration, kyphosis, sarcopenia, cutaneous atrophy, wound healing, hair loss, alopecia, lymphoid atrophy, cataract, corneal endothelial dystrophy, retinal degenerative diseases, and vascular remodeling). Furthermore, several shortcomings in pathologic analysis and descriptions of these models are discussed. Progeroid mouse models are valuable models for aging, but thorough knowledge of both the mouse strain background and the progeria-related phenotype is required to guide interpretation and translation of the pathology data. PMID:26864891

  2. Mouse genome database 2016.

    PubMed

    Bult, Carol J; Eppig, Janan T; Blake, Judith A; Kadin, James A; Richardson, Joel E

    2016-01-01

    The Mouse Genome Database (MGD; http://www.informatics.jax.org) is the primary community model organism database for the laboratory mouse and serves as the source for key biological reference data related to mouse genes, gene functions, phenotypes and disease models with a strong emphasis on the relationship of these data to human biology and disease. As the cost of genome-scale sequencing continues to decrease and new technologies for genome editing become widely adopted, the laboratory mouse is more important than ever as a model system for understanding the biological significance of human genetic variation and for advancing the basic research needed to support the emergence of genome-guided precision medicine. Recent enhancements to MGD include new graphical summaries of biological annotations for mouse genes, support for mobile access to the database, tools to support the annotation and analysis of sets of genes, and expanded support for comparative biology through the expansion of homology data.

  3. Mouse genome database 2016

    PubMed Central

    Bult, Carol J.; Eppig, Janan T.; Blake, Judith A.; Kadin, James A.; Richardson, Joel E.

    2016-01-01

    The Mouse Genome Database (MGD; http://www.informatics.jax.org) is the primary community model organism database for the laboratory mouse and serves as the source for key biological reference data related to mouse genes, gene functions, phenotypes and disease models with a strong emphasis on the relationship of these data to human biology and disease. As the cost of genome-scale sequencing continues to decrease and new technologies for genome editing become widely adopted, the laboratory mouse is more important than ever as a model system for understanding the biological significance of human genetic variation and for advancing the basic research needed to support the emergence of genome-guided precision medicine. Recent enhancements to MGD include new graphical summaries of biological annotations for mouse genes, support for mobile access to the database, tools to support the annotation and analysis of sets of genes, and expanded support for comparative biology through the expansion of homology data. PMID:26578600

  4. Mouse genome database 2016.

    PubMed

    Bult, Carol J; Eppig, Janan T; Blake, Judith A; Kadin, James A; Richardson, Joel E

    2016-01-01

    The Mouse Genome Database (MGD; http://www.informatics.jax.org) is the primary community model organism database for the laboratory mouse and serves as the source for key biological reference data related to mouse genes, gene functions, phenotypes and disease models with a strong emphasis on the relationship of these data to human biology and disease. As the cost of genome-scale sequencing continues to decrease and new technologies for genome editing become widely adopted, the laboratory mouse is more important than ever as a model system for understanding the biological significance of human genetic variation and for advancing the basic research needed to support the emergence of genome-guided precision medicine. Recent enhancements to MGD include new graphical summaries of biological annotations for mouse genes, support for mobile access to the database, tools to support the annotation and analysis of sets of genes, and expanded support for comparative biology through the expansion of homology data. PMID:26578600

  5. DICER Regulates the Formation and Maintenance of Cell-Cell Junctions in the Mouse Seminiferous Epithelium.

    PubMed

    Korhonen, Hanna Maria; Yadav, Ram Prakash; Da Ros, Matteo; Chalmel, Frédéric; Zimmermann, Céline; Toppari, Jorma; Nef, Serge; Kotaja, Noora

    2015-12-01

    The endonuclease DICER that processes micro-RNAs and small interfering RNAs is essential for normal spermatogenesis and male fertility. We previously showed that the deletion of Dicer1 gene in postnatal spermatogonia in mice using Ngn3 promoter-driven Cre expression caused severe defects in the morphogenesis of haploid spermatid to mature spermatozoon, including problems in cell polarization and nuclear elongation. In this study, we further analyzed the same mouse model and revealed that absence of functional DICER in differentiating male germ cells induces disorganization of the cell-cell junctions in the seminiferous epithelium. We detected discontinuous and irregular apical ectoplasmic specializations between elongating spermatids and Sertoli cells. The defective anchoring of spermatids to Sertoli cells caused a premature release of spermatids into the lumen. Our findings may help also explain the abnormal elongation process of remaining spermatids because these junctions and the correct positioning of germ cells in the epithelium are critically important for the progression of spermiogenesis. Interestingly, cell adhesion-related genes were generally upregulated in Dicer1 knockout germ cells. Claudin 5 ( Cldn5 ) was among the most upregulated genes and we show that the polarized localization of CLAUDIN5 in the apical ectoplasmic specializations was lost in Dicer1 knockout spermatids. Our results suggest that DICER-dependent pathways control the formation and organization of cell-cell junctions in the seminiferous epithelium via the regulation of cell adhesion-related genes. PMID:26510868

  6. Distribution of the sex chromosome during mouse spermatogenesis in testis tissue sections.

    PubMed

    Otaka, Kosuke; Hiradate, Yuuki; Kobayashi, Norio; Shirakata, Yoshiki; Tanemura, Kentaro

    2015-01-01

    During mammalian spermatogenesis, spermatogenic cells undergo mitotic division and are subsequently divided into haploid spermatids by meiotic division, but the dynamics of sex chromosomes during spermatogenesis are unclear in vivo. To gain insight into the distribution of sex chromosomes in the testis, we examined the localization of sex chromosomes before and after meiosis in mouse testis sections. Here, we developed a method of fluorescence in situ hybridization (FISH) using specific probes for the X and Y chromosomes to obtain their positional information in histological testis sections. FISH analysis revealed the sex chromosomal position during spermatogenesis in each stage of seminiferous epithelia and in each spermatogenic cell. In the spermatogonia and leptotene spermatocytes, sex chromosomes were distantly positioned in the cell. In the zygotene and pachytene spermatocytes at prophase I, X and Y chromosomes had a random distribution. After meiosis, the X and Y spermatids were random in every seminiferous epithelium. We also detected aneuploidy of sex chromosomes in spermatogenic cells using our developed FISH analysis. Our results provide further insight into the distribution of sex chromosomes during spermatogenesis, which could help to elucidate a specific difference between X and Y spermatids and sex chromosome-specific behavior.

  7. DMRT1 Is Required for Mouse Spermatogonial Stem Cell Maintenance and Replenishment.

    PubMed

    Zhang, Teng; Oatley, Jon; Bardwell, Vivian J; Zarkower, David

    2016-09-01

    Male mammals produce sperm for most of postnatal life and therefore require a robust germ line stem cell system, with precise balance between self-renewal and differentiation. Prior work established doublesex- and mab-3-related transcription factor 1 (Dmrt1) as a conserved transcriptional regulator of male sexual differentiation. Here we investigate the role of Dmrt1 in mouse spermatogonial stem cell (SSC) homeostasis. We find that Dmrt1 maintains SSCs during steady state spermatogenesis, where it regulates expression of Plzf, another transcription factor required for SSC maintenance. We also find that Dmrt1 is required for recovery of spermatogenesis after germ cell depletion. Committed progenitor cells expressing Ngn3 normally do not contribute to SSCs marked by the Id4-Gfp transgene, but do so when spermatogonia are chemically depleted using busulfan. Removal of Dmrt1 from Ngn3-positive germ cells blocks the replenishment of Id4-GFP-positive SSCs and recovery of spermatogenesis after busulfan treatment. Our data therefore reveal that Dmrt1 supports SSC maintenance in two ways: allowing SSCs to remain in the stem cell pool under normal conditions; and enabling progenitor cells to help restore the stem cell pool after germ cell depletion. PMID:27583450

  8. DMRT1 Is Required for Mouse Spermatogonial Stem Cell Maintenance and Replenishment

    PubMed Central

    Zhang, Teng; Oatley, Jon; Bardwell, Vivian J.; Zarkower, David

    2016-01-01

    Male mammals produce sperm for most of postnatal life and therefore require a robust germ line stem cell system, with precise balance between self-renewal and differentiation. Prior work established doublesex- and mab-3-related transcription factor 1 (Dmrt1) as a conserved transcriptional regulator of male sexual differentiation. Here we investigate the role of Dmrt1 in mouse spermatogonial stem cell (SSC) homeostasis. We find that Dmrt1 maintains SSCs during steady state spermatogenesis, where it regulates expression of Plzf, another transcription factor required for SSC maintenance. We also find that Dmrt1 is required for recovery of spermatogenesis after germ cell depletion. Committed progenitor cells expressing Ngn3 normally do not contribute to SSCs marked by the Id4-Gfp transgene, but do so when spermatogonia are chemically depleted using busulfan. Removal of Dmrt1 from Ngn3-positive germ cells blocks the replenishment of Id4-GFP-positive SSCs and recovery of spermatogenesis after busulfan treatment. Our data therefore reveal that Dmrt1 supports SSC maintenance in two ways: allowing SSCs to remain in the stem cell pool under normal conditions; and enabling progenitor cells to help restore the stem cell pool after germ cell depletion. PMID:27583450

  9. Pathophysiology of gene-targeted mouse models for cystic fibrosis.

    PubMed

    Grubb, B R; Boucher, R C

    1999-01-01

    Pathophysiology of Gene-Targeted Mouse Models for Cystic Fibrosis. Physiol. Rev. 79, Suppl.: S193-S214, 1999. - Mutations in the gene causing the fatal disease cystic fibrosis (CF) result in abnormal transport of several ions across a number of epithelial tissues. In just 3 years after this gene was cloned, the first CF mouse models were generated. The CF mouse models generated to date have provided a wealth of information on the pathophysiology of the disease in a variety of organs. Heterogeneity of disease in the mouse models is due to the variety of gene-targeting strategies used in the generation of the CF mouse models as well as the diversity of the murine genetic background. This paper reviews the pathophysiology in the tissues and organs (gastrointestinal, airway, hepatobiliary, pancreas, reproductive, and salivary tissue) involved in the disease in the various CF mouse models. Marked similarities to and differences from the human disease have been observed in the various murine models. Some of the CF mouse models accurately reflect the ion-transport abnormalities and disease phenotype seen in human CF patients, especially in gastrointestinal tissue. However, alterations in airway ion transport, which lead to the devastating lung disease in CF patients, appear to be largely absent in the CF mouse models. Reasons for these unexpected findings are discussed. This paper also reviews pharmacotherapeutic and gene therapeutic studies in the various mouse models. PMID:9922382

  10. Mouse Cleaning Apparatus and Method

    NASA Technical Reports Server (NTRS)

    Williams, Glenn L. (Inventor)

    2005-01-01

    The method of using the mouse pad cleaning apparatus is disclosed and claimed. The method comprises the steps of uncovering the mouse cleaning surface, applying the mouse and ball of the mouse to the cleaning surface, moving the mouse in a rotational pattern on the mouse cleaning surface, removing the mouse form the mouse cleaning surface, washing the cleaning surface, and covering the mouse cleaning surface. A mouse pad cleaning apparatus comprising a plurality of substrates, each said substrate having adhesive thereon, said plurality of substrates residing in and affixed to a receptacle. A single substrate having adhesive, which may be washable or non-washable, thereon may be employed. The washable adhesive may be an organopolysiloxane or gelatinous elastomer.

  11. Effect of different cryoprotectant agents on spermatogenesis efficiency in cryopreserved and grafted neonatal mouse testicular tissue.

    PubMed

    Yildiz, Cengiz; Mullen, Brendan; Jarvi, Keith; McKerlie, Colin; Lo, Kirk C

    2013-08-01

    Restoration of male fertility associated with use of the cryopreserved testicular tissue would be a significant advance in human and animal assisted reproductive technology. The purpose of this study was to test the effects of four different cryoprotectant agents (CPA) on spermatogenesis and steroidogenesis in cryopreserved and allotransplanted neonatal mouse testicular tissue. Hank's balanced salt solution (HBSS) with 5% fetal bovine serum including either 0.7 M dimethyl sulfoxide (DMSO), 0.7 M propylene glycol (PrOH), 0.7 M ethylene glycol (EG), or glycerol was used as the cryoprotectant solution. Donor testes were collected and dissected from neonatal pups of CD-1 mice (one day old). Freezing and seeding of the testicular whole tissues was performed using an automated controlled-rate freezer. Four fresh (non-frozen) or frozen-thawed pieces of testes were subcutaneously grafted onto the hind flank of each castrated male NCr nude recipient mouse and harvested after 3 months. Fresh neonatal testes grafts recovered from transplant sites had the most advanced rate of spermatogenesis with elongated spermatid and spermatozoa in 46.6% of seminiferous tubules and had higher levels of serum testosterone compared to all other frozen-thawed-graft groups (p<0.05). Fresh grafts and frozen-thawed grafts in the DMSO group had the highest rate of tissue survival compared to PrOH, EG, and glycerol after harvesting (p>0.05). The most effective CPA for the freezing and thawing of neonatal mouse testes was DMSO in comparison with EG (p<0.05) in both pre-grafted and post-grafted tissues based on histopathological evaluation. Likewise, the highest level of serum testosterone was obtained from the DMSO CPA group compared to all other cryoprotectants evaluated (p<0.05). The typical damage observed in the frozen-thawed grafts included disruption of the interstitial stroma, intercellular connection ruptures, and detachment of spermatogonia from the basement membrane. These findings

  12. TAF4b is required for mouse spermatogonial stem cell development

    PubMed Central

    Lovasco, Lindsay A.; Gustafson, Eric A.; Seymour, Kimberly A.; de Rooij, Dirk G.; Freiman, Richard N.

    2014-01-01

    Long-term mammalian spermatogenesis requires proper development of spermatogonial stem cells (SSCs) that replenish the testis with germ cell progenitors during adult life. TAF4b is a gonadal-enriched component of the general transcription factor complex, TFIID, which is required for the maintenance of spermatogenesis in the mouse. Successful germ cell transplantation assays into adult TAF4b-deficient host testes suggested that TAF4b performs an essential germ cell autonomous function in SSC establishment and/or maintenance. To elucidate the SSC function of TAF4b, we characterized the initial gonocyte pool and rounds of spermatogenic differentiation in the context of the Taf4b-deficient mouse testis. Here we demonstrate a significant reduction in the late embryonic gonocyte pool and a deficient expansion of this pool soon after birth. Resulting from this reduction of germ cell progenitors is a developmental delay in meiosis initiation, as compared to age-matched controls. While GFRα1+ spermatogonia are appropriately present as Asingle and Apaired in wild type testes, TAF4b-deficient testes display an increased proportion of long and clustered chains of GFRα1+ cells. In the absence of TAF4b, seminiferous tubules in the adult testis either lack germ cells altogether or are found to have missing generations of spermatogenic progenitor cells. Together these data indicate that TAF4b-deficient spermatogenic progenitor cells display a tendency for differentiation at the expense of self-renewal and a renewing pool of SSCs fail to establish during the critical window of SSC development. PMID:25727968

  13. Mouse bladder wall injection.

    PubMed

    Fu, Chi-Ling; Apelo, Charity A; Torres, Baldemar; Thai, Kim H; Hsieh, Michael H

    2011-07-12

    Mouse bladder wall injection is a useful technique to orthotopically study bladder phenomena, including stem cell, smooth muscle, and cancer biology. Before starting injections, the surgical area must be cleaned with soap and water and antiseptic solution. Surgical equipment must be sterilized before use and between each animal. Each mouse is placed under inhaled isoflurane anesthesia (2-5% for induction, 1-3% for maintenance) and its bladder exposed by making a midline abdominal incision with scissors. If the bladder is full, it is partially decompressed by gentle squeezing between two fingers. The cell suspension of interest is intramurally injected into the wall of the bladder dome using a 29 or 30 gauge needle and 1 cc or smaller syringe. The wound is then closed using wound clips and the mouse allowed to recover on a warming pad. Bladder wall injection is a delicate microsurgical technique that can be mastered with practice.

  14. Apoptotic Signaling in Mouse Odontogenesis

    PubMed Central

    Svandova, Eva; Tucker, Abigail S.

    2012-01-01

    Abstract Apoptosis is an important morphogenetic event in embryogenesis as well as during postnatal life. In the last 2 decades, apoptosis in tooth development (odontogenesis) has been investigated with gradually increasing focus on the mechanisms and signaling pathways involved. The molecular machinery responsible for apoptosis exhibits a high degree of conservation but also organ and tissue specific patterns. This review aims to discuss recent knowledge about apoptotic signaling networks during odontogenesis, concentrating on the mouse, which is often used as a model organism for human dentistry. Apoptosis accompanies the entire development of the tooth and corresponding remodeling of the surrounding bony tissue. It is most evident in its role in the elimination of signaling centers within developing teeth, removal of vestigal tooth germs, and in odontoblast and ameloblast organization during tooth mineralization. Dental apoptosis is caspase dependent and proceeds via mitochondrial mediated cell death with possible amplification by Fas-FasL signaling modulated by Bcl-2 family members. PMID:22204278

  15. Apoptotic signaling in mouse odontogenesis.

    PubMed

    Matalova, Eva; Svandova, Eva; Tucker, Abigail S

    2012-01-01

    Apoptosis is an important morphogenetic event in embryogenesis as well as during postnatal life. In the last 2 decades, apoptosis in tooth development (odontogenesis) has been investigated with gradually increasing focus on the mechanisms and signaling pathways involved. The molecular machinery responsible for apoptosis exhibits a high degree of conservation but also organ and tissue specific patterns. This review aims to discuss recent knowledge about apoptotic signaling networks during odontogenesis, concentrating on the mouse, which is often used as a model organism for human dentistry. Apoptosis accompanies the entire development of the tooth and corresponding remodeling of the surrounding bony tissue. It is most evident in its role in the elimination of signaling centers within developing teeth, removal of vestigal tooth germs, and in odontoblast and ameloblast organization during tooth mineralization. Dental apoptosis is caspase dependent and proceeds via mitochondrial mediated cell death with possible amplification by Fas-FasL signaling modulated by Bcl-2 family members.

  16. A strategy for fine-structure functional analysis of a 6- to 11-centimorgan region of mouse chromosome 7 by high-efficiency mutagenesis

    SciTech Connect

    Rinchik, E.M.; Carpenter, D.A.; Selby, P.B. )

    1990-02-01

    A refined functional map of a 6- to 11-centimorgan region surrounding the albino (c) locus in mouse chromosome 7 is being generated by N-ethyl-N-nitrosourea (EtNU) saturation mutagenesis of stem-cell spermatogonia. In the first phase of an experiment that will eventually test at least 3,000 gametes, the authors screened 972 mutagenized gametes for the induction of both lethal and visible mutations with a two-cross breeding protocol. Thirteen mutations mapping within the limits of a segment corresponding to the cytologically visible Df(c Mod-2 sh-1){sup 26DVT} deletion were recovered. They represented three phenotypic groups: prenatal lethality (six mutations); a fitness/runting syndrome (three mutations, provisionally designated as fit variants); and a neurological/balance-defect abnormality (four mutations). Complementation analysis provided evidence for a true repeat mutation at the sh-1 (shaker-1) locus (for the neurological mutations) and another at the here defined fit-1 (fitness-1) locus. The recovery of the repeat mutations suggests that the EtNU-induced mutation rate, estimated from specific-locus tests, should make it possible to achieve saturation mutagenesis of a chromosomal region. This experiment is providing basic logistical and statistical information on which to base strategies for expanding the functional map of larger segments of the mouse genome by experimental mutagenesis.

  17. The Mouse SAGE Site: database of public mouse SAGE libraries.

    PubMed

    Divina, Petr; Forejt, Jirí

    2004-01-01

    The Mouse SAGE Site is a web-based database of all available public libraries generated by the Serial Analysis of Gene Expression (SAGE) from various mouse tissues and cell lines. The database contains mouse SAGE libraries organized in a uniform way and provides web-based tools for browsing, comparing and searching SAGE data with reliable tag-to-gene identification. A modified approach based on the SAGEmap database is used for reliable tag identification. The Mouse SAGE Site is maintained on an ongoing basis at the Institute of Molecular Genetics, Academy of Sciences of the Czech Republic and is accessible at the internet address http://mouse.biomed.cas.cz/sage/.

  18. MPHASYS: a mouse phenotype analysis system

    PubMed Central

    Calder, R Brent; Beems, Rudolf B; van Steeg, Harry; Mian, I Saira; Lohman, Paul HM; Vijg, Jan

    2007-01-01

    Background Systematic, high-throughput studies of mouse phenotypes have been hampered by the inability to analyze individual animal data from a multitude of sources in an integrated manner. Studies generally make comparisons at the level of genotype or treatment thereby excluding associations that may be subtle or involve compound phenotypes. Additionally, the lack of integrated, standardized ontologies and methodologies for data exchange has inhibited scientific collaboration and discovery. Results Here we introduce a Mouse Phenotype Analysis System (MPHASYS), a platform for integrating data generated by studies of mouse models of human biology and disease such as aging and cancer. This computational platform is designed to provide a standardized methodology for working with animal data; a framework for data entry, analysis and sharing; and ontologies and methodologies for ensuring accurate data capture. We describe the tools that currently comprise MPHASYS, primarily ones related to mouse pathology, and outline its use in a study of individual animal-specific patterns of multiple pathology in mice harboring a specific germline mutation in the DNA repair and transcription-specific gene Xpd. Conclusion MPHASYS is a system for analyzing multiple data types from individual animals. It provides a framework for developing data analysis applications, and tools for collecting and distributing high-quality data. The software is platform independent and freely available under an open-source license [1]. PMID:17553167

  19. Targeted disruption of the mouse testis-enriched gene Znf230 does not affect spermatogenesis or fertility

    PubMed Central

    Liu, Yunqiang; Tao, Dachang; Lu, Yongjie; Yang, Yuan; Ma, Yongxin; Zhang, Sizhong

    2014-01-01

    The mouse testis-enriched Znf230 gene, which encodes a type of RING finger protein, is present primarily in the nuclei of spermatogonia, the acrosome and the tail of spermatozoa. To investigate the role of Znf230 in spermatogenesis, we generated Znf230-deficient mice by disrupting Znf230 exon-5 and exon-6 using homologous recombination. The homozygous Znf230-knockout (KO) mice did not exhibit Znf230 mRNA expression and Znf230 protein production. Znf230 KO mice exhibited no obvious impairment in body growth or fertility. Male Znf230 KO mice had integral reproductive systems and mature sperm that were regular in number and shape. The developmental stages of male germ cells of Znf230 KO mice were also normal. We further examined variations in the transcriptomes of testicular tissue between Znf230 KO and wild-type mice through microarray analysis. The results showed that the mRNA level of one unclassified transcript 4921513I08Rik was increased and that the mRNA levels of three other transcripts, i.e., 4930448A20Rik, 4931431B13Rik and potassium channel tetramerisation domain containing 14(Kctd14), were reduced more than two-fold in Znf230 KO mice compared with wild-type mice. Using our current examination techniques, these findings suggested that Znf230 deficiency in mice may not affect growth, fertility or spermatogenesis. PMID:25505846

  20. Neuroanatomy and Neurochemistry of Mouse Cornea

    PubMed Central

    He, Jiucheng; Bazan, Haydee E. P.

    2016-01-01

    Purpose To investigate the entire nerve architecture and content of the two main sensory neuropeptides in mouse cornea to determine if it is a good model with similarities to human corneal innervation. Methods Mice aged 1 to 24 weeks were used. The corneas were stained with neuronal-class βIII-tubulin, calcitonin gene–related peptide (CGRP), and substance P (SP) antibodies; whole-mount images were acquired to build an entire view of corneal innervation. To test the origin of CGRP and SP, trigeminal ganglia (TG) were processed for immunofluorescence. Relative corneal nerve fiber densities or neuron numbers were assessed by computer-assisted analysis. Results Between 1 and 3 weeks after birth, mouse cornea was mainly composed of a stromal nerve network. At 4 weeks, a whorl-like structure (or vortex) appeared that gradually became more defined. By 8 weeks, anatomy of corneal nerves had reached maturity. Epithelial bundles converged into the central area to form the vortex. The number and pattern of whorl-like structures were different. Subbasal nerve density and nerve terminals were greater in the center than the periphery. Nerve fibers and terminals that were CGRP-positive were more abundant than SP-positive nerves and terminals. In trigeminal ganglia, the number of CGRP-positive neurons significantly outnumbered those positive for SP. Conclusions This is the first study to show a complete map of the entire corneal nerves and CGRP and SP sensory neuropeptide distribution in the mouse cornea. This finding shows mouse corneal innervation has many similarities to human cornea and makes the mouse an appropriate model to study pathologies involving corneal nerves. PMID:26906155

  1. Development and testing of a mouse simulated space flight model

    NASA Technical Reports Server (NTRS)

    Sonnenfeld, Gerald

    1987-01-01

    The development and testing of a mouse model for simulating some aspects of weightlessness that occurs during space flight, and the carrying out of immunological experiments on animals undergoing space flight is examined. The mouse model developed was an antiorthostatic, hypokinetic, hypodynamic suspension model similar to one used with rats. The study was divided into two parts. The first involved determination of which immunological parameters should be observed on animals flown during space flight or studied in the suspension model. The second involved suspending mice and determining which of those immunological parameters were altered by the suspension. Rats that were actually flown in Space Shuttle SL-3 were used to test the hypotheses.

  2. A Comparative Encyclopedia of DNA Elements in the Mouse Genome

    PubMed Central

    Yue, Feng; Cheng, Yong; Breschi, Alessandra; Vierstra, Jeff; Wu, Weisheng; Ryba, Tyrone; Sandstrom, Richard; Ma, Zhihai; Davis, Carrie; Pope, Benjamin D.; Shen, Yin; Pervouchine, Dmitri D.; Djebali, Sarah; Thurman, Bob; Kaul, Rajinder; Rynes, Eric; Kirilusha, Anthony; Marinov, Georgi K.; Williams, Brian A.; Trout, Diane; Amrhein, Henry; Fisher-Aylor, Katherine; Antoshechkin, Igor; DeSalvo, Gilberto; See, Lei-Hoon; Fastuca, Meagan; Drenkow, Jorg; Zaleski, Chris; Dobin, Alex; Prieto, Pablo; Lagarde, Julien; Bussotti, Giovanni; Tanzer, Andrea; Denas, Olgert; Li, Kanwei; Bender, M. A.; Zhang, Miaohua; Byron, Rachel; Groudine, Mark T.; McCleary, David; Pham, Long; Ye, Zhen; Kuan, Samantha; Edsall, Lee; Wu, Yi-Chieh; Rasmussen, Matthew D.; Bansal, Mukul S.; Keller, Cheryl A.; Morrissey, Christapher S.; Mishra, Tejaswini; Jain, Deepti; Dogan, Nergiz; Harris, Robert S.; Cayting, Philip; Kawli, Trupti; Boyle, Alan P.; Euskirchen, Ghia; Kundaje, Anshul; Lin, Shin; Lin, Yiing; Jansen, Camden; Malladi, Venkat S.; Cline, Melissa S.; Erickson, Drew T.; Kirkup, Vanessa M; Learned, Katrina; Sloan, Cricket A.; Rosenbloom, Kate R.; de Sousa, Beatriz Lacerda; Beal, Kathryn; Pignatelli, Miguel; Flicek, Paul; Lian, Jin; Kahveci, Tamer; Lee, Dongwon; Kent, W. James; Santos, Miguel Ramalho; Herrero, Javier; Notredame, Cedric; Johnson, Audra; Vong, Shinny; Lee, Kristen; Bates, Daniel; Neri, Fidencio; Diegel, Morgan; Canfield, Theresa; Sabo, Peter J.; Wilken, Matthew S.; Reh, Thomas A.; Giste, Erika; Shafer, Anthony; Kutyavin, Tanya; Haugen, Eric; Dunn, Douglas; Reynolds, Alex P.; Neph, Shane; Humbert, Richard; Hansen, R. Scott; De Bruijn, Marella; Selleri, Licia; Rudensky, Alexander; Josefowicz, Steven; Samstein, Robert; Eichler, Evan E.; Orkin, Stuart H.; Levasseur, Dana; Papayannopoulou, Thalia; Chang, Kai-Hsin; Skoultchi, Arthur; Gosh, Srikanta; Disteche, Christine; Treuting, Piper; Wang, Yanli; Weiss, Mitchell J.; Blobel, Gerd A.; Good, Peter J.; Lowdon, Rebecca F.; Adams, Leslie B.; Zhou, Xiao-Qiao; Pazin, Michael J.; Feingold, Elise A.; Wold, Barbara; Taylor, James; Kellis, Manolis; Mortazavi, Ali; Weissman, Sherman M.; Stamatoyannopoulos, John; Snyder, Michael P.; Guigo, Roderic; Gingeras, Thomas R.; Gilbert, David M.; Hardison, Ross C.; Beer, Michael A.; Ren, Bing

    2014-01-01

    Summary As the premier model organism in biomedical research, the laboratory mouse shares the majority of protein-coding genes with humans, yet the two mammals differ in significant ways. To gain greater insights into both shared and species-specific transcriptional and cellular regulatory programs in the mouse, the Mouse ENCODE Consortium has mapped transcription, DNase I hypersensitivity, transcription factor binding, chromatin modifications, and replication domains throughout the mouse genome in diverse cell and tissue types. By comparing with the human genome, we not only confirm substantial conservation in the newly annotated potential functional sequences, but also find a large degree of divergence of other sequences involved in transcriptional regulation, chromatin state and higher order chromatin organization. Our results illuminate the wide range of evolutionary forces acting on genes and their regulatory regions, and provide a general resource for research into mammalian biology and mechanisms of human diseases. PMID:25409824

  3. A comparative encyclopedia of DNA elements in the mouse genome.

    PubMed

    Yue, Feng; Cheng, Yong; Breschi, Alessandra; Vierstra, Jeff; Wu, Weisheng; Ryba, Tyrone; Sandstrom, Richard; Ma, Zhihai; Davis, Carrie; Pope, Benjamin D; Shen, Yin; Pervouchine, Dmitri D; Djebali, Sarah; Thurman, Robert E; Kaul, Rajinder; Rynes, Eric; Kirilusha, Anthony; Marinov, Georgi K; Williams, Brian A; Trout, Diane; Amrhein, Henry; Fisher-Aylor, Katherine; Antoshechkin, Igor; DeSalvo, Gilberto; See, Lei-Hoon; Fastuca, Meagan; Drenkow, Jorg; Zaleski, Chris; Dobin, Alex; Prieto, Pablo; Lagarde, Julien; Bussotti, Giovanni; Tanzer, Andrea; Denas, Olgert; Li, Kanwei; Bender, M A; Zhang, Miaohua; Byron, Rachel; Groudine, Mark T; McCleary, David; Pham, Long; Ye, Zhen; Kuan, Samantha; Edsall, Lee; Wu, Yi-Chieh; Rasmussen, Matthew D; Bansal, Mukul S; Kellis, Manolis; Keller, Cheryl A; Morrissey, Christapher S; Mishra, Tejaswini; Jain, Deepti; Dogan, Nergiz; Harris, Robert S; Cayting, Philip; Kawli, Trupti; Boyle, Alan P; Euskirchen, Ghia; Kundaje, Anshul; Lin, Shin; Lin, Yiing; Jansen, Camden; Malladi, Venkat S; Cline, Melissa S; Erickson, Drew T; Kirkup, Vanessa M; Learned, Katrina; Sloan, Cricket A; Rosenbloom, Kate R; Lacerda de Sousa, Beatriz; Beal, Kathryn; Pignatelli, Miguel; Flicek, Paul; Lian, Jin; Kahveci, Tamer; Lee, Dongwon; Kent, W James; Ramalho Santos, Miguel; Herrero, Javier; Notredame, Cedric; Johnson, Audra; Vong, Shinny; Lee, Kristen; Bates, Daniel; Neri, Fidencio; Diegel, Morgan; Canfield, Theresa; Sabo, Peter J; Wilken, Matthew S; Reh, Thomas A; Giste, Erika; Shafer, Anthony; Kutyavin, Tanya; Haugen, Eric; Dunn, Douglas; Reynolds, Alex P; Neph, Shane; Humbert, Richard; Hansen, R Scott; De Bruijn, Marella; Selleri, Licia; Rudensky, Alexander; Josefowicz, Steven; Samstein, Robert; Eichler, Evan E; Orkin, Stuart H; Levasseur, Dana; Papayannopoulou, Thalia; Chang, Kai-Hsin; Skoultchi, Arthur; Gosh, Srikanta; Disteche, Christine; Treuting, Piper; Wang, Yanli; Weiss, Mitchell J; Blobel, Gerd A; Cao, Xiaoyi; Zhong, Sheng; Wang, Ting; Good, Peter J; Lowdon, Rebecca F; Adams, Leslie B; Zhou, Xiao-Qiao; Pazin, Michael J; Feingold, Elise A; Wold, Barbara; Taylor, James; Mortazavi, Ali; Weissman, Sherman M; Stamatoyannopoulos, John A; Snyder, Michael P; Guigo, Roderic; Gingeras, Thomas R; Gilbert, David M; Hardison, Ross C; Beer, Michael A; Ren, Bing

    2014-11-20

    The laboratory mouse shares the majority of its protein-coding genes with humans, making it the premier model organism in biomedical research, yet the two mammals differ in significant ways. To gain greater insights into both shared and species-specific transcriptional and cellular regulatory programs in the mouse, the Mouse ENCODE Consortium has mapped transcription, DNase I hypersensitivity, transcription factor binding, chromatin modifications and replication domains throughout the mouse genome in diverse cell and tissue types. By comparing with the human genome, we not only confirm substantial conservation in the newly annotated potential functional sequences, but also find a large degree of divergence of sequences involved in transcriptional regulation, chromatin state and higher order chromatin organization. Our results illuminate the wide range of evolutionary forces acting on genes and their regulatory regions, and provide a general resource for research into mammalian biology and mechanisms of human diseases.

  4. ISOLATION OF MOUSE NEUTROPHILS

    PubMed Central

    Swamydas, Muthulekha; Luo, Yi; Dorf, Martin E.; Lionakis, Michail S.

    2015-01-01

    Neutrophils represent the first line of defense against bacterial and fungal pathogens. Indeed, patients with inherited and acquired qualitative and quantitative neutrophil defects are at high risk for developing bacterial and fungal infections and suffering adverse outcomes from these infections. Therefore, research aiming at defining the molecular factors that modulate neutrophil effector function under homeostatic conditions and during infection is essential for devising strategies to augment neutrophil function and improve the outcome of infected individuals. This unit describes a reproducible density gradient centrifugation-based protocol that can be applied in any laboratory to harvest large numbers of highly enriched and highly viable neutrophils from the bone marrow of mice both at the steady state and following infection with Candida albicans as described in UNIT 19.6. In another protocol, we also present a method that combines gentle enzymatic tissue digestion with a positive immunomagnetic selection technique or Fluorescence-activated cell sorting (FACS) to harvest highly pure and highly viable preparations of neutrophils directly from mouse tissues such as the kidney, the liver or the spleen. Finally, methods for isolating neutrophils from mouse peritoneal fluid and peripheral blood are included. Mouse neutrophils isolated by these protocols can be used for examining several aspects of cellular function ex vivo including pathogen binding, phagocytosis and killing, neutrophil chemotaxis, oxidative burst, degranulation and cytokine production, and for performing neutrophil adoptive transfer experiments. PMID:26237011

  5. RIKEN mouse genome encyclopedia.

    PubMed

    Hayashizaki, Yoshihide

    2003-01-01

    We have been working to establish the comprehensive mouse full-length cDNA collection and sequence database to cover as many genes as we can, named Riken mouse genome encyclopedia. Recently we are constructing higher-level annotation (Functional ANnoTation Of Mouse cDNA; FANTOM) not only with homology search based annotation but also with expression data profile, mapping information and protein-protein database. More than 1,000,000 clones prepared from 163 tissues were end-sequenced to classify into 159,789 clusters and 60,770 representative clones were fully sequenced. As a conclusion, the 60,770 sequences contained 33,409 unique. The next generation of life science is clearly based on all of the genome information and resources. Based on our cDNA clones we developed the additional system to explore gene function. We developed cDNA microarray system to print all of these cDNA clones, protein-protein interaction screening system, protein-DNA interaction screening system and so on. The integrated database of all the information is very useful not only for analysis of gene transcriptional network and for the connection of gene to phenotype to facilitate positional candidate approach. In this talk, the prospect of the application of these genome resourced should be discussed. More information is available at the web page: http://genome.gsc.riken.go.jp/.

  6. Functional genomics in the mouse.

    PubMed

    Perkins, Archibald S

    2002-08-01

    The mouse is the premier genetic model organism for the study of human disease and development. With the recent advances in sequencing of the human and mouse genomes, there is strong interest now in large-scale approaches to decipher the function of mouse genes using various mutagenesis technologies. This review discusses what tools are currently available for manipulating and mutagenizing the mouse genome, such as ethylnitrosourea and gene trap mutagenesis, engineered inversions and deletions using the cre-lox system, and proviral insertional mutagenesis in somatic cells, and how these are being used to uncover gene function.

  7. Isolation and analysis of mouse microglial cells.

    PubMed

    Garcia, Jenny A; Cardona, Sandra M; Cardona, Astrid E

    2014-01-01

    Microglia are mononuclear phagocytes that make up about 10% of the central nervous system (CNS). They are known for their surveillant behavior, which involves continuous monitoring of neural tissue by extending and retracting their processes. Microglial cells are derived from myeloid progenitor cells and play important roles in homeostasis as well as inflammatory and immune responses in the brain. This unit describes several microglial cell isolation protocols that can be easily adapted for projects requiring a rapid and efficient analysis of mouse microglial cells by flow cytometry. Methods for visualizing microglial cells using in situ immunohistochemistry and immunochemistry in free-floating sections are also included.

  8. SIRT1 inhibits the mouse intestinal motility and epithelial proliferation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    SIRT1 inhibits the mouse intestinal motility and epithelial proliferation. Sirtuin 1 (SIRT1), a NAD+-dependent histone deacetylase, is involved in a wide array of cellular processes, including glucose homeostasis, energy metabolism, proliferation and apoptosis, and immune response. However, it is un...

  9. Chandra Catches the `Mouse'

    NASA Technical Reports Server (NTRS)

    2004-01-01

    Astronomers have used an x-ray image to make the first detailed study of the behavior of high-energy particles around a fast moving pulsar. This image, from NASA's Chandra X-Ray Observatory (CXO), shows the shock wave created as a pulsar plows supersonically through interstellar space. These results will provide insight into theories for the production of powerful winds of matter and antimatter by pulsars. Chandra's image of the glowing cloud, known as the Mouse, shows a stubby bright column of high-energy particles, about four light years in length, swept back by the pulsar's interaction with interstellar gas. The intense source at the head of the X-ray column is the pulsar, estimated to be moving through space at about 1.3 million miles per hour. A cone-shaped cloud of radio-wave-emitting particles envelopes the x-ray column. The Mouse, a.k.a. G359.23-0.82, was discovered in 1987 by radio astronomers using the National Science Foundation's Very Large Array in New Mexico. G359.23-0.82 gets its name from its appearance in radio images that show a compact snout, a bulbous body, and a remarkable long, narrow, tail that extends for about 55 light years. NASA's Marshall Space Flight Center in Huntsville, Alabama manages the Chandler program.

  10. A Comprehensive Genetic Map of Murine Chromosome 11 Reveals Extensive Linkage Conservation between Mouse and Human

    PubMed Central

    Buchberg, A. M.; Brownell, E.; Nagata, S.; Jenkins, N. A.; Copeland, N. G.

    1989-01-01

    Interspecific backcross animals from a cross between C57BL/6J and Mus spretus mice were used to generate a comprehensive linkage map of mouse chromosome 11. The relative map positions of genes previously assigned to mouse chromosome 11 by somatic cell hybrid or genetic backcross analysis were determined (Erbb, Rel, Il-3, Csfgm, Trp53-1, Evi-2, Erba, Erbb-2, Csfg, Myhs, Cola-1, Myla, Hox-2 and Pkca). We also analyzed genes that we suspected would map to chromosome 11 by virtue of their location in human chromosomes and the known linkage homologies that exist between murine chromosome 11 and human chromosomes (Mpo, Ngfr, Pdgfr and Fms). Two of the latter genes, Mpo and Ngfr, mapped to mouse chromosome 11. Both genes also mapped to human chromosome 17, extending the degree of linkage conservation observed between human chromosome 17 and mouse chromosome 11. Pdgfr and Fms, which are closely linked to Il-3 and Csfgm in humans on chromosome 5, mapped to mouse chromosome 18 rather than mouse chromosome 11, thereby defining yet another conserved linkage group between human and mouse chromosomes. The mouse chromosome 11 linkage map generated in these studies substantially extends the framework for identifying homologous genes in the mouse that are involved in human disease, for elucidating the genes responsible for several mouse mutations, and for gaining insights into chromosome evolution and genome organization. PMID:2567264

  11. The mouse genome informatics and the mouse genome database

    SciTech Connect

    Maltais, L.J.; Blackburn, R.E.; Bradt, D.W.

    1994-09-01

    The Mouse Genome Database (MGD) is a centralized, comprehensive database of the mouse genome that includes genetic mapping data, comparative mapping data, gene descriptions, mutant phenotype descriptions, strains and allelic polymorphism data, inbred strain characteristics, physical mapping data, and molecular probes and clones data. Data in MGD are obtained from the published literature and by electronic transfer from laboratories working on large backcross panels of mice. MGD provides tools that enable the user to search the database, retrieve data, generate reports, analyze data, annotate records, and build genetic maps. The Encyclopedia of the Mouse Genome provides a graphic user interface to mouse genome data. It consists of software tools including: LinkMap, a graphic display of genetic linkage maps with the ability to magnify regions of high locus density: CytoMap, a graphic display of cytogenetic maps showing banded chromosomes with cytogenetic locations of genes and chromosomal aberrations; CATS, a catalog searching tool for text retrieval of mouse locus descriptions. These software tools provide access to the following data sets: Chromosome Committee Reports, MIT Genome Center data, GBASE reports, Mouse Locus Catalog (MLC), and Mouse Cytogenetic Mapping Data. The MGD is available to the scientific community through the World Wide Web (WWW) and Gopher. In addition GBASE can be accessed via the Internet.

  12. Cloning the laboratory mouse.

    PubMed

    Wakayama, T; Yanagimachi, R

    1999-06-01

    A brief account is given of early attempts to clone mammals (mice) by transferring cells (nuclei) of preimplantation embryos into enucleated oocytes, zygotes or blastomeres of two-cell embryos. This is followed by a brief review of recent successes using adult somatic cells: mammary gland cells for sheep, muscle cells for cattle and cumulus cells for mice. We have developed a technique for cloning the laboratory mouse by transferring cumulus cell nuclei into enucleated oocytes. With this technique, we have produced a population of over 80 cloned animals, and have carried the process over four generations. Development and fertility of these appear normal. However, the yield is very low; only approximately 1% of injected oocytes are carried to term. The challenge is now to understand the reason for this high loss. Is it a problem of technique, genomic reprogramming, somatic mutation, imprinting or incompatible cell cycle phases?

  13. A novel gene, Pog, is necessary for primordial germ cell proliferation in the mouse and underlies the germ cell deficient mutation, gcd.

    PubMed

    Agoulnik, Alexander I; Lu, Baisong; Zhu, Qichao; Truong, Cavatina; Ty, Maria T; Arango, Nelson; Chada, Kiran K; Bishop, Colin E

    2002-11-15

    Primordial germ cells (PGCs) are the precursor of the germ cells in adult gonads. They arise extra-gonadally and migrate through somatic tissues to the presumptive genital ridges, where they proliferate and differentiate into oogonia or spermatogonia cells. Abnormalities in this developmental process can cause embryonic depletion of germ cells leading to infertility in the adult. We report here that the mouse gcd (germ cell deficient) mutant phenotype, characterized by reduced numbers of PGCs and adult sterility, is due to reduced PGC proliferation rather than aberrant migration and is caused by the partial deletion of a single novel gene, Pog (proliferation of germ cells). Pog is critical for normal PGC proliferation, starting between 9.5 and 10.25 dpc when germ cells begin to migrate to the developing genital ridge. Deletion of Pog is also accompanied by reduced embryonic body weight and, on some genetic backgrounds, embryonic lethality. Thus, in addition to being necessary for PGC proliferation, Pog may have a wider significance in early embryonic development.

  14. Apoptosis in the lens anlage of the heritable lens aplastic mouse (lap mouse).

    PubMed

    Aso, S; Tashiro, M; Baba, R; Sawaki, M; Noda, S; Fujita, M

    1998-08-01

    Adult homozygous lap mice show various eye abnormalities, such as aphakia, retinal disorganization, and dysplasia of the cornea and anterior chamber. In the fetal eye of a homozygous lap mouse, the lens placode seems to develop normally. However, the lens vesicle progresses abnormally to form a mass of cells without a cavity, and the mass vanishes soon afterward. We examined cell death in the lens anlage of this mutant. The lens anlagen of homozygous lap and normal mice from days 10 to 12 of gestation were observed by light microscopy after DNA end-labeling by immunohistochemistry and by transmission electron microscopy. By light microscopy, a slight frequency of cell death was detected in the lens anlage encircling the surface ectoderm and in the anlage or in the anlage of both homozygous lap mice and normal mice at day 10 of gestation. Cell death was seen in the lens anlage encircling the surface ectoderm in the normal mouse and sporadically in the anlage of the homozygous lap mouse at day 10.5 of gestation. Cell death was visible at the area of the lens vesicle attached to the surface ectoderm and encircling the surrounding surface ectoderm in the normal mouse, and in the lens anlage encircling the surface ectoderm and the apex areas of the lens anlage in the homozygous lap mouse at day 11 of gestation. At day 12 of gestation, almost no cell death was observed in the lens anlage of the normal mouse. However, extensive areas of cell death were still seen in the lens anlage at its apex, at the inner region, and encircling the surface ectoderm in the homozygous lap mouse. Electron microscopic observation showed that the dead cells observed in the lens anlagen by light microscopy in normal and lap mice are the result of apoptosis. In lap mice, cells with cytoplasmic condensation were observed mainly at days 10 and 10.5 of gestation. Many apoptotic bodies which had been phagocytosed by adjacent cells were seen predominantly at day 11 of gestation. At day 12 of

  15. Retinoic acid regulates Kit translation during spermatogonial differentiation in the mouse

    PubMed Central

    Busada, Jonathan T.; Chappell, Vesna A.; Niedenberger, Bryan A.; Kaye, Evelyn P.; Keiper, Brett D.; Hogarth, Cathryn A.; Geyer, Christopher B.

    2014-01-01

    In the testis, a subset of spermatogonia retains stem cell potential, while others differentiate to eventually become spermatozoa. This delicate balance must be maintained, as defects can result in testicular cancer or infertility. Currently, little is known about the gene products and signaling pathways directing these critical cell fate decisions. Retinoic acid (RA) is a requisite driver of spermatogonial differentiation and entry into meiosis, yet the mechanisms activated downstream are undefined. Here, we determined a requirement for RA in the expression of KIT, a receptor tyrosine kinase essential for spermatogonial differentiation. We found that RA signaling utilized the PI3K/AKT/mTOR signaling pathway to induce the efficient translation of mRNAs for Kit, which are present but not translated in undifferentiated spermatogonia. Our findings provide an important molecular link between a morphogen (RA) and the expression of KIT protein, which together direct the differentiation of spermatogonia throughout the male reproductive lifespan. PMID:25446031

  16. Whole mouse cryo-imaging

    NASA Astrophysics Data System (ADS)

    Wilson, David; Roy, Debashish; Steyer, Grant; Gargesha, Madhusudhana; Stone, Meredith; McKinley, Eliot

    2008-03-01

    The Case cryo-imaging system is a section and image system which allows one to acquire micron-scale, information rich, whole mouse color bright field and molecular fluorescence images of an entire mouse. Cryo-imaging is used in a variety of applications, including mouse and embryo anatomical phenotyping, drug delivery, imaging agents, metastastic cancer, stem cells, and very high resolution vascular imaging, among many. Cryo-imaging fills the gap between whole animal in vivo imaging and histology, allowing one to image a mouse along the continuum from the mouse -> organ -> tissue structure -> cell -> sub-cellular domains. In this overview, we describe the technology and a variety of exciting applications. Enhancements to the system now enable tiled acquisition of high resolution images to cover an entire mouse. High resolution fluorescence imaging, aided by a novel subtraction processing algorithm to remove sub-surface fluorescence, makes it possible to detect fluorescently-labeled single cells. Multi-modality experiments in Magnetic Resonance Imaging and Cryo-imaging of a whole mouse demonstrate superior resolution of cryo-images and efficiency of registration techniques. The 3D results demonstrate the novel true-color volume visualization tools we have developed and the inherent advantage of cryo-imaging in providing unlimited depth of field and spatial resolution. The recent results continue to demonstrate the value cryo-imaging provides in the field of small animal imaging research.

  17. Paracrine Mechanisms Involved in the Control of Early Stages of Mammalian Spermatogenesis

    PubMed Central

    Rossi, Pellegrino; Dolci, Susanna

    2013-01-01

    Within the testis, Sertoli-cell is the primary target of pituitary FSH. Several growth factors have been described to be produced specifically by Sertoli cells and modulate male germ cell development through paracrine mechanisms. Some have been shown to act directly on spermatogonia such as GDNF, which acts on self-renewal of spermatogonial stem cells (SSCs) while inhibiting their differentiation; BMP4, which has both a proliferative and differentiative effect on these cells, and KIT ligand (KL), which stimulates the KIT tyrosine-kinase receptor expressed by differentiating spermatogonia (but not by SSCs). KL not only controls the proliferative cycles of KIT-positive spermatogonia, but it also stimulates the expression of genes that are specific of the early phases of meiosis, whereas the expression of typical spermatogonial markers is down-regulated. On the contrary, FGF9 acts as a meiotic inhibiting substance both in fetal gonocytes and in post-natal spermatogonia through the induction of the RNA-binding protein NANOS2. Vitamin A, which is metabolized to Retinoic Acid in Sertoli cells, controls both SSCs differentiation through KIT induction and NANOS2 inhibition, and meiotic entry of differentiating spermatogonia through STRA8 upregulation. PMID:24324457

  18. Neuregulins are essential for spermatogonial proliferation and meiotic initiation in neonatal mouse testis.

    PubMed

    Zhang, JiDong; Eto, Ko; Honmyou, Asuka; Nakao, Kazuki; Kiyonari, Hiroshi; Abé, Shin-ichi

    2011-08-01

    The transition from mitosis to meiosis is unique to germ cells. In murine embryonic ovaries and juvenile testes, retinoic acid (RA) induces meiosis via the stimulated by retinoic acid gene 8 (Stra8), but its molecular pathway requires elucidation. We present genetic evidence in vivo and in vitro that neuregulins (NRGs) are essential for the proliferation of spermatogonia and the initiation of meiosis. Tamoxifen (TAM) was injected into 14-day post-partum (dpp) Sertoli cell-specific conditional Nrg1(Ser-/-) mutant mice. TAM induced testis degeneration, suppressed BrdU incorporation into spermatogonia and pre-leptotene primary spermatocytes, and decreased and increased the number of STRA8-positive and TUNEL-positive cells, respectively. In testicular organ cultures from 5-6 dpp wild-type mice and cultures of their re-aggregated spermatogonia and Sertoli cells, FSH, RA [all-trans-retinoic acid (ATRA), AM580, 9-cis-RA] and NRG1 promoted spermatogonial proliferation and meiotic initiation. However, TAM treatment of testicular organ cultures from the Nrg1(Ser-/-) mutants suppressed spermatogonial proliferation and meiotic initiation that was promoted by FSH or AM580. In re-aggregated cultures of purified spermatogonia, NRG1, NRG3, ATRA and 9-cis-RA promoted their proliferation and meiotic initiation, but neither AM580 nor FSH did. In addition, FSH, RAs and NRG1 promoted Nrg1 and Nrg3 mRNA expression in Sertoli cells. These results indicate that in juvenile testes RA and FSH induced meiosis indirectly through Sertoli cells when NRG1 and NRG3 were upregulated, as NRG1 amplified itself and NRG3. The amplified NRG1 and NRG3 directly induced meiosis in spermatogonia. In addition, ATRA and 9-cis-RA activated spermatogonia directly and promoted their proliferation and eventually meiotic initiation.

  19. No evidence for transgenerational genomic instability in the F1 or F2 descendants of Muta™Mouse males exposed to N-ethyl-N-nitrosourea.

    PubMed

    O'Brien, Jason M; Williams, Andrew; Gingerich, John; Douglas, George R; Marchetti, Francesco; Yauk, Carole L

    2013-01-01

    Exposure of male mice to genotoxic agents can increase mutation frequencies in their unexposed descendants. This phenomenon, known as transgenerational genomic instability (TGI), can persist for several generations. However, little is known about the underlying mechanisms. Chemically-induced TGI has been demonstrated in non-coding unstable tandem repeat DNA regions, but it is unclear whether it extends to other genetic endpoints. We investigated whether exposure of Muta™Mouse males to a single dose of 75mg/kg N-ethyl-N-nitrosourea (ENU) increased the spontaneous frequency of gene mutations or chromosome damage in their offspring. Treated males were mated with untreated females 3 days, 6 weeks or 10 weeks post-exposure to produce the F1 generation. Offspring were thus conceived from germ cells exposed to ENU as mature spermatozoa, dividing spermatogonia, or spermatogonial stem cells, respectively. F2 mice were generated by mating F1 descendants with untreated partners. Mutations in the lacZ transgene were quantified in bone marrow and micronucleus frequencies were evaluated in red blood cells by flow-cytometry for all F0 and their descendants. LacZ mutant frequencies were also determined in sperm for all exposed males and their male descendants. In F0 males, lacZ mutant frequencies were significantly increased in bone marrow at least 10-fold at all three time points investigated. In sperm, lacZ mutant frequency was significantly increased 7-11-fold after exposure of dividing and stem cell spermatogonia, but not in replication-deficient haploid sperm. Micronucleus frequencies assessed two days after ENU treatment were increased 5-fold in F0 males, but returned to control levels after 10 weeks. Despite the strong mutagenic response in F0 males, pre- and post-meiotic ENU exposure did not significantly increase lacZ mutant or micronucleus frequencies in F1 or F2 offspring. These findings suggest that TGI may not extend to all genetic endpoints and that further

  20. The Mouse That Roared.

    ERIC Educational Resources Information Center

    Paley, Vivian Gussin

    2000-01-01

    Discusses the animal fables of Leo Lionni and recalls a kindergarten class's involvement with his books and their characters. Considers his themes of fairness, friendship, individual decisions versus group decisions, and conformity. (LRW)

  1. Computer Workstation: Pointer/Mouse

    MedlinePlus

    ... and long term use. Potential Hazards: When the sensitivity for the input device is not appropriately set, ... provide adequate control. A mouse that has insufficient sensitivity may require large deviation of the wrist to ...

  2. Mouse models for cancer research

    PubMed Central

    Zhang, Wei; Moore, Lynette; Ji, Ping

    2011-01-01

    Mouse models of cancer enable researchers to learn about tumor biology in complicated and dynamic physiological systems. Since the development of gene targeting in mice, cancer biologists have been among the most frequent users of transgenic mouse models, which have dramatically increased knowledge about how cancers form and grow. The Chinese Journal of Cancer will publish a series of papers reporting the use of mouse models in studying genetic events in cancer cases. This editorial is an overview of the development and applications of mouse models of cancer and directs the reader to upcoming papers describing the use of these models to be published in coming issues, beginning with three articles in the current issue. PMID:21352691

  3. Mouse Models of Gastric Carcinogenesis

    PubMed Central

    Yu, Sungsook; Yang, Mijeong

    2014-01-01

    Gastric cancer is one of the most common cancers in the world. Animal models have been used to elucidate the details of the molecular mechanisms of various cancers. However, most inbred strains of mice have resistance to gastric carcinogenesis. Helicobacter infection and carcinogen treatment have been used to establish mouse models that exhibit phenotypes similar to those of human gastric cancer. A large number of transgenic and knockout mouse models of gastric cancer have been developed using genetic engineering. A combination of carcinogens and gene manipulation has been applied to facilitate development of advanced gastric cancer; however, it is rare for mouse models of gastric cancer to show aggressive, metastatic phenotypes required for preclinical studies. Here, we review current mouse models of gastric carcinogenesis and provide our perspectives on future developments in this field. PMID:25061535

  4. Reactivity of mouse antibodies against bromelain-treated mouse erythrocytes with thrombin-treated mouse platelets.

    PubMed Central

    Kawaguchi, S

    1989-01-01

    The reactivity of mouse antibodies against bromelain-treated mouse erythrocytes (BrMRBC) with mouse platelets before and after thrombin treatment was assessed by flow cytometry. Anti-BrMRBC antibodies could bind to thrombin-treated platelets, although normal platelets were also weakly reactive with the antibodies. The binding of anti-BrMRBC antibodies to platelets was confirmed by complement-dependent lysis. It is suggested that thrombin-activated platelets may be a real target for anti-BrMRBC antibodies. PMID:2467876

  5. Recent segmental and gene duplications in the mouse genome

    PubMed Central

    Cheung, Joseph; Wilson, Michael D; Zhang, Junjun; Khaja, Razi; MacDonald, Jeffrey R; Heng, Henry HQ; Koop, Ben F; Scherer, Stephen W

    2003-01-01

    Background The high quality of the mouse genome draft sequence and its associated annotations are an invaluable biological resource. Identifying recent duplications in the mouse genome, especially in regions containing genes, may highlight important events in recent murine evolution. In addition, detecting recent sequence duplications can reveal potentially problematic regions of the genome assembly. We use BLAST-based computational heuristics to identify large (≥ 5 kb) and recent (≥ 90% sequence identity) segmental duplications in the mouse genome sequence. Here we present a database of recently duplicated regions of the mouse genome found in the mouse genome sequencing consortium (MGSC) February 2002 and February 2003 assemblies. Results We determined that 33.6 Mb of 2,695 Mb (1.2%) of sequence from the February 2003 mouse genome sequence assembly is involved in recent segmental duplications, which is less than that observed in the human genome (around 3.5-5%). From this dataset, 8.9 Mb (26%) of the duplication content consisted of 'unmapped' chromosome sequence. Moreover, we suspect that an additional 18.5 Mb of sequence is involved in duplication artifacts arising from sequence misassignment errors in this genome assembly. By searching for genes that are located within these regions, we identified 675 genes that mapped to duplicated regions of the mouse genome. Sixteen of these genes appear to have been duplicated independently in the human genome. From our dataset we further characterized a 42 kb recent segmental duplication of Mater, a maternal-effect gene essential for embryogenesis in mice. Conclusion Our results provide an initial analysis of the recently duplicated sequence and gene content of the mouse genome. Many of these duplicated loci, as well as regions identified to be involved in potential sequence misassignment errors, will require further mapping and sequencing to achieve accuracy. A Genome Browser database was set up to display the

  6. Development of a mouse-feline chimeric antibody against feline tumor necrosis factor-alpha

    PubMed Central

    DOKI, Tomoyoshi; TAKANO, Tomomi; HOHDATSU, Tsutomu

    2016-01-01

    Feline infectious peritonitis (FIP) is a fatal inflammatory disease caused by FIP virus infection. Feline tumor necrosis factor (fTNF)-alpha is closely involved in the aggravation of FIP pathology. We previously described the preparation of neutralizing mouse anti-fTNF-alpha monoclonal antibody (mAb 2–4) and clarified its role in the clinical condition of cats with FIP using in vitro systems. However, administration of mouse mAb 2–4 to cat may lead to a production of feline anti-mouse antibodies. In the present study, we prepared a mouse-feline chimeric mAb (chimeric mAb 2–4) by fusing the variable region of mouse mAb 2–4 to the constant region of feline antibody. The chimeric mAb 2–4 was confirmed to have fTNF-alpha neutralization activity. Purified mouse mAb 2–4 and chimeric mAb 2–4 were repeatedly administered to cats, and the changes in the ability to induce feline anti-mouse antibody response were investigated. In the serum of cats treated with mouse mAb 2–4, feline anti-mouse antibody production was induced, and the fTNF-alpha neutralization effect of mouse mAb 2–4 was reduced. In contrast, in cats treated with chimeric mAb 2–4, the feline anti-mouse antibody response was decreased compared to that of mouse mAb 2–4-treated cats. PMID:27264736

  7. Construction of mouse adenovirus type 1 mutants.

    PubMed

    Cauthen, Angela N; Welton, Amanda R; Spindler, Katherine R

    2007-01-01

    Mouse adenovirus provides a model for studying adenovirus pathogenesis in the natural host. The ability to make viral mutants allows the investigation of specific mouse adenoviral gene contributions to virus-host interactions. Methods for propagation and titration of wild-type mouse adenovirus, production of viral DNA and viral DNA-protein complex, and transfection of mouse cells to obtain mouse adenovirus mutants are described in this chapter. Plaque purification, propagation, and titration of the mutant viruses are also presented.

  8. Mink-mouse interspecific hybridomas.

    PubMed

    Ufimtseva, E G; Galakhar, N L; Matjakhina, L D; Khlebodarova, T M; Djatchenko, S N

    1991-08-01

    Mink-mouse interspecific hybridomas were produced by fusion of the american mink spleen cells with the NSO cells. Seven cloned lines of the mink-mouse hybridoma were isolated, and their functional mink Ig secretion and karyological characteristics are given. During cytogenetic analysis of mink-mouse hybridoma cell lines, we observed the elimination of mink chromosomes, and inter- and intralineral variability of the numbers of the cells with particular quantities of mink DNA. We did not find that the characteristic peculiarities of mink DNA distribution in the hybridoma cell lines had any bearing upon the secretion or non-secretion of mink Ig. There was no synthesis of lambda-L-chains of mink Ig in line 7 cells because the line lost the lambda-gene. With the aid of in situ hybridization with 3H-labeled total mink DNA, a considerable transformation of hybridoma cell karyotype was observed. Multiple integration of the mink DNA into mouse chromosomes and the appearance of chromosomes not characteristic for either the mink or mouse parent cells were noted. Increasing numbers of cells with translocations of mink chromosomes fragments into mouse chromosomes were found in the hybridoma lines cultivated for lengthy periods. PMID:1937502

  9. Mouse genetics: Catalogue and scissors

    PubMed Central

    Sung, Young Hoon; Baek, In-Jeoung; Seong, Je Kyung; Kim, Jin-Soo; Lee, Han-Woong

    2012-01-01

    Phenotypic analysis of gene-specific knockout (KO) mice has revolutionized our understanding of in vivo gene functions. As the use of mouse embryonic stem (ES) cells is inevitable for conventional gene targeting, the generation of knockout mice remains a very time-consuming and expensive process. To accelerate the large-scale production and phenotype analyses of KO mice, international efforts have organized global consortia such as the International Knockout Mouse Consortium (IKMC) and International Mouse Phenotype Consortium (IMPC), and they are persistently expanding the KO mouse catalogue that is publicly available for the researches studying specific genes of interests in vivo. However, new technologies, adopting zinc-finger nucleases (ZFNs) or Transcription Activator-Like Effector (TALE) Nucleases (TALENs) to edit the mouse genome, are now emerging as valuable and effective shortcuts alternative for the conventional gene targeting using ES cells. Here, we introduce the recent achievement of IKMC, and evaluate the significance of ZFN/TALEN technology in mouse genetics. [BMB Reports 2012; 45(12): 686-692] PMID:23261053

  10. Home Fires Involving Grills

    MedlinePlus

    ... fires were fueled by gas while 13% used charcoal or other solid fuel. Gas grills were involved ... structure fires and 4,300 outdoor fires annually. Charcoal or other solid-fueled grills were involved in ...

  11. Affective Involvement Instrument.

    ERIC Educational Resources Information Center

    Lemlech, Johanna K.

    1970-01-01

    The Affective Involvement Instrument (AII) describes and classifies affective involvement in the process of decision-making as it occurs during classroom activities such as role-playing or group discussions. The thirty-celled instrument behaviorizes the six processes involved in decision-making and combines them with the taxonomic levels of the…

  12. Gubernatorial Involvement in Education.

    ERIC Educational Resources Information Center

    Hines, Edward R.

    This research on 12 States' gubernatorial involvement in State educational policy formation investigates four functional stages of that involvement--issue definition, proposal formulation, support mobilization, and decision enactment. Drawing on the Educational Governance Project information and interviews, a gubernatorial involvement index was…

  13. Targeted Disruption of miR-17-92 Impairs Mouse Spermatogenesis by Activating mTOR Signaling Pathway

    PubMed Central

    Xie, Raoying; Lin, Xiaolin; Du, Tao; Xu, Kang; Shen, Hongfen; Wei, Fang; Hao, Weichao; Lin, Taoyan; Lin, Xia; Qin, Yujuan; Wang, Huiyan; Chen, Lin; Yang, Sheng; Yang, Jie; Rong, Xiaoxiang; Yao, Kaitai; Xiao, Dong; Jia, Junshuang; Sun, Yan

    2016-01-01

    Abstract The miR-17-92 cluster and its 6 different mature microRNAs, including miR-17, miR-18a, miR-19a, miR-20a, miR-19b-1, and miR-92a, play important roles in embryo development, immune system, kidney and heart development, adipose differentiation, aging, and tumorigenicity. Currently, increasing evidence indicates that some members of miR-17-92 cluster may be critical players in spermatogenesis, including miR-17, miR-18a, and miR-20a. However, the roles and underlying mechanisms of miR-17-92 in spermatogenesis remain largely unknown. Our results showed that the targeted disruption of miR-17-92 in the testes of adult mice resulted in severe testicular atrophy, empty seminiferous tubules, and depressed sperm production. This phenotype is partly because of the reduced number of spermatogonia and spermatogonial stem cells, and the significantly increased germ cell apoptosis in the testes of miR-17-92-deficient mice. In addition, overactivation of the mammalian target of rapamycin signaling pathway and upregulation of the pro-apoptotic protein Bim, Stat3, c-Kit, and Socs3 were also observed in miR-17-92-deficient mouse testes, which might be, at least partially if not all, responsible for the aforementioned phenotypic changes in mutant testes. Taken together, these findings suggest that miR-17-92 is essential for normal spermatogenesis in mice. PMID:26886608

  14. Precise and in situ genetic humanization of 6 Mb of mouse immunoglobulin genes.

    PubMed

    Macdonald, Lynn E; Karow, Margaret; Stevens, Sean; Auerbach, Wojtek; Poueymirou, William T; Yasenchak, Jason; Frendewey, David; Valenzuela, David M; Giallourakis, Cosmas C; Alt, Frederick W; Yancopoulos, George D; Murphy, Andrew J

    2014-04-01

    Genetic humanization, which involves replacing mouse genes with their human counterparts, can create powerful animal models for the study of human genes and diseases. One important example of genetic humanization involves mice humanized for their Ig genes, allowing for human antibody responses within a mouse background (HumAb mice) and also providing a valuable platform for the generation of fully human antibodies as therapeutics. However, existing HumAb mice do not have fully functional immune systems, perhaps because of the manner in which they were genetically humanized. Heretofore, most genetic humanizations have involved disruption of the endogenous mouse gene with simultaneous introduction of a human transgene at a new and random location (so-called KO-plus-transgenic humanization). More recent efforts have attempted to replace mouse genes with their human counterparts at the same genetic location (in situ humanization), but such efforts involved laborious procedures and were limited in size and precision. We describe a general and efficient method for very large, in situ, and precise genetic humanization using large compound bacterial artificial chromosome-based targeting vectors introduced into mouse ES cells. We applied this method to genetically humanize 3-Mb segments of both the mouse heavy and κ light chain Ig loci, by far the largest genetic humanizations ever described. This paper provides a detailed description of our genetic humanization approach, and the companion paper reports that the humoral immune systems of mice bearing these genetically humanized loci function as efficiently as those of WT mice.

  15. The Trp53-Trp53inp1-Tnfrsf10b Pathway Regulates the Radiation Response of Mouse Spermatogonial Stem Cells

    PubMed Central

    Ishii, Kei; Ishiai, Masamichi; Morimoto, Hiroko; Kanatsu-Shinohara, Mito; Niwa, Ohtsura; Takata, Minoru; Shinohara, Takashi

    2014-01-01

    Summary Germ cells are thought to exhibit a unique DNA damage response that differs from that of somatic stem cells, and previous studies suggested that Trp53 is not involved in the survival of spermatogonial stem cells (SSCs) after irradiation. Here, we report a critical role for the Trp53-Trp53inp1-Tnfrsf10b pathway during radiation-induced SSC apoptosis. Spermatogonial transplantation revealed that Trp53 deficiency increased the survival of SSCs after irradiation. Although Bbc3, a member of the intrinsic apoptotic pathway, was implicated in apoptosis of germ and somatic stem cells, Bbc3 depletion inhibited apoptosis in committed spermatogonia, but not in SSCs. In contrast, inhibition of Tnfrsf10b, an extrinsic apoptosis regulator, rescued SSCs. Tnfrsf10b, whose deficiency protected SSCs, was upregulated by Trp53inp1 upon irradiation. These results suggest that the Trp53-Trp53inp1-Tnfrsf10b pathway responds to genotoxic damage in SSCs and that stem and progenitor cells exhibit distinct DNA damage responses in self-renewing tissue. PMID:25358794

  16. Mammalian target of rapamycin complex 1 (mTORC1) Is required for mouse spermatogonial differentiation in vivo.

    PubMed

    Busada, Jonathan T; Niedenberger, Bryan A; Velte, Ellen K; Keiper, Brett D; Geyer, Christopher B

    2015-11-01

    Spermatogonial stem cells (SSCs) must balance self-renewal with production of transit-amplifying progenitors that differentiate in response to retinoic acid (RA) before entering meiosis. This self-renewal vs. differentiation spermatogonial fate decision is critical for maintaining tissue homeostasis, as imbalances cause spermatogenesis defects that can lead to human testicular cancer or infertility. A great deal of effort has been exerted to understand how the SSC population is maintained. In contrast, little is known about the essential program of differentiation initiated by retinoic acid (RA) that precedes meiosis, and the pathways and proteins involved are poorly defined. We recently reported a novel role for RA in stimulating the PI3/AKT/mTOR kinase signaling pathway to activate translation of repressed mRNAs such as Kit. Here, we examined the requirement for mTOR complex 1 (mTORC1) in mediating the RA signal to direct spermatogonial differentiation in the neonatal testis. We found that in vivo inhibition of mTORC1 by rapamycin blocked spermatogonial differentiation, which led to an accumulation of undifferentiated spermatogonia. In addition, rapamycin also blocked the RA-induced translational activation of mRNAs encoding KIT, SOHLH1, and SOHLH2 without affecting expression of STRA8. These findings highlight dual roles for RA in germ cell development - transcriptional activation of genes, and kinase signaling to stimulate translation of repressed messages required for spermatogonial differentiation.

  17. Mapping mouse hemangioblast maturation from headfold stages

    PubMed Central

    Rhee, Jerry M.; Iannaccone, Philip M.

    2012-01-01

    The mouse posterior primitive streak at neural plate/headfold stages (NP/HF, ~7.5dpc–8dpc) represents an optimal window from which hemangioblasts can be isolated. We performed immunohistochemistry on this domain using established monoclonal antibodies for proteins that affect blood and endothelial fates. We demonstrate that HoxB4 and GATA1 are the first set of markers that segregate independently to endothelial or blood populations during NP/HF stages of mouse embryonic development. In a subset of cells, both proteins are co-expressed and immunoreactivities appear mutually excluded within nuclear spaces. We searched for this particular state at later sites of hematopoietic stem cell emergence, viz., the aorta-gonadmesonephros (AGM) and the fetal liver at 10.5–11.5dpc, and found that only a rare number of cells displayed this character. Based on this spatial-temporal argument, we propose that the earliest blood progenitors emerge either directly from the epiblast or through segregation within the allantoic core domain (ACD) through reduction of cell adhesion and pSmad1/5 nuclear signaling, followed by a stochastic decision toward a blood or endothelial fate that involves GATA1 and HoxB4, respectively. A third form in which binding distributions are balanced may represent a common condition shared by hemangioblasts and HSCs. We developed a heuristic model of hemangioblast maturation, in part, to be explicit about our assumptions. PMID:22426104

  18. Mouse Models of Frailty: an Emerging Field.

    PubMed

    Seldeen, K L; Pang, M; Troen, B R

    2015-10-01

    Frailty is highly prevalent in the elderly, increasing the risk of poor outcomes that include falls, incident disability, hospitalization, and mortality. Thus, a great need exists to characterize the underlying mechanisms and ultimately identify strategies that prevent, delay, and even reverse frailty. Mouse models can provide insight into molecular mechanisms of frailty by reducing variability in lifestyle and genetic factors that can complicate interpretation of human clinical data. Frailty, generally recognized as a syndrome involving reduced homeostatic reserve in response to physiologic challenges and increasing susceptibility to poor health outcomes, is predominantly assessed using two independent strategies, integrated phenotype and deficit accumulation. The integrated phenotype defines frailty by the presentation of factors affecting functional capacity such as weight loss, exhaustion, low activity levels, slow gait, and grip strength. The deficit accumulation paradigm draws parameters from a greater range of physiological systems, such as the ability to perform daily activities, coordination and gait, mental components, physiological problems, and history and presence of medical morbidities. This strategic division also applies within the emerging field of mouse frailty models, with both methodologies showing usefulness in providing insight into physiologic mechanisms and testing interventions. Our review will explore the strategies used, caveats in methodology, and future directions in the application of animal models for the study of the frailty syndrome.

  19. Calpain 11 Is Unique to Mouse Spermatogenic Cells

    PubMed Central

    Ben-Aharon, Irit; Brown, Paula R.; Shalgi, Ruth; Eddy, Edward M.

    2007-01-01

    The calpains are a family of calcium-dependent thiol proteases involved in intracellular processing of proteins. They occur as heterodimers containing one of various large subunits and a common small subunit. Some of the large subunits are expressed ubiquitously and others are expressed in a restricted set of tissues. We have cloned the cDNA for mouse calpain 11 and demonstrated that it is expressed specifically in the mouse testis. The mRNA begins to accumulate in the testis between days 14 and 16 days after birth, corresponding to the period of pachytene spermatocyte development. The protein is detected by day 18 after birth, during mid to late pachytene spermatocyte development, and is present in the acrosomal region of spermatozoa from the cauda epididymis. The expression of calpain 11 during spermatogenesis and its localization is spermatozoa suggest that it is involved in regulating calcium-dependent signal transduction events during meiosis and sperm functional processes. PMID:16541461

  20. Expression, function and regulation of mouse cytochrome P450 enzymes: comparison with human P450 enzymes.

    PubMed

    Hrycay, E G; Bandiera, S M

    2009-12-01

    The present review focuses on the expression, function and regulation of mouse cytochrome P450 (Cyp) enzymes. Information compiled for mouse Cyp enzymes is compared with data collected for human CYP enzymes. To date, approximately 40 pairs of orthologous mouse-human CYP genes have been identified that encode enzymes performing similar metabolic functions. Recent knowledge concerning the tissue expression of mouse Cyp enzymes from families 1 to 51 is summarized. The catalytic activities of microsomal, mitochondrial and recombinant mouse Cyp enzymes are discussed and their involvement in the metabolism of exogenous and endogenous compounds is highlighted. The role of nuclear receptors, such as the aryl hydrocarbon receptor, constitutive androstane receptor and pregnane X receptor, in regulating the expression of mouse Cyp enzymes is examined. Targeted disruption of selected Cyp genes has generated numerous Cyp null mouse lines used to decipher the role of Cyp enzymes in metabolic, toxicological and biological processes. In conclusion, the laboratory mouse is an indispensable model for exploring human CYP-mediated activities.

  1. Preventive effect of antihistaminics on mouse skin photosensitization with hematoporphyrin derivative

    NASA Astrophysics Data System (ADS)

    Fu, Nai-wu; Yan, Li-xue

    1993-03-01

    Beta-carotene 100 mg/kg per day or vitamin C 50 mg/kg per day was administered orally for two days and did not prevent mouse skin photosensitization caused by hematoporphyrin derivative (HpD). However, (beta) -carotene 100 mg/kg per day administered intramuscularly for two days prevented mouse skin reaction. Cimetidine and benadryl 10 mg/kg per day, P.O.X 2, effectively prevented mouse skin reaction. This suggests histamine may be involved in skin photoreaction induced by HpD.

  2. 10. international mouse genome conference

    SciTech Connect

    Meisler, M.H.

    1996-12-31

    Ten years after hosting the First International Mammalian Genome Conference in Paris in 1986, Dr. Jean-Louis Guenet presided over the Tenth Conference at the Pasteur Institute, October 7--10, 1996. The 1986 conference was a satellite to the Human Gene Mapping Workshop and had approximately 50 attendees. The 1996 meeting was attended by 300 scientists from around the world. In the interim, the number of mapped loci in the mouse increased from 1,000 to over 20,000. This report contains a listing of the program and its participants, and two articles that review the meeting and the role of the laboratory mouse in the Human Genome project. More than 200 papers were presented at the conference covering the following topics: International mouse chromosome committee meetings; Mutant generation and identification; Physical and genetic maps; New technology and resources; Chromatin structure and gene regulation; Rate and hamster genetic maps; Informatics and databases; and Quantitative trait analysis.

  3. Teratology studies in the mouse.

    PubMed

    Marsden, Edward; Leroy, Mariline

    2013-01-01

    The rat is the routine species of choice as the rodent model for regulatory safety testing of xenobiotics such as medicinal products, food additives, and other chemicals. However, the rat is not always suitable for pharmacological, toxicological, immunogenic, pharmacokinetic, or even practical reasons. Under such circumstances, the mouse offers an alternative for finding a suitable rodent model acceptable to the regulatory authorities. Since all essential routes of administration are possible, the short reproductive cycle and large litter size of the mouse make it a species well adapted for use in teratology studies. Given that good quality animals, including virgin mated females, can be acquired relatively easily and inexpensively, the mouse has been used in reproductive toxicity studies for decades and study protocols are well established.

  4. Modeling metastasis in the mouse

    PubMed Central

    Bos, Paula D.; Nguyen, Don X.; Massagué, Joan

    2010-01-01

    Metastasis is a complex clinical and biological problem presently under intense study, and several model systems are in use to experimentally recapitulate and dissect the various steps of the metastatic process. Genetically engineered mouse models provide faithful renditions of events in tumor progression, angiogenesis, and local invasion that set the stage for metastasis, whereas engrafting of human or mouse tumor tissues into mouse hosts has been successfully exploited to investigate metastatic dissemination and colonization of distant organs. Real-time, high-resolution microscopy in live animals, and comprehensive genetic and molecular profiling are effective tools to interrogate diverse metastatic cancer cell phenotypes as well as the metastatic tumor microenvironment in different organs. By integrating the information obtained with these complementary approaches the field is currently obtaining an unprecedented level of understanding of the biology, molecular basis, and therapeutic vulnerabilities of metastasis. PMID:20598638

  5. Involvement of opsins in mammalian sperm thermotaxis

    PubMed Central

    Pérez-Cerezales, Serafín; Boryshpolets, Sergii; Afanzar, Oshri; Brandis, Alexander; Nevo, Reinat; Kiss, Vladimir; Eisenbach, Michael

    2015-01-01

    A unique characteristic of mammalian sperm thermotaxis is extreme temperature sensitivity, manifested by the capacity of spermatozoa to respond to temperature changes of <0.0006 °C as they swim their body-length distance. The identity of the sensing system that confers this exceptional sensitivity on spermatozoa is not known. Here we show that the temperature-sensing system of mammalian spermatozoa involves opsins, known to be G-protein-coupled receptors that act as photosensors in vision. We demonstrate by molecular, immunological, and functional approaches that opsins are present in human and mouse spermatozoa at specific sites, which depend on the species and the opsin type, and that they are involved in sperm thermotaxis via two signalling pathways—the phospholipase C and the cyclic-nucleotide pathways. Our results suggest that, depending on the context and the tissue, mammalian opsins act not only as photosensors but also as thermosensors. PMID:26537127

  6. Mousepox outbreak in a laboratory mouse colony.

    PubMed

    Dick, E J; Kittell, C L; Meyer, H; Farrar, P L; Ropp, S L; Esposito, J J; Buller, R M; Neubauer, H; Kang, Y H; McKee, A E

    1996-12-01

    Mousepox was diagnosed in and eradicated from a laboratory mouse colony at the Naval Medical Research Institute. The outbreak began with increased mortality in a single room; subsequently, small numbers of animals in separate cages in other rooms were involved. Signs of disease were often mild, and overall mortality was low; BALB/cByJ mice were more severely affected, and many of them died spontaneously. Conjunctivitis was the most common clinical sign of disease in addition to occasional small, crusty scabs on sparsely haired or hairless areas of skin. Necropsy findings included conjunctivitis, enlarged spleen, and pale liver. Hemorrhage into the pyloric region of the stomach and proximal portion of the small intestine was observed in experimentally infected animals. In immune competent and immune deficient mice, the most common histologic finding was multifocal to coalescing splenic necrosis; necrosis was seen less frequently in liver, lymph nodes, and Peyer's patches. Necrosis was rarely observed in ovary, vagina, uterus, colon, or lung. Splenic necrosis often involved over 50% of the examined tissue, including white and red pulp. Hepatic necrosis was evident as either large, well-demarcated areas of coagulative necrosis or as multiple, random, interlacing bands of necrosis. Intracytoplasmic eosinophilic inclusion bodies were seen in conjunctival mucosae and haired palpebra. Ectromelia virus was confirmed as the causative agent of the epizootic by electron microscopy, immunohistochemistry, animal inoculations, serologic testing, virus isolation, and polymerase chain reaction. Serologic testing was of little value in the initial stages of the outbreak, although 6 weeks later, orthopoxvirus-specific antibody was detected in colony mice by indirect fluorescent antibody and enzyme-linked immunosorbent assay procedures. The outbreak originated from injection of mice with a contaminated, commercially produced, pooled mouse serum. The most relevant concern may be the

  7. Periodic properties of the histaminergic system of the mouse brain.

    PubMed

    Rozov, Stanislav V; Zant, Janneke C; Karlstedt, Kaj; Porkka-Heiskanen, Tarja; Panula, Pertti

    2014-01-01

    Brain histamine is involved in the regulation of the sleep-wake cycle and alertness. Despite the widespread use of the mouse as an experimental model, the periodic properties of major markers of the mouse histaminergic system have not been comprehensively characterized. We analysed the daily levels of histamine and its first metabolite, 1-methylhistamine, in different brain structures of C57BL/6J and CBA/J mouse strains, and the mRNA level and activity of histidine decarboxylase and histamine-N-methyltransferase in C57BL/6J mice. In the C57BL/6J strain, histamine release, assessed by in vivo microdialysis, underwent prominent periodic changes. The main period was 24 h peaking during the activity period. Additional 8 h periods were also observed. The release was highly positively correlated with active wakefulness, as shown by electroencephalography. In both mouse strains, tissue histamine levels remained steady for 24 h in all structures except for the hypothalamus of CBA/J mice, where 24-h periodicity was observed. Brain tissue 1-methylhistamine levels in both strains reached their maxima in the periods of activity. The mRNA level of histidine decarboxylase in the tuberomamillary nucleus and the activities of histidine decarboxylase and histamine-N-methyltransferase in the striatum and cortex did not show a 24-h rhythm, whereas in the hypothalamus the activities of both enzymes had a 12-h periodicity. These results show that the activities of histamine-metabolizing enzymes are not under simple direct circadian regulation. The complex and non-uniform temporal patterns of the histaminergic system of the mouse brain suggest that histamine is strongly involved in the maintenance of active wakefulness.

  8. MicroRNA expression in the aging mouse thymus.

    PubMed

    Ye, Yaqiong; Li, Daotong; Ouyang, Dan; Deng, Li; Zhang, Yuan; Ma, Yongjiang; Li, Yugu

    2014-09-01

    MicroRNAs (miRNAs) have been implicated in the process of aging in many model organisms, such as Caenorhabditis elegans, and in many organs, such as the mouse lung and human epididymis. However, the role of miRNAs in the thymus tissues of the aging mouse remains unclear. To address this question, we investigated the miRNA expression profiles in the thymuses of 1-, 10- and 19-month-old mice using miRNA array and qRT-PCR assays. A total of 223 mouse miRNAs were screened, and the expression levels of those miRNAs exhibited gradual increases and decreases over the course of thymus aging. Fifty miRNAs in the 10-month-old thymus and 81 miRNAs in the 19-month-old thymus were defined as differentially expressed miRNAs (p<0.05) in comparison with their levels in the 1-month-old mouse, and approximately one-third of these miRNAs were grouped within 11 miRNA clusters. Each miRNA cluster contained 2 to 5 miRNA genes, and most of the cluster members displayed similar expression patterns, being either increased or decreased. In addition, Ingenuity Pathway Analysis (IPA) software and the IPA database were used to analyze the 12 miRNAs that exhibited significant expression changes, revealing that as many as 15 pathways may be involved. Thus, our current study determined the expression profiles of miRNAs in the mouse thymus during the process of aging. The results suggested that these miRNAs could become meaningful biomarkers for studying thymus aging and that the aging-related alternations in miRNA expression may be involved in the regulation of cell proliferation, apoptosis, development and carcinogenesis/tumorigenesis.

  9. The Mouse Genome Database (MGD): mouse biology and model systems.

    PubMed

    Bult, Carol J; Eppig, Janan T; Kadin, James A; Richardson, Joel E; Blake, Judith A

    2008-01-01

    The Mouse Genome Database, (MGD, http://www.informatics.jax.org/), integrates genetic, genomic and phenotypic information about the laboratory mouse, a primary animal model for studying human biology and disease. MGD data content includes comprehensive characterization of genes and their functions, standardized descriptions of mouse phenotypes, extensive integration of DNA and protein sequence data, normalized representation of genome and genome variant information including comparative data on mammalian genes. Data within MGD are obtained from diverse sources including manual curation of the biomedical literature, direct contributions from individual investigator's laboratories and major informatics resource centers such as Ensembl, UniProt and NCBI. MGD collaborates with the bioinformatics community on the development of data and semantic standards such as the Gene Ontology (GO) and the Mammalian Phenotype (MP) Ontology. MGD provides a data-mining platform that enables the development of translational research hypotheses based on comparative genotype, phenotype and functional analyses. Both web-based querying and computational access to data are provided. Recent improvements in MGD described here include the association of gene trap data with mouse genes and a new batch query capability for customized data access and retrieval.

  10. High Involvement Work Teams.

    ERIC Educational Resources Information Center

    1996

    These three papers were presented at a symposium on high-involvement work teams moderated by Michael Leimbach at the 1996 conference of the Academy of Human Resource Development. "Beyond Training to the New Learning Environment: Workers on the High-Involvement Frontline" (Joseph Anthony Ilacqua, Carol Ann Zulauf) shows the link between an…

  11. Building Parent Involvement

    ERIC Educational Resources Information Center

    Nelson, Richard C.; Bloom, John W.

    1973-01-01

    Discussed is the rationale behind parent involvement in guidance and educational activities, together with specific suggestions for involving parents with other adults (parent advisory committees, informal coffees, Transactional analysis (groups etc.), with children (story hours, trips, demonstrations, counseling booths, testing, interviewing,…

  12. Parent Involvement Handbook.

    ERIC Educational Resources Information Center

    Caplan, Arna

    This handbook on parent involvement, designed to be used with preschool programs, was developed by the Jefferson County Public Schools in Lakewood, Colorado. Included are: (1) a general statement about parent involvement in an early childhood program, (2) a description of the Jefferson County Early Childhood Program, (3) a description of the…

  13. Categories of Parent Involvement.

    ERIC Educational Resources Information Center

    Bauch, Jerold P.

    1994-01-01

    The growing interest in effective parent involvement has produced several ways to classify or describe ways parents are or should be involved. This article reviews and evaluates Ira Gordon's systems approach, the California-based System Development Corporation's categories, Eugenia H. Berger's parental role categories, Chavkin and Williams' parent…

  14. Commericial Involvement in Intramurals.

    ERIC Educational Resources Information Center

    Maas, Gerry

    Sport in general has long had ties with commercial interests, the most popular and widespread involving publicity. Intramural sports programs, however, have not cultivated many commercial involvements in publicity. The approach in intramural sports advertising is simple. A commercial interest pays for space or time in a given communication media…

  15. International Mouse Phenotyping Consortium (IMPC) —

    Cancer.gov

    The International Mouse Phenotyping Consortium (IMPC) comprises a group of major mouse genetics research institutions along with national funding organisations formed to address the challenge of developing an encyclopedia of mammalian gene function.

  16. Lipid Extraction from Mouse Feces

    PubMed Central

    Kraus, Daniel; Yang, Qin; Kahn, Barbara B.

    2016-01-01

    The analysis of feces composition is important for the study of energy metabolism, which comprises various measurements of energy intake, energy expenditure, and energy wasting. The current protocol describes how to measure energy-dense lipids in mouse feces using a modification of the method proposed by Folch et al. (1957). PMID:27110587

  17. Mouse Models of Rheumatoid Arthritis.

    PubMed

    Caplazi, P; Baca, M; Barck, K; Carano, R A D; DeVoss, J; Lee, W P; Bolon, B; Diehl, L

    2015-09-01

    Rheumatoid arthritis (RA) is a chronic debilitating autoimmune disorder characterized by synovitis that leads to cartilage and bone erosion by invading fibrovascular tissue. Mouse models of RA recapitulate many features of the human disease. Despite the availability of medicines that are highly effective in many patient populations, autoimmune diseases (including RA) remain an area of active biomedical research, and consequently mouse models of RA are still extensively used for mechanistic studies and validation of therapeutic targets. This review aims to integrate morphologic features with model biology and cover the key characteristics of the most commonly used induced and spontaneous mouse models of RA. Induced models emphasized in this review include collagen-induced arthritis and antibody-induced arthritis. Collagen-induced arthritis is an example of an active immunization strategy, whereas antibody- induced arthritis models, such as collagen antibody-induced arthritis and K/BxN antibody transfer arthritis, represent examples of passive immunization strategies. The coverage of spontaneous models in this review is focused on the TNFΔ (ARE) mouse, in which arthritis results from overexpression of TNF-α, a master proinflammatory cytokine that drives disease in many patients.

  18. APOPTOSIS IN WHOLE MOUSE OVARIES

    EPA Science Inventory

    Apoptosis in Whole Mouse Ovaries
    Robert M. Zucker Susan C. Jeffay and Sally D. Perreault
    Reproductive Toxicology Division, National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina, 27711.

  19. Mouse Models of Diabetic Neuropathy

    PubMed Central

    O'Brien, Phillipe D.; Sakowski, Stacey A.; Feldman, Eva L.

    2014-01-01

    Diabetic peripheral neuropathy (DPN) is the most common complication of diabetes and is associated with significant morbidity and mortality. DPN is characterized by progressive, distal-to-proximal degeneration of peripheral nerves that leads to pain, weakness, and eventual loss of sensation. The mechanisms underlying DPN pathogenesis are uncertain, and other than tight glycemic control in type 1 patients, there is no effective treatment. Mouse models of type 1 (T1DM) and type 2 diabetes (T2DM) are critical to improving our understanding of DPN pathophysiology and developing novel treatment strategies. In this review, we discuss the most widely used T1DM and T2DM mouse models for DPN research, with emphasis on the main neurologic phenotype of each model. We also discuss important considerations for selecting appropriate models for T1DM and T2DM DPN studies and describe the promise of novel emerging diabetic mouse models for DPN research. The development, characterization, and comprehensive neurologic phenotyping of clinically relevant mouse models for T1DM and T2DM will provide valuable resources for future studies examining DPN pathogenesis and novel therapeutic strategies. PMID:24615439

  20. Mouse models of myelodysplastic syndromes

    PubMed Central

    Beachy, Sarah H.; Aplan, Peter D.

    2010-01-01

    Synopsis Three general approaches have been used in an attempt to model myelodysplastic syndrome (MDS) in mice, including treatment with mutagens or carcinogens, xenotransplantation of human MDS cells, and genetic engineering of mouse hematopoietic cells. Xenotransplantation of cells from MDS patients has proved difficult, possibly due to the innate characteristics of the MDS clone and microenvironmental influences, including adverse effects of a host immune response. Genetic engineering of hematopoietic cells or mice has been accomplished by in vitro transfer of genes to mouse hematopoietic cells with subsequent transplantation into an irradiated host, or by modification of the mouse germline to generate mice with altered expression of genes of interest. A number of genes have been studied using these approaches, including RUNX1, Evi1, Npm1, SALL4B, NUP98-HOXD13, BCL2/NRAS, Arid4a, Polg and Dido. This review discusses the phenotypes observed in available mouse models for MDS with a concentration on a model that leads to aberrant expression of conserved homeobox (HOX) genes that are important regulators of normal hematopoiesis. Utilizing these models of MDS should allow a more complete understanding of the disease process and provide a platform for pre-clinical testing of therapeutic approaches. PMID:20359631

  1. Mouse Cochlear Whole Mount Immunofluorescence

    PubMed Central

    Akil, Omar; Lustig, Lawrence R.

    2016-01-01

    This protocol comprises the entire process of immunofluorescence staining on mouse cochlea whole mount, starting from tissue preparation to the mounting of the tissue. This technique provides “three-dimensional” views of the stained components in order to determine the localization of a protein of interest in the tissue in its natural state and environment. PMID:27547786

  2. Postnatal histomorphogenesis of the mandible in the house mouse

    PubMed Central

    Martinez-Maza, Cayetana; Montes, Laëtitia; Lamrous, Hayat; Ventura, Jacint; Cubo, Jorge

    2012-01-01

    The mandible of the house mouse, Mus musculus, is a model structure for the study of the development and evolution of complex morphological systems. This research describes the histomorphogenesis of the house mouse mandible and analyses its biological significance from the first to the eighth postnatal weeks. Histological data allowed us to test a hypothesis concerning modularity in this structure. We measured the bone growth rates by fluorescent labelling and identified the bone tissue types through microscopic analysis of histological cross-sections of the mandible during its postnatal development. The results provide evidence for a modular structure of the mouse mandible, as the alveolar region and the ascending ramus show histological differences throughout ontogeny. The alveolar region increases in length during the first two postnatal weeks by bone growth in the posterior region, while horizontally positioned incisors preclude bone growth in the anterior region. In the fourth postnatal week, growth dynamics shows a critical change. The alveolar region drifts laterally and the ramus becomes more vertical due to the medial growth direction of the coronoid region and the lateral growth of the ventral region of the ramus. Diet changes after weaning are probably involved in these morphological changes. In this way, the development of the masticatory muscles that insert on the ascending ramus may be particularly related to this shape modeling of the house mouse mandible. PMID:22372819

  3. Eye Involvement in TSC

    MedlinePlus

    ... what we see to the brain via the optic nerve. Retinal and optic nerve involvement in TSC are well known today, ... hamartomas (non-cancerous tumors) of the retina or optic nerve. The most common type of retinal hamartoma ...

  4. CYP1A1 and CYP1A2 expression: Comparing 'humanized' mouse lines and wild-type mice; comparing human and mouse hepatoma-derived cell lines

    SciTech Connect

    Uno, Shigeyuki; Endo, Kaori; Ishida, Yuji; Tateno, Chise; Makishima, Makoto; Yoshizato, Katsutoshi; Nebert, Daniel W.

    2009-05-15

    Human and rodent cytochrome P450 (CYP) enzymes sometimes exhibit striking species-specific differences in substrate preference and rate of metabolism. Human risk assessment of CYP substrates might therefore best be evaluated in the intact mouse by replacing mouse Cyp genes with human CYP orthologs; however, how 'human-like' can human gene expression be expected in mouse tissues? Previously a bacterial-artificial-chromosome-transgenic mouse, carrying the human CYP1A1{sub C}YP1A2 locus and lacking the mouse Cyp1a1 and Cyp1a2 orthologs, was shown to express robustly human dioxin-inducible CYP1A1 and basal versus inducible CYP1A2 (mRNAs, proteins, enzyme activities) in each of nine mouse tissues examined. Chimeric mice carrying humanized liver have also been generated, by transplanting human hepatocytes into a urokinase-type plasminogen activator(+/+){sub s}evere-combined-immunodeficiency (uPA/SCID) line with most of its mouse hepatocytes ablated. Herein we compare basal and dioxin-induced CYP1A mRNA copy numbers, protein levels, and four enzymes (benzo[a]pyrene hydroxylase, ethoxyresorufin O-deethylase, acetanilide 4-hydroxylase, methoxyresorufin O-demethylase) in liver of these two humanized mouse lines versus wild-type mice; we also compare these same parameters in mouse Hepa-1c1c7 and human HepG2 hepatoma-derived established cell lines. Most strikingly, mouse liver CYP1A1-specific enzyme activities are between 38- and 170-fold higher than human CYP1A1-specific enzyme activities (per unit of mRNA), whereas mouse versus human CYP1A2 enzyme activities (per unit of mRNA) are within 2.5-fold of one another. Moreover, both the mouse and human hepatoma cell lines exhibit striking differences in CYP1A mRNA levels and enzyme activities. These findings are relevant to risk assessment involving human CYP1A1 and CYP1A2 substrates, when administered to mice as environmental toxicants or drugs.

  5. CYP1A1 and CYP1A2 expression: comparing 'humanized' mouse lines and wild-type mice; comparing human and mouse hepatoma-derived cell lines.

    PubMed

    Uno, Shigeyuki; Endo, Kaori; Ishida, Yuji; Tateno, Chise; Makishima, Makoto; Yoshizato, Katsutoshi; Nebert, Daniel W

    2009-05-15

    Human and rodent cytochrome P450 (CYP) enzymes sometimes exhibit striking species-specific differences in substrate preference and rate of metabolism. Human risk assessment of CYP substrates might therefore best be evaluated in the intact mouse by replacing mouse Cyp genes with human CYP orthologs; however, how "human-like" can human gene expression be expected in mouse tissues? Previously a bacterial-artificial-chromosome-transgenic mouse, carrying the human CYP1A1_CYP1A2 locus and lacking the mouse Cyp1a1 and Cyp1a2 orthologs, was shown to express robustly human dioxin-inducible CYP1A1 and basal versus inducible CYP1A2 (mRNAs, proteins, enzyme activities) in each of nine mouse tissues examined. Chimeric mice carrying humanized liver have also been generated, by transplanting human hepatocytes into a urokinase-type plasminogen activator(+/+)_severe-combined-immunodeficiency (uPA/SCID) line with most of its mouse hepatocytes ablated. Herein we compare basal and dioxin-induced CYP1A mRNA copy numbers, protein levels, and four enzymes (benzo[a]pyrene hydroxylase, ethoxyresorufin O-deethylase, acetanilide 4-hydroxylase, methoxyresorufin O-demethylase) in liver of these two humanized mouse lines versus wild-type mice; we also compare these same parameters in mouse Hepa-1c1c7 and human HepG2 hepatoma-derived established cell lines. Most strikingly, mouse liver CYP1A1-specific enzyme activities are between 38- and 170-fold higher than human CYP1A1-specific enzyme activities (per unit of mRNA), whereas mouse versus human CYP1A2 enzyme activities (per unit of mRNA) are within 2.5-fold of one another. Moreover, both the mouse and human hepatoma cell lines exhibit striking differences in CYP1A mRNA levels and enzyme activities. These findings are relevant to risk assessment involving human CYP1A1 and CYP1A2 substrates, when administered to mice as environmental toxicants or drugs. PMID:19285097

  6. [Pulmonary involvements of sarcoidosis].

    PubMed

    Ohmichi, M; Hiraga, Y; Hirasawa, M

    1990-01-01

    We reported about intrathoracic changes and prognosis of 686 patients with sarcoidosis diagnosed in our hospital between 1963 and 1988. We evaluated CT findings in 135 patients with sarcoidosis and found pulmonary involvements in 81. We analyzed CT findings according to the classification by Tuengerthal which classified radiographic findings combining ILO classification of pneumoconiosis and characteristic findings of bronchovascular sheath with sarcoidosis. The CT findings were as follows: small opacities (44 out of 81 cases, 54.3%), large opacities (37 cases, 46.7%). Additional findings were as follows: peribronchial marking (42 cases, 51.9%), contraction (17 cases, 21.0%), pleural involvement (9 cases, 11.1%), bulla (5 cases, 6.2%). The characteristic CT findings of serious sarcoidosis were extasis of bronchus, thickening of the bronchial wall, unclearness of vascular shadow, atelectasis and thickening of pleura. Concerning the prognosis of pulmonary involvement, according to age, patients younger than 30 years old at initial diagnosis were better than those of 30 years and over in terms of disappearance of pulmonary involvements. According to stage, patients of stage I and stage II were better than those of stage III. Among the patients we were able to observe chest X-ray findings during five years according to the character of shadow, ill-defined shadow of small opacities and rounded shadows of large opacities had a higher disappearance rate of pulmonary involvements than irregular shadows of large opacities, atelectasis and contraction.

  7. Musculoskeletal involvement in sarcoidosis*, **

    PubMed Central

    Nessrine, Akasbi; Zahra, Abourazzak Fatima; Taoufik, Harzy

    2014-01-01

    Sarcoidosis is a multisystem inflammatory disorder of unknown cause. It most commonly affects the pulmonary system but can also affect the musculoskeletal system, albeit less frequently. In patients with sarcoidosis, rheumatic involvement is polymorphic. It can be the presenting symptom of the disease or can appear during its progression. Articular involvement is dominated by nonspecific arthralgia, polyarthritis, and Löfgren's syndrome, which is defined as the presence of lung adenopathy, arthralgia (or arthritis), and erythema nodosum. Skeletal manifestations, especially dactylitis, appear mainly as complications of chronic, multiorgan sarcoidosis. Muscle involvement in sarcoidosis is rare and usually asymptomatic. The diagnosis of rheumatic sarcoidosis is based on X-ray findings and magnetic resonance imaging findings, although the definitive diagnosis is made by anatomopathological study of biopsy samples. Musculoskeletal involvement in sarcoidosis is generally relieved with nonsteroidal anti-inflammatory drugs or corticosteroids. In corticosteroid-resistant or -dependent forms of the disease, immunosuppressive therapy, such as treatment with methotrexate or anti-TNF-α, is employed. The aim of this review was to present an overview of the various types of osteoarticular and muscle involvement in sarcoidosis, focusing on their diagnosis and management. PMID:24831403

  8. Identification of a set of genes showing regionally enriched expression in the mouse brain

    PubMed Central

    D'Souza, Cletus A; Chopra, Vikramjit; Varhol, Richard; Xie, Yuan-Yun; Bohacec, Slavita; Zhao, Yongjun; Lee, Lisa LC; Bilenky, Mikhail; Portales-Casamar, Elodie; He, An; Wasserman, Wyeth W; Goldowitz, Daniel; Marra, Marco A; Holt, Robert A; Simpson, Elizabeth M; Jones, Steven JM

    2008-01-01

    Background The Pleiades Promoter Project aims to improve gene therapy by designing human mini-promoters (< 4 kb) that drive gene expression in specific brain regions or cell-types of therapeutic interest. Our goal was to first identify genes displaying regionally enriched expression in the mouse brain so that promoters designed from orthologous human genes can then be tested to drive reporter expression in a similar pattern in the mouse brain. Results We have utilized LongSAGE to identify regionally enriched transcripts in the adult mouse brain. As supplemental strategies, we also performed a meta-analysis of published literature and inspected the Allen Brain Atlas in situ hybridization data. From a set of approximately 30,000 mouse genes, 237 were identified as showing specific or enriched expression in 30 target regions of the mouse brain. GO term over-representation among these genes revealed co-involvement in various aspects of central nervous system development and physiology. Conclusion Using a multi-faceted expression validation approach, we have identified mouse genes whose human orthologs are good candidates for design of mini-promoters. These mouse genes represent molecular markers in several discrete brain regions/cell-types, which could potentially provide a mechanistic explanation of unique functions performed by each region. This set of markers may also serve as a resource for further studies of gene regulatory elements influencing brain expression. PMID:18625066

  9. Transporting mouse embryos and germplasm as frozen or unfrozen materials.

    PubMed

    Kenyon, Janet; Guan, Mo; Bogani, Debora; Marschall, Susan; Raspa, Marcello; Pickard, Amanda; Takeo, Toru; Nakagata, Naomi; Fray, Martin

    2014-01-01

    The 21st century has seen a huge proliferation in the availability of genetically altered mice. The availability of these resources has been accompanied by ever greater opportunities for international collaborations between laboratories involving the exchange of mouse strains. This exchange can involve significant costs in terms of animal welfare and transportation expenses. In an attempt to mitigate some of these costs, the mouse community has developed a battery of techniques that can be used to avoid transporting live mice. Transporting frozen embryos and sperm at liquid nitrogen (LN2 ) temperatures using dry shippers has been common practice for some time. However, current advances in this field have refined transportation procedures and introduced new techniques for disseminating embryos and sperm: for example, shipping frozen sperm on dry ice, exchanging unfrozen epididymides from which sperm can be extracted, and transporting frozen/thawed embryos in isotonic media. This article discusses some of the current practices used by laboratories to transport mouse strains around the world without having to exchange live mice. PMID:25723918

  10. Genetically modified mouse models addressing gonadotropin function.

    PubMed

    Ratner, Laura D; Rulli, Susana B; Huhtaniemi, Ilpo T

    2014-03-01

    The development of genetically modified animals has been useful to understand the mechanisms involved in the regulation of the gonadotropin function. It is well known that alterations in the secretion of a single hormone is capable of producing profound reproductive abnormalities. Human chorionic gonadotropin (hCG) is a glycoprotein hormone normally secreted by the human placenta, and structurally and functionally it is related to pituitary LH. LH and hCG bind to the same LH/hCG receptor, and hCG is often used as an analog of LH to boost gonadotropin action. There are many physiological and pathological conditions where LH/hCG levels and actions are elevated. In order to understand how elevated LH/hCG levels may impact on the hypothalamic-pituitary-gonadal axis we have developed a transgenic mouse model with chronic hCG hypersecretion. Female mice develop many gonadal and extragonadal phenotypes including obesity, infertility, hyperprolactinemia, and pituitary and mammary gland tumors. This article summarizes recent findings on the mechanisms involved in pituitary gland tumorigenesis and hyperprolactinemia in the female mice hypersecreting hCG, in particular the relationship of progesterone with the hyperprolactinemic condition of the model. In addition, we describe the role of hyperprolactinemia as the main cause of infertility and the phenotypic abnormalities in these mice, and the use of dopamine agonists bromocriptine and cabergoline to normalize these conditions.

  11. Mouse models of intestinal inflammation and cancer.

    PubMed

    Westbrook, Aya M; Szakmary, Akos; Schiestl, Robert H

    2016-09-01

    Chronic inflammation is strongly associated with approximately one-fifth of all human cancers. Arising from combinations of factors such as environmental exposures, diet, inherited gene polymorphisms, infections, or from dysfunctions of the immune response, chronic inflammation begins as an attempt of the body to remove injurious stimuli; however, over time, this results in continuous tissue destruction and promotion and maintenance of carcinogenesis. Here, we focus on intestinal inflammation and its associated cancers, a group of diseases on the rise and affecting millions of people worldwide. Intestinal inflammation can be widely grouped into inflammatory bowel diseases (ulcerative colitis and Crohn's disease) and celiac disease. Long-standing intestinal inflammation is associated with colorectal cancer and small-bowel adenocarcinoma, as well as extraintestinal manifestations, including lymphomas and autoimmune diseases. This article highlights potential mechanisms of pathogenesis in inflammatory bowel diseases and celiac disease, as well as those involved in the progression to associated cancers, most of which have been identified from studies utilizing mouse models of intestinal inflammation. Mouse models of intestinal inflammation can be widely grouped into chemically induced models; genetic models, which make up the bulk of the studied models; adoptive transfer models; and spontaneous models. Studies in these models have lead to the understanding that persistent antigen exposure in the intestinal lumen, in combination with loss of epithelial barrier function, and dysfunction and dysregulation of the innate and adaptive immune responses lead to chronic intestinal inflammation. Transcriptional changes in this environment leading to cell survival, hyperplasia, promotion of angiogenesis, persistent DNA damage, or insufficient repair of DNA damage due to an excess of proinflammatory mediators are then thought to lead to sustained malignant transformation. With

  12. Involve physicians in marketing.

    PubMed

    Randolph, G T; Baker, K M; Laubach, C A

    1984-01-01

    Many everyday problems in medical group practice can be attacked by a marketing approach. To be successful, however, this kind of approach must have the full support of those involved, especially the physicians, since they are the principal providers of healthcare services. When marketing is presented in a broad context, including elements such as patient mix, population distribution, and research, physicians are more likely to be interested and supportive. The members of Geisinger Medical Center's Department of Cardiovascular Medicine addressed their patient appointment backlog problem with a marketing approach. Their method is chronicled here and serves as a fine example of how physician involvement in marketing can lead to a positive outcome.

  13. Aging Research Using Mouse Models

    PubMed Central

    Ackert-Bicknell, Cheryl L.; Anderson, Laura; Sheehan, Susan; Hill, Warren G.; Chang, Bo; Churchill, Gary A.; Chesler, Elissa J.; Korstanje, Ron; Peters, Luanne L.

    2015-01-01

    Despite the dramatic increase in human lifespan over the past century, there remains pronounced variability in “health-span”, or the period of time in which one is generally healthy and free of disease. Much of the variability in health-span and lifespan is thought to be genetic in origin. Understanding the genetic mechanisms of aging and identifying ways to boost longevity is a primary goal in aging research. Here, we describe a pipeline of phenotypic assays for assessing mouse models of aging. This pipeline includes behavior/cognition testing, body composition analysis, and tests of kidney function, hematopoiesis, immune function and physical parameters. We also describe study design methods for assessing lifespan and health-span, and other important considerations when conducting aging research in the laboratory mouse. The tools and assays provided can assist researchers with understanding the correlative relationships between age-associated phenotypes and, ultimately, the role of specific genes in the aging process. PMID:26069080

  14. Genealogies of mouse inbred strains.

    PubMed

    Beck, J A; Lloyd, S; Hafezparast, M; Lennon-Pierce, M; Eppig, J T; Festing, M F; Fisher, E M

    2000-01-01

    The mouse is a prime organism of choice for modelling human disease. Over 450 inbred strains of mice have been described, providing a wealth of different genotypes and phenotypes for genetic and other studies. As new strains are generated and others become extinct, it is useful to review periodically what strains are available and how they are related to each other, particularly in the light of available DNA polymorphism data from microsatellite and other markers. We describe the origins and relationships of inbred mouse strains, 90 years after the generation of the first inbred strain. Given the large collection of inbred strains available, and that published information on these strains is incomplete, we propose that all genealogical and genetic data on inbred strains be submitted to a common electronic database to ensure this valuable information resource is preserved and used efficiently.

  15. Abundant alkali-sensitive sites in DNA of human and mouse sperm

    SciTech Connect

    Singh, N.P.; Danner, D.B.; McCoy, M.T.; Collins, G.D.; Schneider, E.L. ); Tice, R.R. )

    1989-10-01

    The DNA of human and mouse sperm cells was analyzed by single-cell microgel electrophoresis, by agarose gel electrophoresis, and by alkaline elution-three techniques that can detect single-strand DNA breaks and/or labile sites. Under these conditions a surprisingly large number of single-strand DNA breaks, approximately 10{sup 6} to 10{sup 7} per genome, were detected in human and mouse sperm but not in human lymphocytes or in mouse bone marrow cells. These breaks were also present in chicken erythrocyte DNA, which is also highly condensed. These breaks were not observed under neutral pH conditions nor under denaturing conditions not involving alkali, suggesting that these sites are alkali-sensitive and do not represent preexisting single-strand breaks. The high frequency of such sites in sperm from healthy mouse and human donors suggest that they represent a functional characteristic of condensed chromatin rather than DNA damage.

  16. Jute batching oil: a tumor promoter on mouse skin

    SciTech Connect

    Mehrotra, N.K.; Kumar, S.; Agarwal, R.; Antony, M.

    1987-02-01

    A mineral oil essentially used in the jute industry for the batching of jute fibers, and earlier reported to be nontumorigenic on mouse skin, has been found to be a tumor promoter following a two-stage mouse-skin bioassay protocol. The types of tumors developed after initiation with a single dose of urethane or 3-methylcholanthrene (subcutaneously), followed by repeated skin painting with jute batching oil (JBO) included benign papillomas, keratoacanthomas, and fibrosarcomas. Chemical analysis of this oil indicated the total aromatic content was 11.71% and the amount of fluoranthene, pyrene, chrysene, and triphenylene was in the range of 192.54 to 227.79 mg/kg in the test sample. The underlying biochemical mechanism for the tumor-promoting effect of JBO seemed to operate through a different pathway rather than involving the induction of cytochrome-dependent monoxygenase and N-demethylase activities in the tissue.

  17. Jute batching oil: a tumor promoter on mouse skin.

    PubMed

    Mehrotra, N K; Kumar, S; Agarwal, R; Antony, M

    1987-02-01

    A mineral oil essentially used in the jute industry for the "batching" of jute fibers, and earlier reported to be nontumorigenic on mouse skin, has been found to be a tumor promoter following a two-stage mouse-skin bioassay protocol. The types of tumors developed after initiation with a single dose of urethane or 3-methylcholanthrene (subcutaneously), followed by repeated skin painting with jute batching oil (JBO) included benign papillomas, keratoacanthomas, and fibrosarcomas. Chemical analysis of this oil indicated the total aromatic content was 11.71% and the amount of fluoranthene, pyrene, chrysene, and triphenylene was in the range of 192.54 to 227.79 mg/kg in the test sample. The underlying biochemical mechanisms for the tumor-promoting effect of JBO seemed to operate through a different pathway rather than involving the induction of cytochrome-dependent monoxygenase and N-demethylase activities in the tissue.

  18. Metformin prevents methylglyoxal-induced apoptosis of mouse Schwann cells

    SciTech Connect

    Ota, Kimiko; Nakamura, Jiro; Li, Weiguo; Kozakae, Mika; Watarai, Atsuko; Nakamura, Nobuhisa; Yasuda, Yutaka; Nakashima, Eirtaro; Naruse, Keiko; Watabe, Kazuhiko; Kato, Koichi; Oiso, Yutaka; Hamada, Yoji . E-mail: yhama@med.nagoya-u.ac.jp

    2007-05-25

    Methylglyoxal (MG) is involved in the pathogenesis of diabetic complications via the formation of advanced glycation end products (AGEs) and reactive oxygen species (ROS). To clarify whether the antidiabetic drug metformin prevents Schwann cell damage induced by MG, we cultured mouse Schwann cells in the presence of MG and metformin. Cell apoptosis was evaluated using Hoechst 33342 nuclear staining, caspase-3 activity, and c-Jun-N-terminal kinase (JNK) phosphorylation. Intracellular ROS formation was determined by flow cytometry, and AMP-activated kinase (AMPK) phosphorylation was also examined. MG treatment resulted in blunted cell proliferation, an increase in the number of apoptotic cells, and the activation of caspase-3 and JNK along with enhanced intracellular ROS formation. All of these changes were significantly inhibited by metformin. No significant activation of AMPK by MG or metformin was observed. Taken together, metformin likely prevents MG-induced apoptotic signals in mouse Schwann cells by inhibiting the formation of AGEs and ROS.

  19. Retinofugal Projections in the Mouse

    PubMed Central

    Morin, Lawrence P.; Studholme, Keith M.

    2014-01-01

    The laboratory mouse is increasingly a subject for visual system investigation, but there has been no comprehensive evaluation of this species’ visual projections. Here, projections were visualized and mapped following intraocular injection of cholera toxin B subunit. Tissue was processed using standard procedures applied to 30 Am free floating sections with diaminobenzidine as the chromogen. The mouse retina projects to approximately 46 brain regions, including 14 not previously described in this species. These include two amygdaloid nuclei, the horizontal limb of the diagonal band, the paraventricular hypothalamic nucleus, several visual thalamic nuclei, the paranigral nucleus, several pretectal nuclei, and the dorsal cortex of the inferior colliculus. Dense retinal patches were also observed in a narrow portion of the ipsilateral intermediate layer of the superior colliculus. The superior fasciculus of the accessory optic tract, which innervates the medial terminal nucleus, was also determined to be a terminal zone throughout its length. The results are compared with previous descriptions of projections from mouse intrinsically photoreceptive retinal ganglion cells, and with data from the hamster, Nile grass rat and laboratory rat. The retinal projection patterns are similar in all four species, although there are many differences with respect to the details. The specific visual functions of most retinorecipient areas are unknown, but there is substantial convergence of retinal projections onto regions concerned with olfaction and audition. PMID:24889098

  20. Mouse Models of Gastric Cancer

    PubMed Central

    Hayakawa, Yoku; Fox, James G.; Gonda, Tamas; Worthley, Daniel L.; Muthupalani, Sureshkumar; Wang, Timothy C.

    2013-01-01

    Animal models have greatly enriched our understanding of the molecular mechanisms of numerous types of cancers. Gastric cancer is one of the most common cancers worldwide, with a poor prognosis and high incidence of drug-resistance. However, most inbred strains of mice have proven resistant to gastric carcinogenesis. To establish useful models which mimic human gastric cancer phenotypes, investigators have utilized animals infected with Helicobacter species and treated with carcinogens. In addition, by exploiting genetic engineering, a variety of transgenic and knockout mouse models of gastric cancer have emerged, such as INS-GAS mice and TFF1 knockout mice. Investigators have used the combination of carcinogens and gene alteration to accelerate gastric cancer development, but rarely do mouse models show an aggressive and metastatic gastric cancer phenotype that could be relevant to preclinical studies, which may require more specific targeting of gastric progenitor cells. Here, we review current gastric carcinogenesis mouse models and provide our future perspectives on this field. PMID:24216700

  1. Strengthening Parent Involvement.

    ERIC Educational Resources Information Center

    Williams, David L., Jr.; Chavkin, Nancy Feyl

    1986-01-01

    Recent studies have verified Secretary of Education William Bennett's observation on the importance of home and family life. The most successful students are those whose parents become actively engaged in the educational process at home and at school. To capitalize on potential parent involvement, principals need to understand the kinds of…

  2. Parent Involvement. Research Brief

    ERIC Educational Resources Information Center

    Walker, Karen

    2007-01-01

    What are some ways in which to get parents meaningfully involved in their child's high school? According to the research, the most successful programs are those that provide a variety of ways in which parents can be actively engaged in their child's academic life. Joyce Epstein, Director of the National Network of Partnership Schools, out of Johns…

  3. Apolipoprotein gene involved in lipid metabolism

    DOEpatents

    Rubin, Edward; Pennacchio, Len A.

    2007-07-03

    Methods and materials for studying the effects of a newly identified human gene, APOAV, and the corresponding mouse gene apoAV. The sequences of the genes are given, and transgenic animals which either contain the gene or have the endogenous gene knocked out are described. In addition, single nucleotide polymorphisms (SNPs) in the gene are described and characterized. It is demonstrated that certain SNPs are associated with diseases involving lipids and triglycerides and other metabolic diseases. These SNPs may be used alone or with SNPs from other genes to study individual risk factors. Methods for intervention in lipid diseases, including the screening of drugs to treat lipid-related or diabetic diseases are also disclosed.

  4. Genomic structure and chromosomal assignment of the mouse Ku70 gene

    SciTech Connect

    Takiguchi, Yuichi |; Kurimasa, Akihiro; Chen, Fanqing

    1996-07-01

    DNA-dependent protein kinase (DNA-PK) consists of three polypeptide subunits: Ku70, Ku80, and the DNA-PK catalytic subunit (DNA-PKcs). Mammalian mutants deficient in either Ku80 or DNA-PKcs function have been shown to be lacking in DNA double-strand break repair and V(D)J recombination, respectively. The precise role of the Ku70 gene in this process has not yet been determined, in part because no cell lines, animals, or human diseases involved with deficiencies in this gene have yet been identified. Both the human and the mouse Ku70 cDNAs have been cloned, and the human gene has been mapped to chromosome 22q13. The original mouse cDNA clones, however, lacked a complete 5{prime}-region, and none of the mammalian Ku70 genomic sequences have been characterized. This report contains an analysis of the 5{prime}-region of the mouse cDNA sequence, a characterization of the mouse Ku70 genomic structure, and fluorescence in situ hybridization data that map the mouse gene to chromosome 15. The deduced amino acid sequence of the mouse gene consists of 608 amino acids compared to 609 for the human gene. The genomic sequence is 24 kb and consists of 13 exons, including an untranslated first exon. Sequences form the upstream region of exon 1 revealed four consensus GC box sequences and a strong transcription initiation site at a reasonable location. The assignment of the mouse Ku70 gene to chromosome 15 is consistent with the syntenic relationship of this gene in human (chromosome 22q13) and mouse and adds to the comparative mapping data for the genes involved in the SCID phenotype. 39 refs., 3 figs.

  5. Expression pattern of Sox2 during mouse tooth development.

    PubMed

    Zhang, Li; Yuan, Guohua; Liu, Huan; Lin, Heng; Wan, Chunyan; Chen, Zhi

    2012-01-01

    The transcription factor Sox2 plays important roles in maintaining the pluripotency of embryonic stem cells and adult progenitors. However, whether Sox2 is involved in odontogenesis has not been reported. In this study, we examined the expression pattern of Sox2 during mouse incisor and molar development using real-time PCR, in situ hybridization and immunohistochemistry. Sox2 mRNA was expressed in the dental epithelium and mesenchyme while Sox2 protein was mainly detected in the epithelium from embryonic day (E) 11.5 to postnatal (PN) day 20. In the case of incisor, Sox2 mRNA and protein were expressed in most of dental epithelial cells from E11.5 to E14.5, and they were both highly expressed in the labial cervical loop area from E16.5 to PN20. During molar development, we observed an asymmetrical distribution of Sox2 protein in the epithelium from E13.5 to E16.5, with stronger signals in the lingual side. From E18.5 to PN2, Sox2 was expressed within the cervical loop area, and the stellate intermediate layer. From PN6 to PN14, Sox2 expression was confined mainly to the apical end of hertwig's epithelium root sheath (HERS) cells. Sox2 was also detected within the perivascular region of the dental pulp at PN14 and PN20. Our results suggested that: (1) Sox2 was involved in mouse odontogenesis, and (2) it might participate in maintaining the pluripotency of the epithelial stem cells of labial cervical loop in mouse incisor development and the epithelium progenitors during molar development, (3) Sox2 might be regulated at post-transcription level during mouse odontogenesis.

  6. Mouse embryonic stem cells with a multi-integrase mouse artificial chromosome for transchromosomic mouse generation.

    PubMed

    Yoshimura, Yuki; Nakamura, Kazuomi; Endo, Takeshi; Kajitani, Naoyo; Kazuki, Kanako; Kazuki, Yasuhiro; Kugoh, Hiroyuki; Oshimura, Mitsuo; Ohbayashi, Tetsuya

    2015-08-01

    The mouse artificial chromosome (MAC) has several advantages as a gene delivery vector, including stable episomal maintenance of the exogenous genetic material and the ability to carry large and/or multiple gene inserts including their regulatory elements. Previously, a MAC containing multi-integration site (MI-MAC) was generated to facilitate transfer of multiple genes into desired cells. To generate transchromosomic (Tc) mice containing a MI-MAC with genes of interest, the desired genes were inserted into MI-MAC in CHO cells, and then the MI-MAC was transferred to mouse embryonic stem (mES) cells via microcell-mediated chromosome transfer (MMCT). However, the efficiency of MMCT from CHO to mES cells is very low (<10(-6)). In this study, we constructed mES cell lines containing a MI-MAC vector to directly insert a gene of interest into the MI-MAC in mES cells via a simple transfection method for Tc mouse generation. The recombination rate of the GFP gene at each attachment site (FRT, PhiC31attP, R4attP, TP901-1attP and Bxb1attP) on MI-MAC was greater than 50% in MI-MAC mES cells. Chimeric mice with high coat colour chimerism were generated from the MI-MAC mES cell lines and germline transmission from the chimera was observed. As an example for the generation of Tc mice with a desired gene by the MI-MAC mES approach, a Tc mouse strain ubiquitously expressing Emerald luciferase was efficiently established. Thus, the findings suggest that this new Tc strategy employing mES cells and a MI-MAC vector is efficient and useful for animal transgenesis.

  7. Mouse embryonic stem cells with a multi-integrase mouse artificial chromosome for transchromosomic mouse generation.

    PubMed

    Yoshimura, Yuki; Nakamura, Kazuomi; Endo, Takeshi; Kajitani, Naoyo; Kazuki, Kanako; Kazuki, Yasuhiro; Kugoh, Hiroyuki; Oshimura, Mitsuo; Ohbayashi, Tetsuya

    2015-08-01

    The mouse artificial chromosome (MAC) has several advantages as a gene delivery vector, including stable episomal maintenance of the exogenous genetic material and the ability to carry large and/or multiple gene inserts including their regulatory elements. Previously, a MAC containing multi-integration site (MI-MAC) was generated to facilitate transfer of multiple genes into desired cells. To generate transchromosomic (Tc) mice containing a MI-MAC with genes of interest, the desired genes were inserted into MI-MAC in CHO cells, and then the MI-MAC was transferred to mouse embryonic stem (mES) cells via microcell-mediated chromosome transfer (MMCT). However, the efficiency of MMCT from CHO to mES cells is very low (<10(-6)). In this study, we constructed mES cell lines containing a MI-MAC vector to directly insert a gene of interest into the MI-MAC in mES cells via a simple transfection method for Tc mouse generation. The recombination rate of the GFP gene at each attachment site (FRT, PhiC31attP, R4attP, TP901-1attP and Bxb1attP) on MI-MAC was greater than 50% in MI-MAC mES cells. Chimeric mice with high coat colour chimerism were generated from the MI-MAC mES cell lines and germline transmission from the chimera was observed. As an example for the generation of Tc mice with a desired gene by the MI-MAC mES approach, a Tc mouse strain ubiquitously expressing Emerald luciferase was efficiently established. Thus, the findings suggest that this new Tc strategy employing mES cells and a MI-MAC vector is efficient and useful for animal transgenesis. PMID:26055730

  8. Qualitative analysis of mouse specific-locus mutations: information on genetic organization, gene expression, and the chromosomal nature of induced lesions

    SciTech Connect

    Russell, L.B.

    1982-01-01

    Analysis of mouse specific-locus (SL) mutations at three loci has identified over 33 distinct complementation groups - most of which are probably overlapping deficiencies - and 13 to 14 new functional units. The complementation maps that have been generated for the d-se and c regions include numerous vital functions; however, some of the genes in these regions are non-vital. At such loci, hypomorphic mutants must represent intragenic alterations, and some viable nulls could conceivably be intragenic lesions also. Analysis of SL mutations has provided information on genetic expression. Homozygous deficiencies can be completely viable or can kill at any one of a range of developmental stages. Heterozygonus deficiencies of up to 6 cM or more in genetic length have been recovered and propagated. The time of death of homozygous and the degree of inviability of heterozygous deficiencies are related more to specific content of the missing segment than to its length. Combinations of deficiencies with x-autosome translocations that inactivate the homologous region in a mosaic fashion have shown that organismic lethals are not necessarily cell lethal. The spectrum of mutations induced depends on the nature of the mutagen and the type of germ cell exposed. Radiation of spermatogonia produces intragenic as well as null mutations. Spontaneous mutations have an admixture of types not present in populations of mutations induced in germ cells, and this raises doubts concerning the accuracy of doubling-dose calculations in genetic risk estimation. The analysis of SL mutations has yielded genetic tools for the construction of detailed gene-dosage series, cis-trans comparisons, the mapping of known genes and identification of new genes, genetic rescue of various types, and the identification and isolation of DNA sequences. (ERB)

  9. ATM localization and gene expression in the adult mouse eye

    PubMed Central

    Leemput, Julia; Masson, Christel; Bigot, Karine; Errachid, Abdelmounaim; Dansault, Anouk; Provost, Alexandra; Gadin, Stéphanie; Aoufouchi, Said; Menasche, Maurice

    2009-01-01

    Purpose High levels of metabolism and oxygen consumption in most adult murine ocular compartments, combined with exposure to light and ultraviolet (UV) radiation, are major sources of oxidative stress, causing DNA damage in ocular cells. Of all mammalian body cells, photoreceptor cells consume the largest amount of oxygen and generate the highest levels of oxidative damage. The accumulation of such damage throughout life is a major factor of aging tissues. Several multiprotein complexes have recently been identified as the major sensors and mediators involved in the maintenance of DNA integrity. The activity of these complexes initially seemed to be restricted to dividing cells, given their ultimate role in major cell cycle checkpoints. However, it was later established that they are also active in post-mitotic cells. Recent findings demonstrate that the DNA damage response (DDR) is essential for the development, maintenance, and normal functioning of the adult central nervous system. One major molecular factor in the DDR is the protein, ataxia telangiectasia mutated (ATM). It is required for the rapid induction of cellular responses to DNA double-strand breaks. These cytotoxic DNA lesions may be caused by oxidative damage. To understand how ATM prevents oxidative stress and participates in the maintenance of genomic integrity and cell viability of the adult retina, we determined the ATM expression patterns and studied its localization in the adult mouse eye. Methods Atm gene expression was analyzed by RT–PCR experiments and its localization by in situ hybridization on adult mouse ocular and cerebellar tissue sections. ATM protein expression was determined by western blot analysis of proteins homogenates extracted from several mouse tissues and its localization by immunohistochemistry experiments performed on adult mouse ocular and cerebellar tissue sections. In addition, subcellular localization was realized by confocal microscopy imaging of ocular tissue

  10. Mouse mammary tumor biology: a short history.

    PubMed

    Cardiff, Robert D; Kenney, Nicholas

    2007-01-01

    For over a century, mouse mammary tumor biology and the associated Mouse mammary tumor virus (MMTV) have served as the foundation for experimental cancer research, in general, and, in particular, experimental breast cancer research. Spontaneous mouse mammary tumors were the basis for studies of the natural history of neoplasia, oncogenic viruses, host responses, endocrinology, and neoplastic progression. However, lacking formal proof of a human mammary tumor virus, the preeminence of the mouse model faded in the 1980s. Since the late 1980s, genetically engineered mice (GEM) have proven extremely useful for studying breast cancer and have become the animal model for human breast cancer. Hundreds of mouse models of human breast cancer have been developed since the first demonstration, in 1984, that the mouse mammary gland could be molecularly targeted and used to test the oncogenicity of candidate human genes. Now, very few scientists can avoid using a mouse model to test the biology of their favorite gene. The GEM have attracted a new generation of molecular and cellular biologists eager to apply their skills to these surrogates of the human disease. Newcomers often enter the field without an appreciation of the origins of mouse mammary tumor biology and the basis for many of the prevailing concepts. Our purpose in writing this short history of mouse mammary tumor biology is to provide a historical perspective for the benefit of the newcomers. If Einstein was correct in that "we stand on the shoulders of giants," the neophytes should meet their giants.

  11. The human and mouse homologs of the yeat RAD52 gene: cDNA cloning, sequence analysis, assignment to human chromosome 12p12.2-p13, and mRNA expression in mouse tissues

    SciTech Connect

    Shen, Z.; Chen, D.J.; Denison, K.

    1995-01-01

    The yeast Saccharomyces cerevisiae RAD52 gene is involved in DNA double-strand break repair and mitotic/meiotic recombination. The N-terminal amino acid sequence of yeast S. cerevisiae, Schizosaccharomyces pombe, and Kluyveromyces lactis and chicken is highly conserved. Using the technology of mixed oligonucleotide primed amplification of cDNA (MOPAC), two mouse RAD52 homologous cDNA fragments were amplified and sequenced. Subsequently, we have cloned the cDNA of the human and mouse homologs of yeast RAD52 gene by screening cDNA libraries using the identified mouse cDNA fragments. Sequence analysis of cDNA derived amino acid revealed a highly conserved N-terminus among human, mouse, chicken, and yeast RAD52 genes. The human RAD52 gene was assigned to chromosome 12p12.2-p13 by fluorescence in situ hybridization, R-banding, and DNA analysis of somatic cell hybrids. Unlike chicken RAD52 and mouse RAD51, no significant difference in mouse RAD52 mRNA level was found among mouse heart, brain, spleen, lung, liver, skeletal muscle, kidney, and testis. In addition to an {approximately}1.9-kb RAD52 mRNA band that is present in all of the tested tissues, an extra mRNA species of {approximately}0.85 kb was detectable in mouse testis. 40 refs., 7 figs., 1 tab.

  12. Therapeutic cloning in the mouse.

    PubMed

    Mombaerts, Peter

    2003-09-30

    Nuclear transfer technology can be applied to produce autologous differentiated cells for therapeutic purposes, a concept termed therapeutic cloning. Countless articles have been published on the ethics and politics of human therapeutic cloning, reflecting the high expectations from this new opportunity for rejuvenation of the aging or diseased body. Yet the research literature on therapeutic cloning, strictly speaking, is comprised of only four articles, all in the mouse. The efficiency of derivation of embryonic stem cell lines via nuclear transfer is remarkably consistent among these reports. However, the efficiency is so low that, in its present form, the concept is unlikely to become widespread in clinical practice.

  13. Endocannabinoid involvement in endometriosis

    PubMed Central

    Dmitrieva, Natalia; Nagabukuro, Hiroshi; Resuehr, David; Zhang, Guohua; McAllister, Stacy L.; McGinty, Kristina A.; Mackie, Ken; Berkley, Karen J.

    2010-01-01

    Endometriosis is a disease common in women that is defined by abnormal extrauteral growths of uterine endometrial tissue and associated with severe pain. Partly because how the abnormal growths become associated with pain is poorly understood, the pain is difficult to alleviate without resorting to hormones or surgery, which often produce intolerable side effects or fail to help. Recent studies in a rat model and women showed that sensory and sympathetic nerve fibers sprout branches to innervate the abnormal growths. This situation, together with knowledge that the endocannabinoid system is involved in uterine function and dysfunction and that exogenous cannabinoids were once used to alleviate endometriosis-associated pain, suggests that the endocannabinoid system is involved in both endometriosis and its associated pain. Here, using a rat model, we found that CB1 cannabinoid receptors are expressed on both the somata and fibers of both the sensory and sympathetic neurons that innervate endometriosis’s abnormal growths. We further found that CB1 receptor agonists decrease, whereas CB1 receptor antagonists increase, endometriosis-associated hyperalgesia. Together these findings suggest that the endocannabinoid system contributes to mechanisms underlying both the peripheral innervation of the abnormal growths and the pain associated with endometriosis, thereby providing a novel approach for the development of badly-needed new treatments. PMID:20833475

  14. Myostatin - From the Mighty Mouse to cardiovascular disease and cachexia.

    PubMed

    Dschietzig, Thomas Bernd

    2014-06-10

    In 1997, McPherron et al. created the so-called Mighty Mouse: owing to the knock-out of a new member of the TGF-β superfamily of peptides, this mouse line was extremely hypermuscular and also characterized by very low body fat. The new peptide, a powerful negative muscle regulator, was named myostatin. Apart from regulating skeletal muscle growth, myostatin has recently been reported to be significantly involved in different cardio-vascular and metabolic pathologies. This review is focused on these non-muscular myostatin actions. First, myostatin is intricately involved in regulating metabolism: it causes insulin resistance, and the advantageous metabolic profile achieved by myostatin inhibition is mainly attributable to its effects on skeletal muscle. Myostatin is further expressed in myocardium where it exerts anti-hypertrophic, but pro-fibrotic effects. Circulating and local myostatin is elevated in chronic heart failure and poses a major player in cardiac cachexia. Eventually, the current body of evidence regarding myostatin's significant involvement in different entities of the cachexia syndrome is summarized. Activin type-2 receptor antagonism and/or inhibitory myostatin antibodies have emerged as a promising therapeutic approach to treat the cachexia syndrome although the general applicability of this therapeutic approach to the human clinical situation has still to be demonstrated.

  15. Overexpression of mouse TTF-2 gene causes cleft palate

    PubMed Central

    Meng, Tian; Shi, Jia-Yu; Wu, Min; Wang, Yan; Li, Ling; Liu, Yan; Zheng, Qian; Huang, Lei; Shi, Bing

    2012-01-01

    In humans, mutations of the gene encoding for thyroid transcription factor-2 (TTF-2 or FOXE1) result in Bamforth syndrome. Bamforth syndrome is characterized by agenesis, cleft palate, spiky hair and choanal atresia. TTF-2 null mice (TTF-2−/−) also exhibit cleft palate, suggesting its involvement in the palatogenesis. However, the molecular pathology and genetic regulation by TTF2 remain largely unknown. In the present study, the recombinant expression vector pBROAD3-TTF-2 containing the promoter of the mouse ROSA26 gene was created to form the structural gene of mouse TTF-2 and was microinjected into the male pronuclei of fertilized ova. Sequence analysis confirmed that the TTF-2 transgenic mouse model was established successfully. The transgenic mice displayed a phenotype of cleft palate. In addition, we found that TTF-2 was highly expressed in the medial edge epithelium (MEE) from the embryonic day 12.5 (E12.5) to E14.5 in TTF-2 transgenic mice. These observations suggest that overexpression of TTF-2 during palatogenesis may contribute to formation of cleft palate. PMID:22304410

  16. Transgenic Rescue of the LARGEmyd Mouse: A LARGE Therapeutic Window?

    PubMed Central

    Hildyard, J. C. W.; Lacey, E.; Booler, H.; Hopkinson, M.; Wells, D. J.; Brown, S. C.

    2016-01-01

    LARGE is a glycosyltransferase involved in glycosylation of α-dystroglycan (α-DG). Absence of this protein in the LARGEmyd mouse results in α-DG hypoglycosylation, and is associated with central nervous system abnormalities and progressive muscular dystrophy. Up-regulation of LARGE has previously been proposed as a therapy for the secondary dystroglycanopathies: overexpression in cells compensates for defects in multiple dystroglycanopathy genes. Counterintuitively, LARGE overexpression in an FKRP-deficient mouse exacerbates pathology, suggesting that modulation of α-DG glycosylation requires further investigation. Here we demonstrate that transgenic expression of human LARGE (LARGE-LV5) in the LARGEmyd mouse restores α-DG glycosylation (with marked hyperglycosylation in muscle) and that this corrects both the muscle pathology and brain architecture. By quantitative analyses of LARGE transcripts we also here show that levels of transgenic and endogenous LARGE in the brains of transgenic animals are comparable, but that the transgene is markedly overexpressed in heart and particularly skeletal muscle (20–100 fold over endogenous). Our data suggest LARGE overexpression may only be deleterious under a forced regenerative context, such as that resulting from a reduction in FKRP: in the absence of such a defect we show that systemic expression of LARGE can indeed act therapeutically, and that even dramatic LARGE overexpression is well-tolerated in heart and skeletal muscle. Moreover, correction of LARGEmyd brain pathology with only moderate, near-physiological LARGE expression suggests a generous therapeutic window. PMID:27467128

  17. Transgenic Rescue of the LARGEmyd Mouse: A LARGE Therapeutic Window?

    PubMed

    Hildyard, J C W; Lacey, E; Booler, H; Hopkinson, M; Wells, D J; Brown, S C

    2016-01-01

    LARGE is a glycosyltransferase involved in glycosylation of α-dystroglycan (α-DG). Absence of this protein in the LARGEmyd mouse results in α-DG hypoglycosylation, and is associated with central nervous system abnormalities and progressive muscular dystrophy. Up-regulation of LARGE has previously been proposed as a therapy for the secondary dystroglycanopathies: overexpression in cells compensates for defects in multiple dystroglycanopathy genes. Counterintuitively, LARGE overexpression in an FKRP-deficient mouse exacerbates pathology, suggesting that modulation of α-DG glycosylation requires further investigation. Here we demonstrate that transgenic expression of human LARGE (LARGE-LV5) in the LARGEmyd mouse restores α-DG glycosylation (with marked hyperglycosylation in muscle) and that this corrects both the muscle pathology and brain architecture. By quantitative analyses of LARGE transcripts we also here show that levels of transgenic and endogenous LARGE in the brains of transgenic animals are comparable, but that the transgene is markedly overexpressed in heart and particularly skeletal muscle (20-100 fold over endogenous). Our data suggest LARGE overexpression may only be deleterious under a forced regenerative context, such as that resulting from a reduction in FKRP: in the absence of such a defect we show that systemic expression of LARGE can indeed act therapeutically, and that even dramatic LARGE overexpression is well-tolerated in heart and skeletal muscle. Moreover, correction of LARGEmyd brain pathology with only moderate, near-physiological LARGE expression suggests a generous therapeutic window.

  18. Mouse Models Recapitulating Human Adrenocortical Tumors: What Is Lacking?

    PubMed

    Leccia, Felicia; Batisse-Lignier, Marie; Sahut-Barnola, Isabelle; Val, Pierre; Lefrançois-Martinez, A-Marie; Martinez, Antoine

    2016-01-01

    Adrenal cortex tumors are divided into benign forms, such as primary hyperplasias and adrenocortical adenomas (ACAs), and malignant forms or adrenocortical carcinomas (ACCs). Primary hyperplasias are rare causes of adrenocorticotropin hormone-independent hypercortisolism. ACAs are the most common type of adrenal gland tumors and they are rarely "functional," i.e., producing steroids. When functional, adenomas result in endocrine disorders, such as Cushing's syndrome (hypercortisolism) or Conn's syndrome (hyperaldosteronism). By contrast, ACCs are extremely rare but highly aggressive tumors that may also lead to hypersecreting syndromes. Genetic analyses of patients with sporadic or familial forms of adrenocortical tumors (ACTs) led to the identification of potentially causative genes, most of them being involved in protein kinase A (PKA), Wnt/β-catenin, and P53 signaling pathways. Development of mouse models is a crucial step to firmly establish the functional significance of candidate genes, to dissect mechanisms leading to tumors and endocrine disorders, and in fine to provide in vivo tools for therapeutic screens. In this article, we will provide an overview on the existing mouse models (xenografted and genetically engineered) of ACTs by focusing on the role of PKA and Wnt/β-catenin pathways in this context. We will discuss the advantages and limitations of models that have been developed heretofore and we will point out necessary improvements in the development of next generation mouse models of adrenal diseases. PMID:27471492

  19. Host Genetic and Environmental Effects on Mouse Cecum Microbiota

    SciTech Connect

    Campbell, James H; Foster, Carmen M; Vishnivetskaya, Tatiana A; Campbell, Alisha G; Yang, Zamin Koo; Wymore, Ann; Palumbo, Anthony Vito; Podar, Mircea

    2012-01-01

    The mammalian gut harbors complex and variable microbial communities, across both host phylogenetic space and conspecific individuals. A synergy of host genetic and environmental factors shape these communities and account for their variability, but their individual contributions and the selective pressures involved are still not well understood. We employed barcoded pyrosequencing of V1-2 and V4 regions of bacterial small subunit ribosomal RNA genes to characterize the effects of host genetics and environment on cecum assemblages in 10 genetically distinct, inbred mouse strains. Eight of these strains are the foundation of the Collaborative Cross (CC), a panel of mice derived from a genetically diverse set of inbred founder strains, designed specifically for complex trait analysis. Diversity of gut microbiota was characterized by complementing phylogenetic and distance-based, sequence-clustering approaches. Significant correlations were found between the mouse strains and their gut microbiota, reflected by distinct bacterial communities. Cohabitation and litter had a reduced, although detectable effect, and the microbiota response to these factors varied by strain. We identified bacterial phylotypes that appear to be discriminative and strain-specific to each mouse line used. Cohabitation of different strains of mice revealed an interaction of host genetic and environmental factors in shaping gut bacterial consortia, in which bacterial communities became more similar but retained strain specificity. This study provides a baseline analysis of intestinal bacterial communities in the eight CC progenitor strains and will be linked to integrated host genotype, phenotype and microbiota research on the resulting CC panel.

  20. Mouse Models Recapitulating Human Adrenocortical Tumors: What Is Lacking?

    PubMed Central

    Leccia, Felicia; Batisse-Lignier, Marie; Sahut-Barnola, Isabelle; Val, Pierre; Lefrançois-Martinez, A-Marie; Martinez, Antoine

    2016-01-01

    Adrenal cortex tumors are divided into benign forms, such as primary hyperplasias and adrenocortical adenomas (ACAs), and malignant forms or adrenocortical carcinomas (ACCs). Primary hyperplasias are rare causes of adrenocorticotropin hormone-independent hypercortisolism. ACAs are the most common type of adrenal gland tumors and they are rarely “functional,” i.e., producing steroids. When functional, adenomas result in endocrine disorders, such as Cushing’s syndrome (hypercortisolism) or Conn’s syndrome (hyperaldosteronism). By contrast, ACCs are extremely rare but highly aggressive tumors that may also lead to hypersecreting syndromes. Genetic analyses of patients with sporadic or familial forms of adrenocortical tumors (ACTs) led to the identification of potentially causative genes, most of them being involved in protein kinase A (PKA), Wnt/β-catenin, and P53 signaling pathways. Development of mouse models is a crucial step to firmly establish the functional significance of candidate genes, to dissect mechanisms leading to tumors and endocrine disorders, and in fine to provide in vivo tools for therapeutic screens. In this article, we will provide an overview on the existing mouse models (xenografted and genetically engineered) of ACTs by focusing on the role of PKA and Wnt/β-catenin pathways in this context. We will discuss the advantages and limitations of models that have been developed heretofore and we will point out necessary improvements in the development of next generation mouse models of adrenal diseases. PMID:27471492

  1. Expression and Function of Kisspeptin during Mouse Decidualization

    PubMed Central

    Lin, Yan; Zong, Teng; Zhong, Chengxue; Zhang, BaoPing; Ren, Min; Kuang, HaiBin

    2014-01-01

    Background Plasma kisspeptin levels dramatically increased during the first trimester of human pregnancy, which is similar to pregnancy specific glycoprotein-human chorionic gonadotropin. However, its particular role in the implantation and decidualization has not been fully unraveled. Here, the study was conducted to investigate the expression and function of kisspeptin in mouse uterus during early pregnancy and decidualization. Methodology/Principal Findings Quantitative PCR results demonstrated that Kiss1 and GPR54 mRNA levels showed dynamic increase in the mouse uterus during early pregnancy and artificially induced decidualization in vivo. KISS-1 and GPR54 proteins were spatiotemporally expressed in decidualizing stromal cells in intact pregnant females, as well as in pseudopregnant mice undergoing artificially induced decidualization. In the ovariectomized mouse uterus, the expression of Kiss1 mRNA was upregulated after progesterone or/and estradiol treatment. Moreover, in a stromal cell culture model, the expression of Kiss1 and GPR54 mRNA gradually rise with the progression of stromal cell decidualization, whereas the attenuated expression of Kiss1 using small interfering RNA approaches significantly blocked the progression of stromal cell decidualization. Conclusion our results demonstrated that Kiss1/GPR54 system was involved in promoting uterine decidualization during early pregnancy in mice. PMID:24830702

  2. Organization and evolution of D region class I genes in the mouse major histocompatibility complex

    PubMed Central

    1986-01-01

    Chromosome walking has been used to study the organization of the class I genes in the D and Qa regions of the MHC of the BALB/c mouse and in the D region of the AKR mouse. Five and eight class I genes are found in the D and Qa regions of the BALB/c mouse, respectively, while the AKR mouse contains only a single class I D region gene that has been identified by transfection as the Dk gene. Restriction map homologies and crosshybridization experiments suggest that the multiple class I genes in the D region of the BALB/c mouse have been generated by unequal crossing-over involving class I genes from the Qa region. The expanded D region of BALB/c and other H-2d haplotype mouse strains appears to be metastable, since evidence for gene contraction in the Dd region has been found in two mutant strains. Thus the D region and also the Qa region class I genes are in a dynamic state, evolving by gene expansion and contraction. PMID:3701254

  3. Effective protein inhibition in intact mouse oocytes through peptide nanoparticle-mediated antibody transfection

    PubMed Central

    Li, Ruichao; Jin, Zhen; Gao, Leilei; Liu, Peng

    2016-01-01

    Female meiosis is a fundamental area of study in reproductive medicine, and the mouse oocyte model of in vitro maturation (IVM) is most widely used to study female meiosis. To investigate the probable role(s) of an unknown protein in female meiosis, the method traditionally used involves microinjecting a specific antibody into mouse oocytes. Recently, in studies on somatic cells, peptide nanoparticle-mediated antibody transfection has become a popular tool because of its high efficiency, low toxicity, good stability, and strong serum compatibility. However, untill now no researchers have tried using this technique on mouse oocytes because the zona pellucida surrounding the oocyte membrane (vitelline membrane) is usually thought or proved to be a tough barrier to macromolecules such as antibodies and proteins. Therefore, we attempted to introduce an antibody into mouse oocytes using a peptide nanoparticle. Here we show for the first time that with our optimized method, an antibody can be effectively delivered into mouse oocytes and inhibit its target protein with high specificity. We obtained significant results using small GTPase Arl2 as a test subject protein. We propose peptide nanoparticle-mediated antibody transfection to be a superior alternative to antibody microinjection for preliminary functional studies of unknown proteins in mouse oocytes. PMID:27114861

  4. Mouse ENU Mutagenesis to Understand Immunity to Infection: Methods, Selected Examples, and Perspectives.

    PubMed

    Caignard, Grégory; Eva, Megan M; van Bruggen, Rebekah; Eveleigh, Robert; Bourque, Guillaume; Malo, Danielle; Gros, Philippe; Vidal, Silvia M

    2014-01-01

    Infectious diseases are responsible for over 25% of deaths globally, but many more individuals are exposed to deadly pathogens. The outcome of infection results from a set of diverse factors including pathogen virulence factors, the environment, and the genetic make-up of the host. The completion of the human reference genome sequence in 2004 along with technological advances have tremendously accelerated and renovated the tools to study the genetic etiology of infectious diseases in humans and its best characterized mammalian model, the mouse. Advancements in mouse genomic resources have accelerated genome-wide functional approaches, such as gene-driven and phenotype-driven mutagenesis, bringing to the fore the use of mouse models that reproduce accurately many aspects of the pathogenesis of human infectious diseases. Treatment with the mutagen N-ethyl-N-nitrosourea (ENU) has become the most popular phenotype-driven approach. Our team and others have employed mouse ENU mutagenesis to identify host genes that directly impact susceptibility to pathogens of global significance. In this review, we first describe the strategies and tools used in mouse genetics to understand immunity to infection with special emphasis on chemical mutagenesis of the mouse germ-line together with current strategies to efficiently identify functional mutations using next generation sequencing. Then, we highlight illustrative examples of genes, proteins, and cellular signatures that have been revealed by ENU screens and have been shown to be involved in susceptibility or resistance to infectious diseases caused by parasites, bacteria, and viruses.

  5. Sperm Proteome Maturation in the Mouse Epididymis

    PubMed Central

    Skerget, Sheri; Rosenow, Matthew A.; Petritis, Konstantinos; Karr, Timothy L.

    2015-01-01

    In mammals, transit through the epididymis, which involves the acquisition, loss and modification of proteins, is required to confer motility and fertilization competency to sperm. The overall dynamics of maturation is poorly understood, and a systems level understanding of the complex maturation process will provide valuable new information about changes occurring during epididymal transport. We report the proteomes of sperm collected from the caput, corpus and cauda segments of the mouse epididymis, identifying 1536, 1720 and 1234 proteins respectively. This study identified 765 proteins that are present in sperm obtained from all three segments. We identified 1766 proteins that are potentially added (732) or removed (1034) from sperm during epididymal transit. Phenotypic analyses of the caput, corpus and cauda sperm proteomes identified 60 proteins that have known sperm phenotypes when mutated, or absent from sperm. Our analysis indicates that as much as one-third of proteins with known sperm phenotypes are added to sperm during epididymal transit. GO analyses revealed that cauda sperm are enriched for specific functions including sperm-egg recognition and motility, consistent with the observation that sperm acquire motility and fertilization competency during transit through the epididymis. In addition, GO analyses revealed that the immunity protein profile of sperm changes during sperm maturation. Finally, we identified components of the 26S proteasome, the immunoproteasome, and a proteasome activator in mature sperm. PMID:26556802

  6. Plantarflexion Contracture in the mdx Mouse

    PubMed Central

    Garlich, Michael W.; Baltgalvis, Kristen A.; Call, Jarrod A.; Dorsey, Lisa L.; Lowe, Dawn A.

    2012-01-01

    Objective Contractures are a major clinical issue for patients with muscular dystrophies. However, it is unknown whether contractures are present in the widely used mdx mouse model of Duchenne muscular dystrophy. Therefore, the objectives of this study were to develop methods to measure muscle contractures in mice, to determine whether plantarflexion contractures are present in mdx mice, and to analyze the composition of the major muscles involved. Design Hindlimbs of eight wild type and six mdx mice were assessed every 2 wks during the course of a 12-wk study. Assessments included range of motion and in vivo torques about the ankle. At the end of the study, mice were euthanized, and muscles were analyzed for composition. Results The mdx mice had ~10 degrees less dorsiflexion, increased passive torque moving the ankle into dorsiflexion, and an increased passive-to-active torque ratio relative to wild type mice. Gastrocnemius muscle composition alterations included increased wet mass, decreased protein content, and increased collagen. Conclusions The results indicate that mdx mice have plantarflexion contractures similar to those seen in children with Duchenne muscular dystrophy. In future studies, these measures can be used to assess strategies to slow the progression of contractures that occur with muscular dystrophies. PMID:21403594

  7. Combinatorial effects of odorants on mouse behavior

    PubMed Central

    Saraiva, Luis R.; Kondoh, Kunio; Ye, Xiaolan; Yoon, Kyoung-hye; Hernandez, Marcus; Buck, Linda B.

    2016-01-01

    The mechanisms by which odors induce instinctive behaviors are largely unknown. Odor detection in the mouse nose is mediated by >1, 000 different odorant receptors (ORs) and trace amine-associated receptors (TAARs). Odor perceptions are encoded combinatorially by ORs and can be altered by slight changes in the combination of activated receptors. However, the stereotyped nature of instinctive odor responses suggests the involvement of specific receptors and genetically programmed neural circuits relatively immune to extraneous odor stimuli and receptor inputs. Here, we report that, contrary to expectation, innate odor-induced behaviors can be context-dependent. First, different ligands for a given TAAR can vary in behavioral effect. Second, when combined, some attractive and aversive odorants neutralize one another’s behavioral effects. Both a TAAR ligand and a common odorant block aversion to a predator odor, indicating that this ability is not unique to TAARs and can extend to an aversive response of potential importance to survival. In vitro testing of single receptors with binary odorant mixtures indicates that behavioral blocking can occur without receptor antagonism in the nose. Moreover, genetic ablation of a single receptor prevents its cognate ligand from blocking predator odor aversion, indicating that the blocking requires sensory input from the receptor. Together, these findings indicate that innate odor-induced behaviors can depend on context, that signals from a single receptor can block innate odor aversion, and that instinctive behavioral responses to odors can be modulated by interactions in the brain among signals derived from different receptors. PMID:27208093

  8. Mouse models: the ketogenic diet and polyunsaturated fatty acids.

    PubMed

    Borges, Karin

    2008-11-01

    Literature on the anticonvulsant effects of the ketogenic diet (KD) in mouse seizure models is summarized. Recent data show that a KD balanced in vitamin, mineral, and antioxidant content is anticonvulsant in mice, confirming that the KD's effect in mice can be attributed to the composition of the diet and not other dietary factors. Given that the anticonvulsant mechanism of the KD is still unknown, the anticonvulsant profile of the diet in different seizure models may help to decipher this mechanism. The implications of the findings that the KD is anticonvulsant in electrical seizure models are indicated. Further, the potential involvement of polyunsaturated fatty acids (PUFA) in the KD's anticonvulsant mechanism is discussed.

  9. A Mouse Model of Furosemide-Induced Overactive Bladder.

    PubMed

    Saporito, Michael S; Zuvich, Eva; DiCamillo, Amy

    2016-01-01

    Detailed in this unit is a mouse model of overactive bladder and urinary incontinence based on diuretic stress-induced urination. The procedure involves the use of a unique, highly sensitive, and automated urine capturing method to measure urinary latency, frequency, and void volume. Although this method was first described and validated using an anti-muscarinic drug used for treating overactive bladder, subsequent work has shown that effective non-cholinergic agents can be detected. These findings indicate good predictive value for this model regarding the possible clinical utility of test agents as treatments for overactive bladder, regardless of their site of action. © 2016 by John Wiley & Sons, Inc. PMID:27636110

  10. Using Mouse Models to Explore Genotype-Phenotype Relationship in Down Syndrome

    ERIC Educational Resources Information Center

    Salehi, Ahmad; Faizi, Mehrdad; Belichenko, Pavel V.; Mobley, William C.

    2007-01-01

    Down Syndrome (DS) caused by trisomy 21 is characterized by a variety of phenotypes and involves multiple organs. Sequencing of human chromosome 21 (HSA21) and subsequently of its orthologues on mouse chromosome 16 have created an unprecedented opportunity to explore the complex relationship between various DS phenotypes and the extra copy of…

  11. "Mouse Calls:" A Storytelling Approach to Teaching First Aid Skills to Young Children.

    ERIC Educational Resources Information Center

    Marchand, Nancy E.; McDermott, Robert J.

    1986-01-01

    Preschoolers are an important group to teach first aid skills because most of their injuries occur in the home. "Mouse Calls" is a program aimed at four-to-eight year olds that involves puppets who treat frostbite, a bee string, burns, bruises, possible broken bones, and minor lacerations. The program is briefly described. (MT)

  12. Investigation of cranial and other nerves in the mouse with muscular dystrophy.

    PubMed Central

    Biscoe, T J; Caddy, K W; Pallot, D J; Pehrson, U M

    1975-01-01

    In the muscular dystrophic mouse mutant there is an absence of Schwann cells over circumscribed lengths of all cranial nerves except for II (I was not examined) and the lesion involves the sympathetic system. Where present, Schwann cells do not produce myelin of normal thickness. The lesion is similar to that described for the spinal roots. Causation is discussed. Images PMID:1141926

  13. Mouse models of the laminopathies

    SciTech Connect

    Stewart, Colin L. . E-mail: stewartc@ncifcrf.gov; Kozlov, Serguei; Fong, Loren G.; Young, Stephen G. . E-mail: sgyoung@mednet.ucla.edu

    2007-06-10

    The A and B type lamins are nuclear intermediate filament proteins that comprise the bulk of the nuclear lamina, a thin proteinaceous structure underlying the inner nuclear membrane. The A type lamins are encoded by the lamin A gene (LMNA). Mutations in this gene have been linked to at least nine diseases, including the progeroid diseases Hutchinson-Gilford progeria and atypical Werner's syndromes, striated muscle diseases including muscular dystrophies and dilated cardiomyopathies, lipodystrophies affecting adipose tissue deposition, diseases affecting skeletal development, and a peripheral neuropathy. To understand how different diseases arise from different mutations in the same gene, mouse lines carrying some of the same mutations found in the human diseases have been established. We, and others have generated mice with different mutations that result in progeria, muscular dystrophy, and dilated cardiomyopathy. To further our understanding of the functions of the lamins, we also created mice lacking lamin B1, as well as mice expressing only one of the A type lamins. These mouse lines are providing insights into the functions of the lamina and how changes to the lamina affect the mechanical integrity of the nucleus as well as signaling pathways that, when disrupted, may contribute to the disease.

  14. Mouse Models of Tumor Immunotherapy.

    PubMed

    Ngiow, Shin Foong; Loi, Sherene; Thomas, David; Smyth, Mark J

    2016-01-01

    Immunotherapy is now evolving into a major therapeutic option for cancer patients. Such clinical advances also promote massive interest in the search for novel immunotherapy targets, and to understand the mechanism of action of current drugs. It is projected that a series of novel immunotherapy agents will be developed and assessed for their therapeutic activity. In light of this, in vivo experimental mouse models that recapitulate human malignancies serve as valuable tools to validate the efficacy and safety profile of immunotherapy agents, before their transition into clinical trials. In this review, we will discuss the major classes of experimental mouse models of cancer commonly used for immunotherapy assessment and provide examples to guide the selection of appropriate models. We present some new data concerning the utility of a carcinogen-induced tumor model for comparing immunotherapies and combining immunotherapy with chemotherapy. We will also highlight some recent advances in experimental modeling of human malignancies in mice that are leading towards personalized therapy in patients.

  15. Mouse models for liver cancer.

    PubMed

    Bakiri, Latifa; Wagner, Erwin F

    2013-04-01

    Hepatocellular carcinoma (HCC), the most common form of primary liver cancer is the third leading cause of cancer-related cell death in human and the fifth in women worldwide. The incidence of HCC is increasing despite progress in identifying risk factors, understanding disease etiology and developing anti-viral strategies. Therapeutic options are limited and survival after diagnosis is poor. Therefore, better preventive, diagnostic and therapeutic tools are urgently needed, in particular given the increased contribution from systemic metabolic disease to HCC incidence worldwide. In the last three decades, technological advances have facilitated the generation of genetically engineered mouse models (GEMMs) to mimic the alterations frequently observed in human cancers or to conduct intervention studies and assess the relevance of candidate gene networks in tumor establishment, progression and maintenance. Because these studies allow molecular and cellular manipulations impossible to perform in patients, GEMMs have improved our understanding of this complex disease and represent a source of great potential for mechanism-based therapy development. In this review, we provide an overview of the current state of HCC modeling in the mouse, highlighting successes, current challenges and future opportunities.

  16. Mouse Behavior: Conjectures about Adaptations for Survival.

    ERIC Educational Resources Information Center

    Rop, Charles

    2001-01-01

    Presents an experiment on mouse behavior in which students learn to observe, pay attention to details, record field notes, and ask questions about their observations. Uses a white mouse to eliminate the risk of disease that a wild rodent might carry. Lists materials, set up, and procedure. (YDS)

  17. Measuring Viscoelastic Deformation with an Optical Mouse

    ERIC Educational Resources Information Center

    Ng, T. W.

    2004-01-01

    The feasibility of using an optical mouse to track the viscoelastic deformation of low-density polyethylene films that have a fixed attached load is presented. It is seen that using an optical mouse and with rudimentary experiment paraphernalia and arrangement, it is possible to get good measurements of viscoelastic deformation.

  18. Expression of DMP1 in the developing mouse tongue embryo.

    PubMed

    Murata, Hidetaka; Sunohara, Msataka; Sato, Iwao

    2015-07-01

    Dentin matrix protein 1 (DMP-1) is an important factor in the mineralization of hard tissues. However, it has many other functions in addition to the regulation of mineralized tissues. We analyzed the expression and localization of DMP-1 by immunohistochemical staining and in situ hybridization in the developing mouse tongue during embryonic days 12.5 (E12.5), E14.5, E17.5, and E18.5. We also detected the mRNA abundance of tongue morphogenesis markers such as FGF6, TGF-β1, Collagen I, osteocalcin, chondromodulin 1, tenomodulin, Vascular endothelial growth factor (VEGF), caspase-3, and Aifm from embryonic stages by real-time RT-PCR. The antisense probe for DMP-1 was detected in a few mesenchymal cells surrounding blood vessels at E12.5, and faint localization was seen at E18.5 in the embryonic mouse tongue by in situ hybridization. The DMP-1 and osteocalcin abundance levels gradually increased compared with the other tongue markers from E12.5 to E18.5 (p<0.001). Cluster analyses identified the following distinct clusters for mRNA abundance in the tongue: cluster 1, E12.5; cluster 2, E14.5 and E17.5; and cluster 3, E18.5. The positive correlation between DMP-1 and osteocalcin (Pearson's r=0.685; p<0.05) and negative correlation between DMP-1 and Caspase-3 (Pearson's r=-0.632; p<0.05) were analyzed. These data suggested that DMP-1 potentially influences osteocalcin and Caspase-3 during mouse tongue development and morphogenesis. DMP-1 also affects the angiogenic marker VEGF in specific stages and areas, terminating the differentiation of the tongue from other developing tissues. We conclude that DMP-1 may be involved in regulating the temporal expression at embryonic stages in the mouse tongue.

  19. Mechanism of Oligonucleotide Uptake by Cells: Involvement of Specific receptors?

    NASA Astrophysics Data System (ADS)

    Yakubov, Leonid A.; Deeva, Elena A.; Zarytova, Valentina F.; Ivanova, Eugenia M.; Ryte, Antonina S.; Yurchenko, Lyudmila V.; Vlassov, Valentin V.

    1989-09-01

    We have investigated the interaction of oligonucleotides and their alkylating derivatives with mammalian cells. In experiments with L929 mouse fibroblast and Krebs 2 ascites carcinoma cells, it was found that cellular uptake of oligodeoxynucleotide derivatives is achieved by an endocytosis mechanism. Uptake is considerably more efficient at low oligomer concentration (< 1 μ M), because at this concentration a significant percentage of the total oligomer pool is absorbed on the cell surface and internalized by a more efficient absorptive endocytosis process. Two modified proteins were detected in mouse fibroblasts that were treated with the alkylating oligonucleotide derivatives. The binding of the oligomers to the proteins is inhibited by other oligodeoxynucleotides, single- and double-stranded DNA, and RNA. The polyanions heparin and chondroitin sulfates A and B do not inhibit binding. These observations suggest the involvement of specific receptor proteins in binding of oligomers to mammalian cells.

  20. The Mouse Genome Database (MGD): from genes to mice--a community resource for mouse biology.

    PubMed

    Eppig, Janan T; Bult, Carol J; Kadin, James A; Richardson, Joel E; Blake, Judith A; Anagnostopoulos, A; Baldarelli, R M; Baya, M; Beal, J S; Bello, S M; Boddy, W J; Bradt, D W; Burkart, D L; Butler, N E; Campbell, J; Cassell, M A; Corbani, L E; Cousins, S L; Dahmen, D J; Dene, H; Diehl, A D; Drabkin, H J; Frazer, K S; Frost, P; Glass, L H; Goldsmith, C W; Grant, P L; Lennon-Pierce, M; Lewis, J; Lu, I; Maltais, L J; McAndrews-Hill, M; McClellan, L; Miers, D B; Miller, L A; Ni, L; Ormsby, J E; Qi, D; Reddy, T B K; Reed, D J; Richards-Smith, B; Shaw, D R; Sinclair, R; Smith, C L; Szauter, P; Walker, M B; Walton, D O; Washburn, L L; Witham, I T; Zhu, Y

    2005-01-01

    The Mouse Genome Database (MGD) forms the core of the Mouse Genome Informatics (MGI) system (http://www.informatics.jax.org), a model organism database resource for the laboratory mouse. MGD provides essential integration of experimental knowledge for the mouse system with information annotated from both literature and online sources. MGD curates and presents consensus and experimental data representations of genotype (sequence) through phenotype information, including highly detailed reports about genes and gene products. Primary foci of integration are through representations of relationships among genes, sequences and phenotypes. MGD collaborates with other bioinformatics groups to curate a definitive set of information about the laboratory mouse and to build and implement the data and semantic standards that are essential for comparative genome analysis. Recent improvements in MGD discussed here include the enhancement of phenotype resources, the re-development of the International Mouse Strain Resource, IMSR, the update of mammalian orthology datasets and the electronic publication of classic books in mouse genetics.

  1. Immunochemical detection of arylamine N-acetyltransferase during mouse embryonic development and in adult mouse brain.

    PubMed

    Stanley, L A; Copp, A J; Pope, J; Rolls, S; Smelt, V; Perry, V H; Sim, E

    1998-11-01

    Arylamine N-acetyltransferases (NATs) are important in susceptibility to xenobiotic-induced disorders (e.g., drug-induced autoimmune disease, bladder cancer), but their role in endogenous metabolism is yet to be elucidated. The discovery that human NAT1 acts upon p-aminobenzoylgluatamate (p-ABG) to generate p-acetamidobenzoylglutamate (p-AABG), a major urinary metabolite of folic acid, suggests that human NAT1 may play a role in folic acid metabolism and hence in the normal development of the neural tube. In this study we examined the distribution of NAT in neuronal tissue from adult mice and embryos. Immunohistochemical staining of the adult mouse cerebellum revealed NAT2 (the mouse homologue of human NAT1) expression in the cell bodies and dendrites of Purkinje cells and in the neuroglia of the molecular layer. In embryos, NAT2 was detected in developing neuronal tissue on days 9.5, 11.5, and 13.5. It was expressed intensely in the nerual tube around the time of closure. The level of expression subsequently declined in the neuroepithelium but increased in glial cells. In addition, NAT2 was detected in the developing heart and gut. These findings demonstrate that the embryo itself expresses an enzyme which is involved in the metabolism of folic acid, so that the role played by both mother and embryo must be considered when examining the role of folic acid in embryonic development. These findings imply that polymorphisms in NAT genes could play a role in determining susceptibility to neural tube defects (NTD) and orofacial clefting, developmental disorders which can be prevented by dietary administration of folic acid. PMID:9839355

  2. Cardiac involvement in hemochromatosis.

    PubMed

    Gulati, Vinay; Harikrishnan, Prakash; Palaniswamy, Chandrasekar; Aronow, Wilbert S; Jain, Diwakar; Frishman, William H

    2014-01-01

    Cardiac hemochromatosis or primary iron-overload cardiomyopathy is an important and potentially preventable cause of heart failure. This is initially characterized by diastolic dysfunction and arrhythmias and in later stages by dilated cardiomyopathy. Diagnosis of iron overload is established by elevated transferrin saturation (>55%) and elevated serum ferritin (>300 ng/mL). Genetic testing for mutations in the HFE (high iron) gene and other proteins, such as hemojuvelin, transferrin receptor, and ferroportin, should be performed if secondary causes of iron overload are ruled out. Patients should undergo comprehensive 2D and Doppler echocardiography to evaluate their systolic and diastolic function. Newer modalities like strain imaging and speckle-tracking echocardiography hold promise for earlier detection of cardiac involvement. Cardiac magnetic resonance imaging with measurement of T2* relaxation times can help quantify myocardial iron overload. In addition to its value in diagnosis of cardiac iron overload, response to iron reduction therapy can be assessed by serial imaging. Therapeutic phlebotomy and iron chelation are the cornerstones of therapy. The average survival is less than a year in untreated patients with severe cardiac impairment. However, if treated early and aggressively, the survival rate approaches that of the regular heart failure population.

  3. Probing interfaces involving liquids.

    PubMed

    Robinson, A L

    1987-04-10

    Last month in Washington, D.C., the National Academy of Sciences held the first of what it hopes will be a series of seminars in forefront fields of science, technology, and medicine. The idea is to bring the academy closer to the frontlines of research and to help spread the word to federal science policy-makers. The subject of the 23 and 24 March seminar was interfaces and thin films, and the talks, though tutorial in nature, contained a pleasantly large number of still unpublished results. Interfaces, such as the surface of a solid exposed to a liquid or gas, and thin films, whose properties are heavily influenced by interfaces, have long been of considerable technological importance and have always been so in biological processes, but researchers are now getting access to the experimental and theoretical tools needed to explore these complex physical systems that are neither ideally two-dimensional nor fully three-dimensional. The briefings that follow give a peek at three ways to probe interfaces involving liquids.

  4. The Mouse Forced Swim Test

    PubMed Central

    Can, Adem; Dao, David T.; Arad, Michal; Terrillion, Chantelle E.; Piantadosi, Sean C.; Gould, Todd D.

    2012-01-01

    The forced swim test is a rodent behavioral test used for evaluation of antidepressant drugs, antidepressant efficacy of new compounds, and experimental manipulations that are aimed at rendering or preventing depressive-like states. Mice are placed in an inescapable transparent tank that is filled with water and their escape related mobility behavior is measured. The forced swim test is straightforward to conduct reliably and it requires minimal specialized equipment. Successful implementation of the forced swim test requires adherence to certain procedural details and minimization of unwarranted stress to the mice. In the protocol description and the accompanying video, we explain how to conduct the mouse version of this test with emphasis on potential pitfalls that may be detrimental to interpretation of results and how to avoid them. Additionally, we explain how the behaviors manifested in the test are assessed. PMID:22314943

  5. The mouse forced swim test.

    PubMed

    Can, Adem; Dao, David T; Arad, Michal; Terrillion, Chantelle E; Piantadosi, Sean C; Gould, Todd D

    2012-01-29

    The forced swim test is a rodent behavioral test used for evaluation of antidepressant drugs, antidepressant efficacy of new compounds, and experimental manipulations that are aimed at rendering or preventing depressive-like states. Mice are placed in an inescapable transparent tank that is filled with water and their escape related mobility behavior is measured. The forced swim test is straightforward to conduct reliably and it requires minimal specialized equipment. Successful implementation of the forced swim test requires adherence to certain procedural details and minimization of unwarranted stress to the mice. In the protocol description and the accompanying video, we explain how to conduct the mouse version of this test with emphasis on potential pitfalls that may be detrimental to interpretation of results and how to avoid them. Additionally, we explain how the behaviors manifested in the test are assessed.

  6. Applying Employee Involvement in Schools.

    ERIC Educational Resources Information Center

    Mohrman, Susan Albers; And Others

    1992-01-01

    The applicability of employee-involvement approaches to the management of schools is explored, describing three approaches (parallel-suggestion involvement, job involvement, and high involvement). Design issues (technology; organizational structure; leadership; organizational boundaries, customer definition, and relation to stakeholder; measures;…

  7. Mouse model of intracerebellar haemorrhage.

    PubMed

    Tijjani Salihu, Abubakar; Muthuraju, Sangu; Aziz Mohamed Yusoff, Abdul; Ahmad, Farizan; Zulkifli Mustafa, Mohd; Jaafar, Hasnan; Idris, Zamzuri; Rahman Izaini Ghani, Abdul; Malin Abdullah, Jafri

    2016-10-01

    The present study aimed to investigate the behavior and neuronal morphological changes in the perihaemorrhagic tissue of the mouse intracerebellar haemorrhage experimental model. Adult male Swiss albino mice were stereotactically infused with collagenase type VII (0.4U/μl of saline) unilaterally in to the cerebellum, following anaesthesia. Motor deficits were assessed using open field and composite score for evaluating the mouse model of cerebellar ataxia at 1, 3, 7, 14 and 21 days after collagenase infusion. The animals were sacrificed at the same time interval for evaluation of perihaematomal neuronal degeneration using haematoxylin and eosin staining and Annexin V-FITC/Propidium iodide assay. At the end of the study, it was found that infusion of 0.4U collagenase produces significant locomotor and ataxic deficit in the mice especially within the first week post surgery, and that this gradually improved within three weeks. Neuronal degeneration evident by cytoplasmic shrinkage and nuclear pyknosis was observed at the perihaematomal area after one day; especially at 3 and 7 days post haemorrhage. By 21 days, both the haematoma and degenerating neurons in the perihaematomal area were phagocytosed and the remaining neuronal cells around the scar tissue appeared normal. Moreover, Annexin-V/propidium iodide-positive cells were observed at the perihaematomal area at 3 and 7 days implying that the neurons likely die via apoptosis. It was concluded that a population of potentially salvageable neurons exist in the perihaematomal area after cerebellar haemorrhage throughout a wide time window that could be amenable to treatment. PMID:27327104

  8. Prion infection of mouse neurospheres

    PubMed Central

    Giri, Ranjit K.; Young, Rebecca; Pitstick, Rose; DeArmond, Stephen J.; Prusiner, Stanley B.; Carlson, George A.

    2006-01-01

    Only a few cell lines have been infected with prions, offering limited genetic diversity and sensitivity to several strains. Here we report that cultured neurospheres expressing cellular prion protein (PrPC) can be infected with prions. Neurosphere lines isolated from the brains of mice at embryonic day 13–15 grow as aggregates and contain CNS stem cells. We produced neurosphere cultures from FVB/NCr (FVB) mice, from transgenic (Tg) FVB mice that overexpress mouse PrP-A (Tg4053), and from congenic FVB mice with a targeted null mutation in the PrP gene (Prnp0/0) and incubated them with the Rocky Mountain Laboratory prion strain. While monitoring the levels of disease-causing PrP (PrPSc) at each passage, we observed a dramatic rise in PrPSc levels with time in the Tg4053 neurosphere cells, whereas the level of PrPSc decayed to undetectable levels in cell cultures lacking PrP. PrPSc levels in cultures from FVB mice initially declined but then increased with passage. Prions produced in culture were transmissible to mice and produced disease pathology. Intracellular aggregates of PrPSc were present in cells from infected cultures. The susceptibility of neurosphere cultures to prions mirrored that of the mice from which they were derived. Neurosphere lines from Tg4053 mice provide a sensitive in vitro bioassay for mouse prions; neurosphere lines from other Tg mice overexpressing PrP might be used to assay prions from other species, including humans. PMID:16495413

  9. Establishment of crown-root domain borders in mouse incisor.

    PubMed

    Juuri, Emma; Saito, Kan; Lefebvre, Sylvie; Michon, Frederic

    2013-10-01

    Teeth are composed of two domains, the enamel-covered crown and the enamel-free root. The understanding of the initiation and regulation of crown and root domain formation is important for the development of bioengineered teeth. In most teeth the crown develops before the root, and erupts to the oral cavity whereas the root anchors the tooth to the jawbone. However, in the continuously growing mouse incisor the crown and root domains form simultaneously, the crown domain forming the labial and the root domain the lingual part of the tooth. While the crown-root border on the incisor distal side supports the distal enamel extent, reflecting an evolutionary diet adaptation, on the incisor mesial side the root-like surface is necessary for the attachment of the interdental ligament between the two incisors. Therefore, the mouse incisor exhibits a functional distal-mesial asymmetry. Here, we used the mouse incisor as a model to understand the mechanisms involved in the crown-root border formation. We analyzed the cellular origins and gene expression patterns leading to the development of the mesial and distal crown-root borders. We discovered that Barx2, En1, Wnt11, and Runx3 were exclusively expressed on the mesial crown-root border. In addition, the distal border of the crown-root domain might be established by cells from a different origin and by an early Follistatin expression, factor known to be involved in the root domain formation. The use of different mechanisms to establish domain borders gives indications of the incisor functional asymmetry.

  10. Mineral metabolism in isolated mouse long bones: Opposite effects of microgravity on mineralization and resorption

    NASA Technical Reports Server (NTRS)

    Veldhuijzen, Jean Paul; Vanloon, Jack J. W. A.

    1994-01-01

    An experiment using isolated skeletal tissues under microgravity, is reported. Fetal mouse long bones (metatarsals) were cultured for 4 days in the Biorack facility of Spacelab during the IML-1 (International Microgravity Laboratory) mission of the Space Shuttle. Overall growth was not affected, however glucose consumption was significantly reduced under microgravity. Mineralization of the diaphysis was also strongly reduced under microgravity as compared to the on-board 1 g group. In contrast, mineral resorption by osteoclasts was signficantly increased. These results indicate that these fetal mouse long bones are a sensitive and useful model to further study the cellular mechanisms involved in the changed mineral metabolism of skeletal tissues under microgravity.

  11. Ultrasound biomicroscopy in mouse cardiovascular development

    NASA Astrophysics Data System (ADS)

    Turnbull, Daniel H.

    2001-05-01

    The mouse is the preferred animal model for studying mammalian cardiovascular development and many human congenital heart diseases. Ultrasound biomicroscopy (UBM), utilizing high-frequency (40-50-MHz) ultrasound, is uniquely capable of providing in vivo, real-time microimaging and Doppler blood velocity measurements in mouse embryos and neonates. UBM analyses of normal and abnormal mouse cardiovascular function will be described to illustrate the power of this microimaging approach. In particular, real-time UBM images have been used to analyze dimensional changes in the mouse heart from embryonic to neonatal stages. UBM-Doppler has been used recently to examine the precise timing of onset of a functional circulation in early-stage mouse embryos, from the first detectable cardiac contractions. In other experiments, blood velocity waveforms have been analyzed to characterize the functional phenotype of mutant mouse embryos having defects in cardiac valve formation. Finally, UBM has been developed for real-time, in utero image-guided injection of mouse embryos, enabling cell transplantation and genetic gain-of-function experiments with transfected cells and retroviruses. In summary, UBM provides a unique and powerful approach for in vivo analysis and image-guided manipulation in normal and genetically engineered mice, over a wide range of embryonic to neonatal developmental stages.

  12. Spatial gene expression in the T-stage mouse metanephros.

    PubMed

    Caruana, Georgina; Cullen-McEwen, Luise; Nelson, Amy L; Kostoulias, Xenia; Woods, Kyra; Gardiner, Brooke; Davis, Melissa J; Taylor, Darrin F; Teasdale, Rohan D; Grimmond, Sean M; Little, Melissa H; Bertram, John F

    2006-10-01

    The E11.5 mouse metanephros is comprised of a T-stage ureteric epithelial tubule sub-divided into tip and trunk cells surrounded by metanephric mesenchyme (MM). Tip cells are induced to undergo branching morphogenesis by the MM. In contrast, signals within the mesenchyme surrounding the trunk prevent ectopic branching of this region. In order to identify novel genes involved in the molecular regulation of branching morphogenesis we compared the gene expression profiles of isolated tip, trunk and MM cells using Compugen mouse long oligo microarrays. We identified genes enriched in the tip epithelium, sim-1, Arg2, Tacstd1, Crlf-1 and BMP7; genes enriched in the trunk epithelium, Innp1, Itm2b, Mkrn1, SPARC, Emu2 and Gsta3 and genes spatially restricted to the mesenchyme surrounding the trunk, CSPG2 and CV-2, with overlapping and complimentary expression to BMP4, respectively. This study has identified genes spatially expressed in regions of the developing kidney involved in branching morphogenesis, nephrogenesis and the development of the collecting duct system, calyces, renal pelvis and ureter.

  13. Isolation and Characterization of Pluripotent Human Spermatogonial Stem Cell-Derived Cells

    PubMed Central

    Kossack, Nina; Meneses, Juanito; Shefi, Shai; Nguyen, Ha Nam; Chavez, Shawn; Nicholas, Cory; Gromoll, Joerg; Turek, Paul J; Reijo-Pera, Renee A

    2009-01-01

    Several reports have documented the derivation of pluripotent cells (multipotent germline stem cells) from spermatogonial stem cells obtained from the adult mouse testis. These spermatogonia-derived stem cells express embryonic stem cell markers and differentiate to the three primary germ layers, as well as the germline. Data indicate that derivation may involve reprogramming of endogenous spermatogonia in culture. Here, we report the derivation of human multipotent germline stem cells (hMGSCs) from a testis biopsy. The cells express distinct markers of pluripotency, form embryoid bodies that contain derivatives of all three germ layers, maintain a normal XY karyotype, are hypomethylated at the H19 locus, and express high levels of telomerase. Teratoma assays indicate the presence of human cells 8 weeks post-transplantation but limited teratoma formation. Thus, these data suggest the potential to derive pluripotent cells from human testis biopsies but indicate a need for novel strategies to optimize hMGSC culture conditions and reprogramming. PMID:18927477

  14. Humanization of the mouse mammary gland.

    PubMed

    Wronski, A; Arendt, L M; Kuperwasser, Charlotte

    2015-01-01

    Although mouse models have provided invaluable information on the mechanisms of mammary gland development, anatomical and developmental differences between human and mice limit full understanding of this fundamental process. Humanization of the mouse mammary gland by injecting immortalized human breast stromal cells into the cleared murine mammary fat pad enables the growth and development of human mammary epithelial cells or tissue. This facilitates the characterization of human mammary gland development or tumorigenesis by utilizing the mouse mammary fat pad. Here we describe the process of isolating human mammary stromal and epithelial cells as well as their introduction into the mammary fat pads of immunocompromised mice.

  15. Augmented Computer Mouse Would Measure Applied Force

    NASA Technical Reports Server (NTRS)

    Li, Larry C. H.

    1993-01-01

    Proposed computer mouse measures force of contact applied by user. Adds another dimension to two-dimensional-position-measuring capability of conventional computer mouse; force measurement designated to represent any desired continuously variable function of time and position, such as control force, acceleration, velocity, or position along axis perpendicular to computer video display. Proposed mouse enhances sense of realism and intuition in interaction between operator and computer. Useful in such applications as three-dimensional computer graphics, computer games, and mathematical modeling of dynamics.

  16. Spinal Cord Ventral Horns and Lymphoid Organ Involvement in Powassan Virus Infection in a Mouse Model.

    PubMed

    Santos, Rodrigo I; Hermance, Meghan E; Gelman, Benjamin B; Thangamani, Saravanan

    2016-01-01

    Powassan virus (POWV) belongs to the family Flaviviridae and is a member of the tick-borne encephalitis serogroup. Transmission of POWV from infected ticks to humans has been documented in the USA, Canada, and Russia, causing fatal encephalitis in 10% of human cases and significant neurological sequelae in survivors. We used C57BL/6 mice to investigate POWV infection and pathogenesis. After footpad inoculation, infected animals exhibited rapid disease progression and 100% mortality. Immunohistochemistry and immunofluorescence revealed a very strong neuronal tropism of POWV infection. The central nervous system infection appeared as a meningoencephalitis with perivascular mononuclear infiltration and microglial activation in the brain, and a poliomyelitis-like syndrome with high level of POWV antigen at the ventral horn of the spinal cord. Pathological studies also revealed substantial infection of splenic macrophages by POWV, which suggests that the spleen plays a more important role in pathogenesis than previously realized. This report provides a detailed description of the neuroanatomical distribution of the lesions produced by POWV infection in C57BL/6 mice. PMID:27529273

  17. Spinal Cord Ventral Horns and Lymphoid Organ Involvement in Powassan Virus Infection in a Mouse Model

    PubMed Central

    Santos, Rodrigo I.; Hermance, Meghan E.; Gelman, Benjamin B.; Thangamani, Saravanan

    2016-01-01

    Powassan virus (POWV) belongs to the family Flaviviridae and is a member of the tick-borne encephalitis serogroup. Transmission of POWV from infected ticks to humans has been documented in the USA, Canada, and Russia, causing fatal encephalitis in 10% of human cases and significant neurological sequelae in survivors. We used C57BL/6 mice to investigate POWV infection and pathogenesis. After footpad inoculation, infected animals exhibited rapid disease progression and 100% mortality. Immunohistochemistry and immunofluorescence revealed a very strong neuronal tropism of POWV infection. The central nervous system infection appeared as a meningoencephalitis with perivascular mononuclear infiltration and microglial activation in the brain, and a poliomyelitis-like syndrome with high level of POWV antigen at the ventral horn of the spinal cord. Pathological studies also revealed substantial infection of splenic macrophages by POWV, which suggests that the spleen plays a more important role in pathogenesis than previously realized. This report provides a detailed description of the neuroanatomical distribution of the lesions produced by POWV infection in C57BL/6 mice. PMID:27529273

  18. Involvement of PrP(C) in kainate-induced excitotoxicity in several mouse strains.

    PubMed

    Carulla, Patricia; Llorens, Franc; Matamoros-Angles, Andreu; Aguilar-Calvo, Patricia; Espinosa, Juan Carlos; Gavín, Rosalina; Ferrer, Isidre; Legname, Giuseppe; Torres, Juan Maria; del Río, José A

    2015-01-01

    The cellular prion protein (PrP(C)) has been associated with a plethora of cellular functions ranging from cell cycle to neuroprotection. Mice lacking PrP(C) show an increased susceptibility to epileptic seizures; the protein, then, is neuroprotective. However, lack of experimental reproducibility has led to considering the possibility that other factors besides PrP(C) deletion, such as the genetic background of mice or the presence of so-called "Prnp flanking genes", might contribute to the reported susceptibility. Here, we performed a comparative analysis of seizure-susceptibility using characterized Prnp(+/+) and Prnp(0/0) mice of B6129, B6.129, 129/Ola or FVB/N genetic backgrounds. Our study indicates that PrP(C) plays a role in neuroprotection in KA-treated cells and mice. For this function, PrP(C) should contain the aa32-93 region and needs to be linked to the membrane. In addition, some unidentified "Prnp-flanking genes" play a role parallel to PrP(C) in the KA-mediated responses in B6129 and B6.129 Prnp(0/0) mice. PMID:26155834

  19. Involvement of Mouse Constitutive Androstane Receptor in Acifluorfen-Induced Liver Injury and Subsequent Tumor Development.

    PubMed

    Kuwata, Kazunori; Inoue, Kaoru; Ichimura, Ryohei; Takahashi, Miwa; Kodama, Yukio; Shibutani, Makoto; Yoshida, Midori

    2016-06-01

    Acifluorfen (ACI), a protoporphyrinogen oxidase (PROTOX) inhibitor herbicide, promotes the accumulation of protoporphyrin IX (PPIX), and induces tumors in the rodent liver. Porphyria is a risk factor for liver tumors in humans; however, the specific mechanisms through which ACI induces hepatocarcinogenesis in rodents are unclear. Here, we investigated the mode of action of ACI-induced hepatocarcinogenesis, focusing on constitutive androstane receptor (CAR, NR1I3), which is essential for the development of rodent liver tumors in response to certain cytochrome P450 (CYP) 2B inducers. Dietary treatment with 2500 ppm ACI for up to 13 weeks increased Cyp2b10 expression in the livers of wild-type (WT) mice, but not in CAR-knockout (CARKO) mice. Microscopically, ACI treatment-induced cytotoxic changes, including hepatocellular necrosis and inflammation, and caused regenerative changes accompanied by prolonged increases in the numbers of proliferating cell nuclear antigen-positive hepatocytes in WT mice. In contrast, these cytotoxic and regenerative changes in hepatocytes were significantly attenuated, but still observed, in CARKO mice. ACI treatment also increased liver PPIX levels similarly in both genotypes; however, no morphological evidence of porphyrin deposition was found in hepatocytes from either genotype. Treatment with 2500 ppm ACI for 26 weeks after initiation with diethylnitrosamine increased the incidence and multiplicities of altered foci and adenomas in hepatocytes from WT mice; these effects were significantly reduced in CARKO mice. These results indicated that prolonged cytotoxicity in the liver was a key factor for ACI-induced hepatocarcinogenesis, and that CAR played an important role in ACI-induced liver injury and tumor development in mice. PMID:26928356

  20. Involvement of PrPC in kainate-induced excitotoxicity in several mouse strains

    PubMed Central

    Carulla, Patricia; Llorens, Franc; Matamoros-Angles, Andreu; Aguilar-Calvo, Patricia; Espinosa, Juan Carlos; Gavín, Rosalina; Ferrer, Isidre; Legname, Giuseppe; Torres, Juan Maria; del Río, José A.

    2015-01-01

    The cellular prion protein (PrPC) has been associated with a plethora of cellular functions ranging from cell cycle to neuroprotection. Mice lacking PrPC show an increased susceptibility to epileptic seizures; the protein, then, is neuroprotective. However, lack of experimental reproducibility has led to considering the possibility that other factors besides PrPC deletion, such as the genetic background of mice or the presence of so-called “Prnp flanking genes”, might contribute to the reported susceptibility. Here, we performed a comparative analysis of seizure-susceptibility using characterized Prnp+/+ and Prnp0/0 mice of B6129, B6.129, 129/Ola or FVB/N genetic backgrounds. Our study indicates that PrPC plays a role in neuroprotection in KA-treated cells and mice. For this function, PrPC should contain the aa32–93 region and needs to be linked to the membrane. In addition, some unidentified “Prnp-flanking genes” play a role parallel to PrPC in the KA-mediated responses in B6129 and B6.129 Prnp0/0 mice. PMID:26155834

  1. An integrase of endogenous retrovirus is involved in maternal mitochondrial DNA inheritance of the mouse.

    PubMed

    Hayashida, Kenji; Omagari, Katsuhisa; Masuda, Jun-Ichi; Kohno, Shigeru

    2008-02-01

    The mechanism of maternal mitochondrial DNA (mtDNA) inheritance in animals can be said to be the selective elimination of sperm mtDNA via the elimination factor of the egg and a sperm mitochondria-specific factor. In 2005, we clarified that t-tpis (Spag1 isoform 1) is a mitochondria-specific translocator and the sperm factor, and furthermore estimated that the elimination factors of the egg are the divalent cation-dependent endonuclease and s-tpis (Spag1 isoform 2 and isoform 3) as the elimination system-specific chaperone [K. Hayashida, K. Omagari, J. Masuda, H. Hazama, Y. Kadokawa, K. Ohba, S. Kohno, The sperm mitochondria-specific translocator has a key role in maternal mitochondrial inheritance, Cell Biol. Int. 29 (2005) 472-481]. This time, using a recombinant Spag1 isoform 1 protein, a pull-down assay of ovary cytosol was performed and the elimination factors searched for. Surprisingly, an endogenous retroviral integrase fragment (Eri15) was identified using mass spectrometry of the electrophoresis band of the pull-down protein. Eri15 was detected as a complex of approximately 500kDa with Spag1 isoform 2 or isoform 3 in native PAGE of the ovary cytosol. This strongly suggested that Eri15 is selectively transported into the sperm mitochondria matrix by Spag1 isoform 2 and 3 via Spag1 isoform 1 and that sperm mtDNA is destroyed, thus causing the establishment of maternal mtDNA inheritance.

  2. Finding mouse models of human lymphomas and leukemia's using the Jackson laboratory mouse tumor biology database.

    PubMed

    Begley, Dale A; Sundberg, John P; Krupke, Debra M; Neuhauser, Steven B; Bult, Carol J; Eppig, Janan T; Morse, Herbert C; Ward, Jerrold M

    2015-12-01

    Many mouse models have been created to study hematopoietic cancer types. There are over thirty hematopoietic tumor types and subtypes, both human and mouse, with various origins, characteristics and clinical prognoses. Determining the specific type of hematopoietic lesion produced in a mouse model and identifying mouse models that correspond to the human subtypes of these lesions has been a continuing challenge for the scientific community. The Mouse Tumor Biology Database (MTB; http://tumor.informatics.jax.org) is designed to facilitate use of mouse models of human cancer by providing detailed histopathologic and molecular information on lymphoma subtypes, including expertly annotated, on line, whole slide scans, and providing a repository for storing information on and querying these data for specific lymphoma models. PMID:26302176

  3. Mouse homologue of yeast Prp19 interacts with mouse SUG1, the regulatory subunit of 26S proteasome

    SciTech Connect

    Sihn, Choong-Ryoul; Cho, Si Young; Lee, Jeong Ho; Lee, Tae Ryong; Kim, Sang Hoon . E-mail: shkim@khu.ac.kr

    2007-04-27

    Yeast Prp19 has been shown to involve in pre-mRNA splicing and DNA repair as well as being an ubiquitin ligase. Mammalian homologue of yeast Prp19 also plays on similar functional activities in cells. In the present study, we isolated mouse SUG1 (mSUG1) as binding partner of mouse Prp19 (mPrp19) by the yeast two-hybrid system. We confirmed the interaction of mPrp9 with mSUG1 by GST pull-down assay and co-immunoprecipitation assay. The N-terminus of mPrp19 including U-box domain was associated with the C-terminus of mSUG1. Although, mSUG1 is a regulatory subunit of 26S proteasome, mPrp19 was not degraded in the proteasome-dependent pathway. Interestingly, GFP-mPrp19 fusion protein was co-localized with mSUG1 protein in cytoplasm as the formation of the speckle-like structures in the presence of a proteasome inhibitor MG132. In addition, the activity of proteasome was increased in cells transfected with mPrp19. Taken together, these results suggest that mPrp19 involves the regulation of protein turnover and may transport its substrates to 26S proteasome through mSUG1 protein.

  4. Galantamine improves olfactory learning in the Ts65Dn mouse model of Down syndrome

    PubMed Central

    Simoes de Souza, Fabio M.; Busquet, Nicolas; Blatner, Megan; Maclean, Kenneth N.; Restrepo, Diego

    2011-01-01

    Down syndrome (DS) is the most common form of congenital intellectual disability. Although DS involves multiple disturbances in various tissues, there is little doubt that in terms of quality of life cognitive impairment is the most serious facet and there is no effective treatment for this aspect of the syndrome. The Ts65Dn mouse model of DS recapitulates multiple aspects of DS including cognitive impairment. Here the Ts65Dn mouse model of DS was evaluated in an associative learning paradigm based on olfactory cues. In contrast to disomic controls, trisomic mice exhibited significant deficits in olfactory learning. Treatment of trisomic mice with the acetylcholinesterase inhibitor galantamine resulted in a significant improvement in olfactory learning. Collectively, our study indicates that olfactory learning can be a sensitive tool for evaluating deficits in associative learning in mouse models of DS and that galantamine has therapeutic potential for improving cognitive abilities. PMID:22355654

  5. Did the house mouse (Mus musculus L.) shape the evolutionary trajectory of wheat (Triticum aestivum L.)?

    PubMed Central

    Morris, C F; Fuerst, E P; Beecher, B S; Mclean, D J; James, C P; Geng, H W

    2013-01-01

    Wheat (Triticum aestivum L.) is one of the most successful domesticated plant species in the world. The majority of wheat carries mutations in the Puroindoline genes that result in a hard kernel phenotype. An evolutionary explanation, or selective advantage, for the spread and persistence of these hard kernel mutations has yet to be established. Here, we demonstrate that the house mouse (Mus musculus L.) exerts a pronounced feeding preference for soft over hard kernels. When allele frequencies ranged from 0.5 to 0.009, mouse predation increased the hard allele frequency as much as 10-fold. Studies involving a single hard kernel mixed with ∼1000 soft kernels failed to recover the mutant kernel. Nevertheless, the study clearly demonstrates that the house mouse could have played a role in the evolution of wheat, and therefore the cultural trajectory of humankind. PMID:24223281

  6. Cardiac-Autonomic Imbalance and Baroreflex Dysfunction in the Renovascular Angiotensin-Dependent Hypertensive Mouse

    PubMed Central

    Campagnaro, Bianca P.; Gava, Agata L.; Meyrelles, Silvana S.; Vasquez, Elisardo C.

    2012-01-01

    Mouse models provide powerful tools for studying the mechanisms underlying the dysfunction of the autonomic reflex control of cardiovascular function and those involved in cardiovascular diseases. The established murine model of two-kidney, one-clip (2K1C) angiotensin II-dependent hypertension represents a useful tool for studying the neural control of cardiovascular function. In this paper, we discuss the main contributions from our laboratory and others regarding cardiac-autonomic imbalance and baroreflex dysfunction. We show recent data from the angiotensin-dependent hypertensive mouse demonstrating DNA damage and oxidative stress using the comet assay and flow cytometry, respectively. Finally, we highlight the relationships between angiotensin and peripheral and central nervous system areas of cardiovascular control and oxidative stress in the 2K1C hypertensive mouse. PMID:23193440

  7. Development of the vestigial tooth primordia as part of mouse odontogenesis.

    PubMed

    Peterková, R; Peterka, M; Viriot, L; Lesot, H

    2002-01-01

    The mouse functional dentition comprises one incisor separated from three molars by a toothless diastema in each dental quadrant. Between the incisor and molars, the embryonic tooth pattern also includes vestigial dental primordia, which undergo regression involving apoptosis in their epithelium. Apoptosis appears to play an important role in achieving the specific tooth pattern in the mouse. We documented similarities in the folding mechanism allowing the formation of the dental lamina in mice as well as in reptiles. While further budding on this dental lamina gives rise to many individual simple tooth primordia in crocodiles and lizards, budding morphogenesis of several simple tooth primordia appears to be integrated in the mouse, giving rise to enamel organs of a complex nature. The differentiation of a mammalian tooth germ during both ontogeny and phylogeny might thus include the concrescence (connation) of more primordia, putatively corresponding to simple teeth in mammalian ancestors.

  8. Genomic organization of mouse gene zfp162.

    PubMed

    Wrehlke, C; Wiedemeyer, W R; Schmitt-Wrede, H P; Mincheva, A; Lichter, P; Wunderlich, F

    1999-05-01

    We report the cloning and characterization of the alternatively spliced mouse gene zfp162, formerly termed mzfm, the homolog of the human ZFM1 gene encoding the splicing factor SF1 and a putative signal transduction and activation of RNA (STAR) protein. The zfp162 gene is about 14 kb long and consists of 14 exons and 13 introns. Comparison of zfp162 with the genomic sequences of ZFM1/SF1 revealed that the exon-intron structure and exon sequences are well conserved between the genes, whereas the introns differ in length and sequence composition. Using fluorescent in situ hybridization, the zfp162 gene was assigned to chromosome 19, region B. Screening of a genomic library integrated in lambda DASH II resulted in the identification of the 5'-flanking region of zfp162. Sequence analysis of this region showed that zfp162 is a TATA-less gene containing an initiator control element and two CCAAT boxes. The promoter exhibits the following motifs: AP-2, CRE, Ets, GRE, HNF5, MRE, SP-1, TRE, TCF1, and PU.1. The core promoter, from position -331 to -157, contains the motifs CRE, SP-1, MRE, and AP-2, as determined in transfected CHO-K1 cells and IC-21 cells by reporter gene assay using a secreted form of human placental alkaline phosphatase. The occurrence of PU.1/GRE supports the view that the zfp162 gene encodes a protein involved not only in nuclear RNA metabolism, as the human ZFM1/SF1, but also in as yet unknown macrophage-inherent functions. PMID:10360842

  9. Sequence Diversity, Intersubgroup Relationships, and Origins of the Mouse Leukemia Gammaretroviruses of Laboratory and Wild Mice

    PubMed Central

    Bamunusinghe, Devinka; Naghashfar, Zohreh; Buckler-White, Alicia; Plishka, Ronald; Baliji, Surendranath; Liu, Qingping; Kassner, Joshua; Oler, Andrew J.; Hartley, Janet

    2016-01-01

    house mouse evolution. We did this because the laboratory mouse viruses derive directly from specific ERVs or arise by recombination between different ERVs. The six distinctively different wild mouse viruses appear to be recombinants, often involving different host range subgroups, and most are related to a distinctive, largely Y-chromosome-linked MLV ERV subtype. MLVs with ecotropic host ranges show the greatest variability with extensive inter- and intrasubtype envelope differences and with homologies to other host range subgroups outside the envelope. The sequence diversity among these wild mouse isolates helps define their relationships and origins and emphasizes the importance of recombination in their evolution. PMID:26865715

  10. Human and mouse ABCA1 comparative sequencing and transgenesis studies identify regulatory elements

    SciTech Connect

    Qiu, Yang; Cavelier, L.; Chiu, Sally; Rubin, Edward; Cheng, Jan-Fang

    2000-08-01

    The expression of ABCA1, a major participant in apolipoprotein mediated cholesterol efflux is highly regulated by a variety of factors including intracellular cholesterol concentration. To analyze its genomic organization and identify those sequences involved in its regulation we sequenced and compared approximately 200 Kb of orthologous DNA from mice and humans containing the ABCA1 gene and significant flanking DNA. The comparison revealed a variety of mouse human conserved sequences including 50 conserved ABCA1 exons over 147Kb of human and 124Kb of mouse genomic DNA as well as multiple mouse human conserved noncoding sequences. Using as a criteria for identifying putative regulatory elements in non-coding sequence, human and mouse sequences that were &62;75% identical for over 120 bp were screened for resulting in the identification of 34 elements. The two most highly conserved human mouse noncoding elements (CNS1: 88% identity over 498 bp, CNS2: 81% identity over 214 bp)! were also highly conserved in the ABCA1 genes of rats, dogs, cows, rabbits and pigs. Two independent studies have demonstrated that the DNA segments containing CNS2 function in vitro as a sterol response promoter. Support for the inclusion of major ABCA1 regulatory elements in the human genomic sequence examined was the demonstration that mice containing a human BAC transgene containing sequences exclusively from the analyzed interval, expressed human ABCA1 in a tissue distribution mimicking expression of endogenous mouse ABC1. These studies using a comparative genomic approach has characterized the structure of the human and mouse ABCA1 genes and has helped identify sequences participating in its expression.

  11. Characterization of the Mouse Brain Proteome Using Global Proteomic Analysis Complemented with Cysteinyl-Peptide Enrichment

    PubMed Central

    Wang, Haixing; Qian, Wei-Jun; Chin, Mark H.; Petyuk, Vladislav A.; Barry, Richard C.; Liu, Tao; Gritsenko, Marina A.; Mottaz, Heather M.; Moore, Ronald J.; Camp, David G.; Khan, Arshad H.; Smith, Desmond J.; Smith, Richard D.

    2007-01-01

    Given the growing interest in applying genomic and proteomic approaches for studying the mammalian brain using mouse models, we hereby present a global proteomic approach for analyzing brain tissue and for the first time a comprehensive characterization of the whole mouse brain proteome. Preparation of the whole brain sample incorporated a highly efficient cysteinyl-peptide enrichment (CPE) technique to complement a global enzymatic digestion method. Both the global and the cysteinyl-enriched peptide samples were analyzed by SCX fractionation coupled with reversed phase LC-MS/MS analysis. A total of 48,328 different peptides were confidently identified (>98% confidence level), covering 7792 non-redundant proteins (∼34% of the predicted mouse proteome). 1564 and 1859 proteins were identified exclusively from the cysteinyl-peptide and the global peptide samples, respectively, corresponding to 25% and 31% improvements in proteome coverage compared to analysis of only the global peptide or cysteinyl-peptide samples. The identified proteins provide a broad representation of the mouse proteome with little bias evident due to protein pI, molecular weight, and/or cellular localization. Approximately 26% of the identified proteins with gene ontology (GO) annotations were membrane proteins, with 1447 proteins predicted to have transmembrane domains, and many of the membrane proteins were found to be involved in transport and cell signaling. The MS/MS spectrum count information for the identified proteins was used to provide a measure of relative protein abundances. The mouse brain peptide/protein database generated from this study represents the most comprehensive proteome coverage for the mammalian brain to date, and the basis for future quantitative brain proteomic studies using mouse models. The proteomic approach presented here may have broad applications for rapid proteomic analyses of various mouse models of human brain diseases. PMID:16457602

  12. The Mouse Gene Expression Database (GXD)

    PubMed Central

    Ringwald, Martin; Eppig, Janan T.; Begley, Dale A.; Corradi, John P.; McCright, Ingeborg J.; Hayamizu, Terry F.; Hill, David P.; Kadin, James A.; Richardson, Joel E.

    2001-01-01

    The Gene Expression Database (GXD) is a community resource of gene expression information for the laboratory mouse. By combining the different types of expression data, GXD aims to provide increasingly complete information about the expression profiles of genes in different mouse strains and mutants, thus enabling valuable insights into the molecular networks that underlie normal development and disease. GXD is integrated with the Mouse Genome Database (MGD). Extensive interconnections with sequence databases and with databases from other species, and the development and use of shared controlled vocabularies extend GXD’s utility for the analysis of gene expression information. GXD is accessible through the Mouse Genome Informatics web site at http://www.informatic s.jax.org/ or directly at http://www.informatics.jax.org/me nus/expression_menu.shtml. PMID:11125060

  13. Integration of Mouse Phenome Data Resources

    SciTech Connect

    Hancock, John M; Adams, Neils; Aidinis, Vassilis; Blake, Judith A; Bogue, Molly; Brown, Steve D M; Chesler, Elissa J; Davidson, Duncan; Duran, Christopher; Eppig, Janan T; Gailus-Durner, Valerie; Gkoutos, Georgios V; Greenaway, Simon; Angelis, Martin Hrabe de; Kollias, George; Leblanc, Sophie; Lee, Kirsty; Lengger, Christoph; Maier, Holger; Mallon, Ann-Marie; Masuya, Hiroshi; Melvin, David; Muller, Werner; Parkinson, Helen; Proctor, Glenn; Reuveni, Eli; Schofield, Paul; Shukla, Aadya; Smith, Cynthia; Toyoda, Tetsuro; Vasseur, Laurent; Wakana, Shigeharu; Walling, Alison; White, Jacqui; Wood, Joe; Zouberakis, Michalis

    2008-01-01

    Understanding the functions encoded in the mouse genome will be central to an understanding of the genetic basis of human disease. To achieve this it will be essential to be able to characterise the phenotypic consequences of variation and alterations in individual genes. Data on the phenotypes of mouse strains are currently held in a number of different forms (detailed descriptions of mouse lines, first line phenotyping data on novel mutations, data on the normal features of inbred lines, etc.) at many sites worldwide. For the most efficient use of these data sets, we have set in train a process to develop standards for the description of phenotypes (using ontologies), and file formats for the description of phenotyping protocols and phenotype data sets. This process is ongoing, and needs to be supported by the wider mouse genetics and phenotyping communities to succeed. We invite interested parties to contact us as we develop this process further.

  14. A catalog of the mouse gut metagenome.

    PubMed

    Xiao, Liang; Feng, Qiang; Liang, Suisha; Sonne, Si Brask; Xia, Zhongkui; Qiu, Xinmin; Li, Xiaoping; Long, Hua; Zhang, Jianfeng; Zhang, Dongya; Liu, Chuan; Fang, Zhiwei; Chou, Joyce; Glanville, Jacob; Hao, Qin; Kotowska, Dorota; Colding, Camilla; Licht, Tine Rask; Wu, Donghai; Yu, Jun; Sung, Joseph Jao Yiu; Liang, Qiaoyi; Li, Junhua; Jia, Huijue; Lan, Zhou; Tremaroli, Valentina; Dworzynski, Piotr; Nielsen, H Bjørn; Bäckhed, Fredrik; Doré, Joël; Le Chatelier, Emmanuelle; Ehrlich, S Dusko; Lin, John C; Arumugam, Manimozhiyan; Wang, Jun; Madsen, Lise; Kristiansen, Karsten

    2015-10-01

    We established a catalog of the mouse gut metagenome comprising ∼2.6 million nonredundant genes by sequencing DNA from fecal samples of 184 mice. To secure high microbiome diversity, we used mouse strains of diverse genetic backgrounds, from different providers, kept in different housing laboratories and fed either a low-fat or high-fat diet. Similar to the human gut microbiome, >99% of the cataloged genes are bacterial. We identified 541 metagenomic species and defined a core set of 26 metagenomic species found in 95% of the mice. The mouse gut microbiome is functionally similar to its human counterpart, with 95.2% of its Kyoto Encyclopedia of Genes and Genomes (KEGG) orthologous groups in common. However, only 4.0% of the mouse gut microbial genes were shared (95% identity, 90% coverage) with those of the human gut microbiome. This catalog provides a useful reference for future studies.

  15. Mouse models for human otitis media

    PubMed Central

    Trune, Dennis R.; Zheng, Qing Yin

    2010-01-01

    Otitis media (OM) remains the most common childhood disease and its annual costs exceed $5 billion. Its potential for permanent hearing impairment also emphasizes the need to better understand and manage this disease. The pathogenesis of OM is multifactorial and includes infectious pathogens, anatomy, immunologic status, genetic predisposition, and environment. Recent progress in mouse model development is helping to elucidate the respective roles of these factors and to significantly contribute toward efforts of OM prevention and control. Genetic predisposition is recognized as an important factor in OM and increasing numbers of mouse models are helping to uncover the potential genetic bases for human OM. Furthermore, the completion of the mouse genome sequence has offered a powerful set of tools for investigating gene function and is generating a rich resource of mouse mutants for studying the genetic factors underlying OM. PMID:19272362

  16. Acute pharmacokinetics of memantine in the mouse.

    PubMed

    Saab, Bechara J; Roder, John C

    2011-01-01

    The pharmacokinetics of memantine, a widely prescribed medication in the United States and the European Union for the treatment of moderate-to-severe Alzheimer's disease (AD), have not been well explored in the mouse. Memantine is a highly unspecific blocker of many channels and how memantine may be of benefit in AD remains a mystery. Therefore, the investigation of memantine in the mouse, the most commonly chosen subject for modeling AD, has strong potential to lead to better therapies. Here, we present an acute pharmacokinetic analysis of memantine in mouse brain tissue and blood serum for a variety of experimentally relevant doses. The data help shed light on the mechanism of memantine action in vivo, and demonstrate that subcutaneous doses above 10 mg/kg in the mouse are most likely not therapeutically relevant to the human.

  17. Effects of endotoxin on the lactating mouse

    SciTech Connect

    Carr, J.K.

    1985-01-01

    The regulation of endogenous mouse mammary tumor virus (MMTV) sequences in trans by a host gene, the Lps locus on mouse chromosome 4, was suspected from a genetic linkage analysis. The Lps locus mediates the mouse's response to the injection of lipopolysaccharide (LPS) in the responder mouse while mice with the deficient allele are incapable of responding. Others have found that endotoxin exposure reduces milk production in lactating animals. This observation was confirmed in mice and extended by examining /sup 125/I-prolactin binding to liver membranes of lactating mice. Endotoxin treatment of responder mice increases liver prolactin binding within 15 minutes, followed by a decline over 6 hours. Scatchard analysis shows that the immediate increase comes from both increased affinity and abundance of the prolactin receptor. No such change in prolactin binding is seen in the non-responder following endotoxin treatment nor in /sup 125/I-insulin binding in responders.

  18. Features of adenosine metabolism of mouse heart.

    PubMed

    Deussen, Andreas; Weichsel, Johannes; Pexa, Annette

    2006-11-01

    Adenosine metabolism and transport were evaluated in the isolated perfused mouse heart and compared with the well-established model of isolated perfused guinea pig heart. Coronary venous release of adenosine under well-oxygenated conditions in the mouse exceeds that in the guinea pig threefold when related to tissue mass. Total myocardial adenosine production rate under this condition was approximately 2 nmol/min per gramme and similar in both species. Coronary resistance vessels of mice are highly sensitive to exogenous adenosine, and the threshold for adenosine-induced vasodilation is approximately 30 nmol/l. Adenosine membrane transport was largely insensitive to nitrobenzyl-thioinosine (NBTI) in mouse heart, which is in contrast to guinea pig and several other species. This indicates the dominance of NBTI-insensitive transporters in mouse heart. For future studies, the assessment of cytosolic and extracellular adenosine metabolism and its relationship with coronary flow will require the use of more effective membrane transport blockers.

  19. Melatonin receptors: latest insights from mouse models

    PubMed Central

    Tosini, Gianluca; Owino, Sharon; Guillame, Jean-Luc; Jockers, Ralf

    2014-01-01

    Summary Melatonin, the neuro-hormone synthesized during the night, has recently seen an unexpected extension of its functional implications towards type 2 diabetes development, visual functions, sleep disturbances and depression. Transgenic mouse models were instrumental for the establishment of the link between melatonin and these major human diseases. Most of the actions of melatonin are mediated by two types of G protein-coupled receptors, named MT1 and MT2, which are expressed in many different organs and tissues. Understanding the pharmacology and function of mouse MT1 and MT2 receptors, including MT1/MT2 heteromers, will be of crucial importance to evaluate the relevance of these mouse models for future therapeutic developments. This review will critically discuss these aspects, and give some perspectives including the generation of new mouse models. PMID:24903552

  20. A brain-specific gene cluster isolated from the region of the mouse obesity locus is expressed in the adult hypothalamus and during mouse development

    SciTech Connect

    Laig-Webster, M.; Lim, M.E.; Chehab, F.F.

    1994-09-01

    The molecular defect underlying an autosomal recessive form of genetic obesity in a classical mouse model C57 BL/6J-ob/ob has not yet been elucidated. Whereas metabolic and physiological disturbances such as diabetes and hypertension are associated with obesity, the site of expression and the nature of the primary lesion responsible for this cascade of events remains elusive. Our efforts aimed at the positional cloning of the ob gene by YAC contig mapping and gene identification have resulted in the cloning of a brain-specific gene cluster from the ob critical region. The expression of this gene cluster is remarkably complex owing to the multitude of brain-specific mRNA transcripts detected on Northern blots. cDNA cloning of these transcripts suggests that they are expressed from different genes as well as by alternate splicing mechanisms. Furthermore, the genomic organization of the cluster appears to consist of at least two identical promoters displaying CpG islands characteristic of housekeeping genes, yet clearly involving tissue-specific expression. Sense and anti-sense synthetic RNA probes were derived from a common DNA sequence on 3 cDNA clones and hybridized to 8-16 days mouse embryonic stages and mouse adult brain sections. Expression in development was noticeable as of the 11th day of gestation and confined to the central nervous system mainly in the telencephalon and spinal cord. Coronal and sagittal sections of the adult mouse brain showed expression only in 3 different regions of the brain stem. In situ hybridization to mouse hypothalamus sections revealed the presence of a localized and specialized group of cells expressing high levels of mRNA, suggesting that this gene cluster may also be involved in the regulation of hypothalamic activities. The hypothalamus has long been hypothesized as a primary candidate tissue for the expression of the obesity gene mainly because of its well-established role in the regulation of energy metabolism and food intake.

  1. Adrenomedullin increases the short-circuit current in the mouse seminal vesicle: actions on chloride secretion.

    PubMed

    Liao, S B; Cheung, K H; O, W S; Tang, Fai

    2014-08-01

    Adrenomedullin (ADM) may regulate seminal vesicle fluid secretion, and this may affect sperm quality. In this study, we investigated the effect of ADM on chloride secretion in the mouse seminal vesicle. The presence of ADM in mouse seminal vesicle was confirmed using immunostaining, and the molecular species was determined using gel filtration chromatography coupled with enzyme-linked assay for ADM. The effects of ADM on chloride secretion were studied by short-circuit current technique in a whole-mount preparation of mouse seminal vesicle in an Ussing chamber. The effects of specific ADM and calcitonin gene-related peptide (CGRP) receptor antagonists were investigated. Whether the ADM effect depended on the cAMP- and/or calcium-activated chloride channel was also studied using specific chloride channel blockers. The results showed that ADM was present in seminal vesicle epithelial cells. The major molecular species was precursor in the mouse seminal vesicle. ADM increased short-circuit current through the calcium-activated chloride channel in mouse seminal vesicle, and CGRP receptor was involved. We conclude that ADM may regulate chloride and fluid secretion from the seminal vesicle, which may affect the composition of the seminal plasma bathing the sperm and, hence, fertility.

  2. Activation of farnesoid X receptor induces RECK expression in mouse liver

    SciTech Connect

    Peng, Xiaomin; Wu, Weibin; Zhu, Bo; Sun, Zhichao; Ji, Lingling; Ruan, Yuanyuan; Zhou, Meiling; Zhou, Lei; Gu, Jianxin

    2014-01-03

    Highlights: •RECK is a novel transcriptional target gene of FXR in mouse liver. •The FXR response element is located within the intron 1 of RECK gene. •FXR agonist reverses the down-regulation of RECK in the liver in mouse NASH model. -- Abstract: Farnesoid X receptor (FXR) belongs to the ligand-activated nuclear receptor superfamily, and functions as a transcription factor regulating the transcription of numerous genes involved in bile acid homeostasis, lipoprotein and glucose metabolism. In the present study, we identified RECK, a membrane-anchored inhibitor of matrix metalloproteinases, as a novel target gene of FXR in mouse liver. We found that FXR agonist substantially augmented hepatic RECK mRNA and protein expression in vivo and in vitro. FXR regulated the transcription of RECK through directly binding to FXR response element located within intron 1 of the mouse RECK gene. Moreover, FXR agonist reversed the down-regulation of RECK in the livers from mice fed a methionine and choline deficient diet. In summary, our data suggest that RECK is a novel transcriptional target of FXR in mouse liver, and provide clues to better understanding the function of FXR in liver.

  3. Mapping the mouse brain with rs-fMRI: An optimized pipeline for functional network identification.

    PubMed

    Zerbi, Valerio; Grandjean, Joanes; Rudin, Markus; Wenderoth, Nicole

    2015-12-01

    The use of resting state fMRI (rs-fMRI) in translational research is a powerful tool to assess brain connectivity and investigate neuropathology in mouse models. However, despite encouraging initial results, the characterization of consistent and robust resting state networks in mice remains a methodological challenge. One key reason is that the quality of the measured MR signal is degraded by the presence of structural noise from non-neural sources. Notably, in the current pipeline of the Human Connectome Project, a novel approach has been introduced to clean rs-fMRI data, which involves automatic artifact component classification and data cleaning (FIX). FIX does not require any external recordings of physiology or the segmentation of CSF and white matter. In this study, we evaluated the performance of FIX for analyzing mouse rs-fMRI data. Our results showed that FIX can be easily applied to mouse datasets and detects true signals with 100% accuracy and true noise components with very high accuracy (>98%), thus reducing both within- and between-subject variability of rs-fMRI connectivity measurements. Using this improved pre-processing pipeline, maps of 23 resting state circuits in mice were identified including two networks that displayed default mode network-like topography. Hierarchical clustering grouped these neural networks into meaningful larger functional circuits. These mouse resting state networks, which are publicly available, might serve as a reference for future work using mouse models of neurological disorders.

  4. Human and Mouse Mononuclear Phagocyte Networks: A Tale of Two Species?

    PubMed Central

    Reynolds, Gary; Haniffa, Muzlifah

    2015-01-01

    Dendritic cells (DCs), monocytes, and macrophages are a heterogeneous population of mononuclear phagocytes that are involved in antigen processing and presentation to initiate and regulate immune responses to pathogens, vaccines, tumor, and tolerance to self. In addition to their afferent sentinel function, DCs and macrophages are also critical as effectors and coordinators of inflammation and homeostasis in peripheral tissues. Harnessing DCs and macrophages for therapeutic purposes has major implications for infectious disease, vaccination, transplantation, tolerance induction, inflammation, and cancer immunotherapy. There has been a paradigm shift in our understanding of the developmental origin and function of the cellular constituents of the mononuclear phagocyte system. Significant progress has been made in tandem in both human and mouse mononuclear phagocyte biology. This progress has been accelerated by comparative biology analysis between mouse and human, which has proved to be an exceptionally fruitful strategy to harmonize findings across species. Such analyses have provided unexpected insights and facilitated productive reciprocal and iterative processes to inform our understanding of human and mouse mononuclear phagocytes. In this review, we discuss the strategies, power, and utility of comparative biology approaches to integrate recent advances in human and mouse mononuclear phagocyte biology and its potential to drive forward clinical translation of this knowledge. We also present a functional framework on the parallel organization of human and mouse mononuclear phagocyte networks. PMID:26124761

  5. Families Get Involved! Learning Partners.

    ERIC Educational Resources Information Center

    Office of Educational Research and Improvement (ED), Washington, DC. Media and Information Services.

    Noting that families who are involved in their children's education make a difference in their child's performance, this two-page information sheet encourages families to get involved by listing the benefits of family involvement on one side and the ways adult family members can help in the school on the other. As a result of family participation:…

  6. Parent Involvement: Barriers and Opportunities.

    ERIC Educational Resources Information Center

    Mannan, Golam; Blackwell, Jacqueline

    1992-01-01

    Explores issues of parent involvement in light of current educational reform movements, asserts that parent involvement as a voluntary effort may not be effective, and argues that businesses and industries interested in reform are not focusing sufficiently on work-related variables that might encourage involvement of parents and other adults. (SLD)

  7. Measuring Involvement with Social Issues.

    ERIC Educational Resources Information Center

    Nowak, Glen J.; Salmon, Charles T.

    A study applied research concepts from consumer product involvement to test a model for research on involvement with social issues. Issue involvement was defined as the state or level of perceived importance and/or interest evoked by a stimulus (issue) within a specific situation. Attitudes on four social issues--abortion, pornography, the…

  8. Youth Maltreatment and Gang Involvement.

    ERIC Educational Resources Information Center

    Thompson, Kevin M.; Braaten-Antrim, Rhonda

    1998-01-01

    Examines whether physical and sexual maltreatment raises the risk of gang involvement among secondary school students. Findings show that maltreatment increases the probability of gang involvement, independent of demographic factors. When youth are physically and sexually abused their odds of gang involvement are four times higher than those who…

  9. Parental Involvement in High Schools.

    ERIC Educational Resources Information Center

    Brian, Donna JG

    Although parental involvement is recommended at all levels of schooling, involvement of parents at the secondary level has not been well defined in the literature. This paper presents findings of a case study that examined three high schools with varying levels of parental involvement--the first, a large high school with a predominantly working…

  10. Mouse Models of Diabetic Neuropathy

    PubMed Central

    Sullivan, Kelli A.; Hayes, John M.; Wiggin, Timothy D.; Backus, Carey; Oh, Sang Su; Lentz, Stephen I.; Brosius, Frank; Feldman, Eva L.

    2007-01-01

    Diabetic neuropathy (DN) is a debilitating complication of type 1 and type 2 diabetes. Rodent models of DN do not fully replicate the pathology observed in human patients. We examined DN in streptozotocin (STZ)-induced [B6] and spontaneous type 1 diabetes [B6Ins2Akita] and spontaneous type 2 diabetes [B6-db/db, BKS-db/db]. DN was defined using the criteria of the Animal Models of Diabetic Complications Consortium (http://www.amdcc.org). Despite persistent hyperglycemia, the STZ-treated B6 and B6Ins2Akita mice were resistant to the development of DN. In contrast, DN developed in both type 2 diabetes models: the B6-db/db and BKS-db/db mice. The persistence of hyperglycemia and development of DN in the B6-db/db mice required an increased fat diet while the BKS-db/db mice developed severe DN and remained hyperglycemic on standard mouse chow. Our data support the hypothesis that genetic background and diet influence the development of DN and should be considered when developing new models of DN. PMID:17804249

  11. Ethical Considerations in Mouse Experiments.

    PubMed

    Baertschi, Bernard; Gyger, Marcel

    2011-01-01

    Mice count morally because they can be harmed. This raises a moral issue in animal experimentation. Three main ethical attitudes towards animals are reviewed here. The Kantian view denies moral value to animals because they lack reason. The second view, by Singer, considers animals as sentient creatures (i.e., able to suffer). Finally, Regan considers that animals are subjects of their own life; they are autonomous and therefore have moral rights. Singer is a reformist and allows animal experimentation under certain conditions. Regan is abolitionist, saying that animals have moral rights that cannot be negotiated. Current animal protection legislation strives to put in balance the human and animal interests to decide whether an animal experiment is morally justified or not. An ethical evaluation process is conducted based on the harm-benefit assessment of the experiment. The researcher has to implement the 3Rs (Replacement, Reduction, Refinement) to minimize the harms to the animals and make sure that the outcomes are scientifically significant and that the quality of the science is high, in order to maximize benefits to humans and animals. Curr. Protoc. Mouse Biol. 1:155-167. © 2011 by John Wiley & Sons, Inc. PMID:26068990

  12. Expanded conserved linkage group between human 16p13 and the Scid region of the mouse chromosome 16

    SciTech Connect

    Deng, Z.M.; Siciliano, M.J.; Davisson, M.T.

    1994-09-01

    Knowledge of homologies between human and mouse chromosomes is essential for understanding chromosomal evolution and the development of experimental models for human disease. We have reported the identification of a conserved linkage group between human 16p13 and the centromeric portion of the mouse 16. Defining the extent of this linkage conservation has significant biomedical implications since that region of mouse genome contains the Scid mutation and the human 16p13 contains genes that are involved in DNA repair and certain types of human leukemia as well as other diseases such as Rubinstein-Taybi Syndrome. Here, this conserved linkage group has been defined and expanded. It now contains 5 genetic loci and spans more than 3 Mb in human and 23 cM in mouse. The 5 loci are PRM1,2 (protamine 1 and 2), NOP3 (a subclone of D16S237), GSPT1 (a gene involved in the regulation of G1 to S phase transition), MYH11 (a human smooth muscle myosin heavy chain gene) and MRP (multi-drug resistant-associated protein gene). Using a panel of human-rodent hybrids that are informative for different portions of human 16, we have established the following order on human 16p: telomere-NOP3-PRM1,2-GSPT1-(MYH11,MRP)-centromere. The genes were assigned to the mouse chromosome 16 by a mouse-Chinese hamster somatic cell hybrid panel informative for mouse chromosomes. Linkage analysis using backcross mice informative for the Scid mutation indicated the following order and genetic distance (in cM) in mouse: centromere-Nop3-11.7-Prm1-1.4-Gspt1-8.2-(Myh11,Mrp)-1.4-Scid-telomere.

  13. Oncogenic Radiation Abscopal Effects In Vivo: Interrogating Mouse Skin

    SciTech Connect

    Mancuso, Mariateresa; Leonardi, Simona; Giardullo, Paola; Pasquali, Emanuela; Tanori, Mirella; De Stefano, Ilaria; Casciati, Arianna; Naus, Christian C.; Pazzaglia, Simonetta; Saran, Anna

    2013-08-01

    Purpose: To investigate the tissue dependence in transmission of abscopal radiation signals and their oncogenic consequences in a radiosensitive mouse model and to explore the involvement of gap junction intercellular communication (GJIC) in mediating radiation tumorigenesis in off-target mouse skin. Methods and Materials: Patched1 heterozygous (Ptch1{sup +/−}) mice were irradiated at postnatal day 2 (P2) with 10 Gy of x-rays. Individual lead cylinders were used to protect the anterior two-thirds of the body, whereas the hindmost part was directly exposed to radiation. To test the role of GJICs and their major constituent connexin43 (Cx43), crosses between Ptch1{sup +/−} and Cx43{sup +/−} mice were similarly irradiated. These mouse groups were monitored for their lifetime, and skin basal cell carcinomas (BCCs) were counted and recorded. Early responses to DNA damage - Double Strand Breaks (DSBs) and apoptosis - were also evaluated in shielded and directly irradiated skin areas. Results: We report abscopal tumor induction in the shielded skin of Ptch1{sup +/−} mice after partial-body irradiation. Endpoints were induction of early nodular BCC-like tumors and macroscopic infiltrative BCCs. Abscopal tumorigenesis was significantly modulated by Cx43 status, namely, Cx43 reduction was associated with decreased levels of DNA damage and oncogenesis in out-of-field skin, suggesting a key role of GJIC in transmission of oncogenic radiation signals to unhit skin. Conclusions: Our results further characterize the nature of abscopal responses and the implications they have on pathologic processes in different tissues, including their possible underlying mechanistic bases.

  14. Cannabinoid ligand-receptor signaling in the mouse uterus.

    PubMed

    Das, S K; Paria, B C; Chakraborty, I; Dey, S K

    1995-05-01

    Using RNA (Northern) blot hybridization and reverse transcription-PCR, we demonstrate that the brain-type cannabinoid receptor (CB1-R) mRNA, but not the spleen-type cannabinoid receptor (CB2-R) mRNA, is expressed in the mouse uterus and that this organ has the capacity to synthesize the putative endogenous cannabinoid ligand, anandamide (arachidonylethanolamide). The psychoactive cannabinoid component of marijuana--delta 9-tetrahydrocannabinol (THC)--or anandamide, but not the inactive and nonpsychoactive cannabidiol (CBD), inhibited forskolin-stimulated cyclic AMP formation in the mouse uterus, which was prevented by pertussis toxin pretreatment. These results suggest that uterine CB1-R is coupled to inhibitory guanine nucleotide-binding protein and is biologically active. Autoradiographic studies identified ligand binding sites ([3H]anandamide) in the uterine epithelium and stromal cells, suggesting that these cells are perhaps the targets for cannabinoid action. Scatchard analysis of the binding of [3H]WIN 55212-2, another cannabinoid receptor ligand, showed a single class of high-affinity binding sites in the endometrium with an apparent Kd of 2.4 nM and Bmax of 5.4 x 10(9) molecules per mg of protein. The gene encoding lactoferrin is an estrogen-responsive gene in the mouse uterus that was rapidly and transiently up-regulated by THC, but not by CBD, in ovariectomized mice in the absence of ovarian steroids. This effect, unlike that of 17 beta-estradiol (E2), was not influenced by a pure antiestrogen, ICI 182780, suggesting that the THC-induced uterine lactoferrin gene expression does not involve estrogen receptors. We propose that the uterus is a new target for cannabinoid ligand-receptor signaling.

  15. Detailed characterization of the substrate specificity of mouse wax synthase.

    PubMed

    Miklaszewska, Magdalena; Kawiński, Adam; Banaś, Antoni

    2013-01-01

    Wax synthases are membrane-associated enzymes catalysing the esterification reaction between fatty acyl-CoA and a long chain fatty alcohol. In living organisms, wax esters function as storage materials or provide protection against harmful environmental influences. In industry, they are used as ingredients for the production of lubricants, pharmaceuticals, and cosmetics. Currently the biological sources of wax esters are limited to jojoba oil. In order to establish a large-scale production of desired wax esters in transgenic high-yielding oilseed plants, enzymes involved in wax esters synthesis from different biological resources should be characterized in detail taking into consideration their substrate specificity. Therefore, this study aims at determining the substrate specificity of one of such enzymes -- the mouse wax synthase. The gene encoding this enzyme was expressed heterologously in Saccharomyces cerevisiae. In the in vitro assays (using microsomal fraction from transgenic yeast), we evaluated the preferences of mouse wax synthase towards a set of combinations of 11 acyl-CoAs with 17 fatty alcohols. The highest activity was observed for 14:0-CoA, 12:0-CoA, and 16:0-CoA in combination with medium chain alcohols (up to 5.2, 3.4, and 3.3 nmol wax esters/min/mg microsomal protein, respectively). Unsaturated alcohols longer than 18°C were better utilized by the enzyme in comparison to the saturated ones. Combinations of all tested alcohols with 20:0-CoA, 22:1-CoA, or Ric-CoA were poorly utilized by the enzyme, and conjugated acyl-CoAs were not utilized at all. Apart from the wax synthase activity, mouse wax synthase also exhibited a very low acyl-CoA:diacylglycerol acyltransferase activity. However, it displayed neither acyl-CoA:monoacylglycerol acyltransferase, nor acyl-CoA:sterol acyltransferase activity.

  16. Cannabinoid ligand-receptor signaling in the mouse uterus.

    PubMed Central

    Das, S K; Paria, B C; Chakraborty, I; Dey, S K

    1995-01-01

    Using RNA (Northern) blot hybridization and reverse transcription-PCR, we demonstrate that the brain-type cannabinoid receptor (CB1-R) mRNA, but not the spleen-type cannabinoid receptor (CB2-R) mRNA, is expressed in the mouse uterus and that this organ has the capacity to synthesize the putative endogenous cannabinoid ligand, anandamide (arachidonylethanolamide). The psychoactive cannabinoid component of marijuana--delta 9-tetrahydrocannabinol (THC)--or anandamide, but not the inactive and nonpsychoactive cannabidiol (CBD), inhibited forskolin-stimulated cyclic AMP formation in the mouse uterus, which was prevented by pertussis toxin pretreatment. These results suggest that uterine CB1-R is coupled to inhibitory guanine nucleotide-binding protein and is biologically active. Autoradiographic studies identified ligand binding sites ([3H]anandamide) in the uterine epithelium and stromal cells, suggesting that these cells are perhaps the targets for cannabinoid action. Scatchard analysis of the binding of [3H]WIN 55212-2, another cannabinoid receptor ligand, showed a single class of high-affinity binding sites in the endometrium with an apparent Kd of 2.4 nM and Bmax of 5.4 x 10(9) molecules per mg of protein. The gene encoding lactoferrin is an estrogen-responsive gene in the mouse uterus that was rapidly and transiently up-regulated by THC, but not by CBD, in ovariectomized mice in the absence of ovarian steroids. This effect, unlike that of 17 beta-estradiol (E2), was not influenced by a pure antiestrogen, ICI 182780, suggesting that the THC-induced uterine lactoferrin gene expression does not involve estrogen receptors. We propose that the uterus is a new target for cannabinoid ligand-receptor signaling. Images Fig. 1 Fig. 3 Fig. 4 Fig. 5 PMID:7753807

  17. Orientation-tuned surround suppression in mouse visual cortex.

    PubMed

    Self, Matthew W; Lorteije, Jeannette A M; Vangeneugden, Joris; van Beest, Enny H; Grigore, Mihaela E; Levelt, Christiaan N; Heimel, J Alexander; Roelfsema, Pieter R

    2014-07-01

    The firing rates of neurons in primary visual cortex (V1) are suppressed by large stimuli, an effect known as surround suppression. In cats and monkeys, the strength of suppression is sensitive to orientation; responses to regions containing uniform orientations are more suppressed than those containing orientation contrast. This effect is thought to be important for scene segmentation, but the underlying neural mechanisms are poorly understood. We asked whether it is possible to study these mechanisms in the visual cortex of mice, because of recent advances in technology for studying the cortical circuitry in mice. It is unknown whether neurons in mouse V1 are sensitive to orientation contrast. We measured the orientation selectivity of surround suppression in the different layers of mouse V1. We found strong surround suppression in layer 4 and the superficial layers, part of which was orientation tuned: iso-oriented surrounds caused more suppression than cross-oriented surrounds. Surround suppression was delayed relative to the visual response and orientation-tuned suppression was delayed further, suggesting two separate suppressive mechanisms. Previous studies proposed that surround suppression depends on the activity of inhibitory somatostatin-positive interneurons in the superficial layers. To test the involvement of the superficial layers we topically applied lidocaine. Silencing of the superficial layers did not prevent orientation-tuned suppression in layer 4. These results show that neurons in mouse V1, which lacks orientation columns, show orientation-dependent surround suppression in layer 4 and the superficial layers and that surround suppression in layer 4 does not require contributions from neurons in the superficial layers.

  18. Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III

    SciTech Connect

    Han, Q.; Robinson, H; Cai, T; Tagle, D; Li, J

    2009-01-01

    Kynurenine aminotransferase III (KAT III) has been considered to be involved in the production of mammalian brain kynurenic acid (KYNA), which plays an important role in protecting neurons from overstimulation by excitatory neurotransmitters. The enzyme was identified based on its high sequence identity with mammalian KAT I, but its activity toward kynurenine and its structural characteristics have not been established. In this study, the biochemical and structural properties of mouse KAT III (mKAT III) were determined. Specifically, mKAT III cDNA was amplified from a mouse brain cDNA library, and its recombinant protein was expressed in an insect cell protein expression system. We established that mKAT III is able to efficiently catalyze the transamination of kynurenine to KYNA and has optimum activity at relatively basic conditions of around pH 9.0 and at relatively high temperatures of 50 to 60C. In addition, mKAT III is active toward a number of other amino acids. Its activity toward kynurenine is significantly decreased in the presence of methionine, histidine, glutamine, leucine, cysteine, and 3-hydroxykynurenine. Through macromolecular crystallography, we determined the mKAT III crystal structure and its structures in complex with kynurenine and glutamine. Structural analysis revealed the overall architecture of mKAT III and its cofactor binding site and active center residues. This is the first report concerning the biochemical characteristics and crystal structures of KAT III enzymes and provides a basis toward understanding the overall physiological role of mammalian KAT III in vivo and insight into regulating the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  19. The pathophysiology of mitochondrial disease as modeled in the mouse

    PubMed Central

    Wallace, Douglas C.; Fan, WeiWei

    2009-01-01

    It is now clear that mitochondrial defects are associated with a plethora of clinical phenotypes in man and mouse. This is the result of the mitochondria's central role in energy production, reactive oxygen species (ROS) biology, and apoptosis, and because the mitochondrial genome consists of roughly 1500 genes distributed across the maternal mitochondrial DNA (mtDNA) and the Mendelian nuclear DNA (nDNA). While numerous pathogenic mutations in both mtDNA and nDNA mitochondrial genes have been identified in the past 21 years, the causal role of mitochondrial dysfunction in the common metabolic and degenerative diseases, cancer, and aging is still debated. However, the development of mice harboring mitochondrial gene mutations is permitting demonstration of the direct cause-and-effect relationship between mitochondrial dysfunction and disease. Mutations in nDNA-encoded mitochondrial genes involved in energy metabolism, antioxidant defenses, apoptosis via the mitochondrial permeability transition pore (mtPTP), mitochondrial fusion, and mtDNA biogenesis have already demonstrated the phenotypic importance of mitochondrial defects. These studies are being expanded by the recent development of procedures for introducing mtDNA mutations into the mouse. These studies are providing direct proof that mtDNA mutations are sufficient by themselves to generate major clinical phenotypes. As more different mtDNA types and mtDNA gene mutations are introduced into various mouse nDNA backgrounds, the potential functional role of mtDNA variation in permitting humans and mammals to adapt to different environments and in determining their predisposition to a wide array of diseases should be definitively demonstrated. PMID:19651984

  20. Sox2 and Jagged1 Expression in Normal and Drug-Damaged Adult Mouse Inner Ear

    PubMed Central

    Campbell, Sean; Taylor, Ruth R.; Forge, Andrew; Hume, Clifford R.

    2007-01-01

    Inner ear hair cells detect environmental signals associated with hearing, balance, and body orientation. In humans and other mammals, significant hair cell loss leads to irreversible hearing and balance deficits, whereas hair cell loss in nonmammalian vertebrates is repaired by the spontaneous generation of replacement hair cells. Research in mammalian hair cell regeneration is hampered by the lack of in vivo damage models for the adult mouse inner ear and the paucity of cell-type-specific markers for non-sensory cells within the sensory receptor epithelia. The present study delineates a protocol to drug damage the adult mouse auditory epithelium (organ of Corti) in situ and uses this protocol to investigate Sox2 and Jagged1 expression in damaged inner ear sensory epithelia. In other tissues, the transcription factor Sox2 and a ligand member of the Notch signaling pathway, Jagged1, are involved in regenerative processes. Both are involved in early inner ear development and are expressed in developing support cells, but little is known about their expressions in the adult. We describe a nonsurgical technique for inducing hair cell damage in adult mouse organ of Corti by a single high-dose injection of the aminoglycoside kanamycin followed by a single injection of the loop diuretic furosemide. This drug combination causes the rapid death of outer hair cells throughout the cochlea. Using immunocytochemical techniques, Sox2 is shown to be expressed specifically in support cells in normal adult mouse inner ear and is not affected by drug damage. Sox2 is absent from auditory hair cells, but is expressed in a subset of vestibular hair cells. Double-labeling experiments with Sox2 and calbindin suggest Sox2-positive hair cells are Type II. Jagged1 is also expressed in support cells in the adult ear and is not affected by drug damage. Sox2 and Jagged1 may be involved in the maintenance of support cells in adult mouse inner ear. PMID:18157569

  1. Changes in gene expression associated with retinal degeneration in the rd3 mouse

    PubMed Central

    Cheng, Christiana L.

    2013-01-01

    Purpose To identify and characterize changes in gene expression associated with photoreceptor degeneration in the rd3 mouse model of Leber congenital amaurosis (LCA) type 12. Methods Global genome expression profiling using microarray technology was performed on total RNA extracts from rd3 and wild-type control mouse retinas at postnatal day 21. Quantitative PCR analysis of selected transcripts was performed to validate the microarray results. Results Functional annotation of differentially regulated genes in the rd3 mouse defined key canonical pathways, including phototransduction, glycerophospholipid metabolism, tumor necrosis factor receptor 1 signaling, and endothelin signaling. Overall, 1,140 of approximately 55,800 transcripts were differentially represented. In particular, a large percentage of the upregulated transcripts encode proteins involved in the immune response; whereas the downregulated transcripts encode proteins involved in phototransduction and lipid metabolism. Conclusions This analysis has elucidated several candidate genes and pathways, thus providing insight into the pathogenic mechanisms underlying the photoreceptor degeneration in the rd3 mouse retina and indicating directions for future studies. PMID:23687432

  2. An intron capture strategy used to identify and map a lysyl oxidase-like gene on chromosome 9 in the mouse

    SciTech Connect

    Wydner, K.S.; Passmore, H.C.; Kim, Houngho; Csiszar, K.; Boyd, C.D.

    1997-03-01

    An intron capture strategy involving use of polymerase chain reaction was used to identify and map the mouse homologue of a human lysyl oxidase-like gene (LOXL). Oligonucleotides complementary to conserved domains within exons 4 and 5 of the human lysyl oxidase-like gene were used to amplify the corresponding segment from mouse genomic DNA. Sequencing of the resulting mouse DNA fragment of approximately 1 kb revealed that the exon sequences at the ends of the amplified fragment are highly homologous (90% nucleotide identity) to exons 4 and 5 of the human lysyl oxidase-like gene. An AluI restriction site polymorphism within intron 4 was used to map the mouse lysyl oxidase-like gene (Loxl) to mouse Chromosome 9 in a region that shares linkage conservation with human chromosome 15q24, to which the LOXL was recently mapped. 22 refs., 3 figs.

  3. A false report of product tampering involving a rodent and soft drink can: light microscopy, image analysis and scanning electron microscopy/energy dispersive X-ray analysis.

    PubMed

    Platek, F; Ranieri, N; Wolnik, K A

    1997-11-01

    The "Pepsi Tamperings" of 1993 resulted in a large number of cases involving foreign objects reportedly found inside canned soft drinks. Although the majority of cases involved medical syringes and metallic objects, one case involved the report of a mouse found inside a can of Caffeine-Free Diet Pepsi. Using light and polarized light microscopy and computer-assisted image analysis, trace evidence and tooth structure from the suspect mouse were matched to scratches and indentions on the suspect can. Scanning electron microscopy and energy dispersive X-ray analysis were used to compare and match particles of gnawed metal from the lid of the suspect can to other particles recovered from the muzzle and stomach of the suspect mouse. The forensic analyses in this case proved the mouse could not have been canned in the soft drink product and refuted the defendant's sworn statements.

  4. A process-based review of mouse models of pulmonary hypertension

    PubMed Central

    Das, Mita; Fessel, Joshua; Tang, Haiyang; West, James

    2012-01-01

    Genetically modified mouse models have unparalleled power to determine the mechanisms behind different processes involved in the molecular and physiologic etiology of various classes of human pulmonary hypertension (PH). Processes known to be involved in PH for which there are extensive mouse models available include the following: (1) Regulation of vascular tone through secreted vasoactive factors; (2) regulation of vascular tone through potassium and calcium channels; (3) regulation of vascular remodeling through alteration in metabolic processes, either through alteration in substrate usage or through circulating factors; (4) spontaneous vascular remodeling either before or after development of elevated pulmonary pressures; and (5) models in which changes in tone and remodeling are primarily driven by inflammation. PH development in mice is of necessity faster and with different physiologic ramifications than found in human disease, and so mice make poor models of natural history of PH. However, transgenic mouse models are a perfect tool for studying the processes involved in pulmonary vascular function and disease, and can effectively be used to test interventions designed against particular molecular pathways and processes involved in disease. PMID:23372926

  5. A process-based review of mouse models of pulmonary hypertension.

    PubMed

    Das, Mita; Fessel, Joshua; Tang, Haiyang; West, James

    2012-10-01

    Genetically modified mouse models have unparalleled power to determine the mechanisms behind different processes involved in the molecular and physiologic etiology of various classes of human pulmonary hypertension (PH). Processes known to be involved in PH for which there are extensive mouse models available include the following: (1) Regulation of vascular tone through secreted vasoactive factors; (2) regulation of vascular tone through potassium and calcium channels; (3) regulation of vascular remodeling through alteration in metabolic processes, either through alteration in substrate usage or through circulating factors; (4) spontaneous vascular remodeling either before or after development of elevated pulmonary pressures; and (5) models in which changes in tone and remodeling are primarily driven by inflammation. PH development in mice is of necessity faster and with different physiologic ramifications than found in human disease, and so mice make poor models of natural history of PH. However, transgenic mouse models are a perfect tool for studying the processes involved in pulmonary vascular function and disease, and can effectively be used to test interventions designed against particular molecular pathways and processes involved in disease.

  6. Surfing the internet with a BCI mouse.

    PubMed

    Yu, Tianyou; Li, Yuanqing; Long, Jinyi; Gu, Zhenghui

    2012-06-01

    In this paper, we present a new web browser based on a two-dimensional (2D) brain-computer interface (BCI) mouse, where our major concern is the selection of an intended target in a multi-target web page. A real-world web page may contain tens or even hundreds of targets, including hyperlinks, input elements, buttons, etc. In this case, a target filter designed in our system can be used to exclude most of those targets of no interest. Specifically, the user filters the targets of no interest out by inputting keywords with a P300-based speller, while keeps those containing the keywords. Such filtering largely facilitates the target selection task based on our BCI mouse. When there are only several targets in a web page (either an original sparse page or a target-filtered page), the user moves the mouse toward the target of interest using his/her electroencephalographic signal. The horizontal movement and vertical movement are controlled by motor imagery and P300 potential, respectively. If the mouse encounters a target of no interest, the user rejects it and continues to move the mouse. Otherwise the user selects the target and activates it. With the collaboration of the target filtering and a series of mouse movements and target selections/rejections, the user can select an intended target in a web page. Based on our browser system, common navigation functions, including history rolling forward and backward, hyperlink selection, page scrolling, text input, etc, are available. The system has been tested on seven subjects. Experimental results not only validated the efficacy of the proposed method, but also showed that free internet surfing with a BCI mouse is feasible.

  7. Surfing the internet with a BCI mouse

    NASA Astrophysics Data System (ADS)

    Yu, Tianyou; Li, Yuanqing; Long, Jinyi; Gu, Zhenghui

    2012-06-01

    In this paper, we present a new web browser based on a two-dimensional (2D) brain-computer interface (BCI) mouse, where our major concern is the selection of an intended target in a multi-target web page. A real-world web page may contain tens or even hundreds of targets, including hyperlinks, input elements, buttons, etc. In this case, a target filter designed in our system can be used to exclude most of those targets of no interest. Specifically, the user filters the targets of no interest out by inputting keywords with a P300-based speller, while keeps those containing the keywords. Such filtering largely facilitates the target selection task based on our BCI mouse. When there are only several targets in a web page (either an original sparse page or a target-filtered page), the user moves the mouse toward the target of interest using his/her electroencephalographic signal. The horizontal movement and vertical movement are controlled by motor imagery and P300 potential, respectively. If the mouse encounters a target of no interest, the user rejects it and continues to move the mouse. Otherwise the user selects the target and activates it. With the collaboration of the target filtering and a series of mouse movements and target selections/rejections, the user can select an intended target in a web page. Based on our browser system, common navigation functions, including history rolling forward and backward, hyperlink selection, page scrolling, text input, etc, are available. The system has been tested on seven subjects. Experimental results not only validated the efficacy of the proposed method, but also showed that free internet surfing with a BCI mouse is feasible.

  8. Musculoskeletal involvement in systemic sclerosis.

    PubMed

    Randone, Silvia Bellando; Guiducci, Serena; Cerinic, Marco Matucci

    2008-04-01

    Musculoskeletal involvement is more frequent than expected in patients with systemic sclerosis (SSc) and is a major cause of disability, even if the prognosis of the disease largely depends on visceral involvement. The most common clinical feature of musculoskeletal involvement is arthralgia; less frequent features are arthritis, flexion contractures, stiffness (affecting predominantly fingers, wrists and ankles), proximal muscle weakness (mainly of the shoulder and hip) and tendon sheath involvement. Tendon friction rubs are predictive of poor prognosis. If musculoskeletal involvement is suspected, serum creatinine phosphokinase, aldolase, lactate dehydrogenase, alkaline phosphate, rheumatoid factor and anticyclic citrullinated peptide autoantibodies should be checked routinely. Treatment for muscle involvement has not yet been considered adequately and, in the future, it is to be hoped that clinical trials will identify new drugs to control this aspect of SSc, which seriously compromises patients' quality of life. PMID:18455689

  9. Gene expression and dental enamel structure in developing mouse incisor.

    PubMed

    Sehic, Amer; Risnes, Steinar; Khan, Qalb-E-Saleem; Khuu, Cuong; Osmundsen, Harald

    2010-04-01

    At the mouse incisor tip the initially differentiated ameloblasts produce a thin, prism-free enamel, while further apically, in the immediate adjacent segment, the enamel thickness increases and the four-layered enamel of mouse incisor is formed. Comparative gene-expression profiling was carried out on RNA isolated from these two segments of incisor tooth germs at embryonic day (E)17.5 and at postnatal days (P)0, 1, 2, and 10 using microarrays to measure messenger RNA (mRNA) and microRNA (miRNA) species present in the segments. Validation of expression data was achieved using real-time reverse transcription-polymerase chain reaction (RT-PCR) and western blotting. Bioinformatic data suggested enhanced cellular apoptosis in the incisal tip segment, which, together with diminished expression of the Amelx and Enam genes, may contribute to the production of the thin enamel seen in this tooth segment. For genes exhibiting higher levels of expression in the adjacent segment where complex enamel is being formed, bioinformatic analysis suggested significant associations with cellular functions involving the actin cytoskeleton, cellular development, morphology, and movement. This is suggested to reflect that ameloblasts with Tomes' process are being organized in transverse rows, facilitating the transverse movement that results in prism decussation in the inner enamel of the adjacent segment. Bioinformatic analysis of miRNA expression data lends support to these suggestions.

  10. In vitro effects of retinoic acid on mouse incisor development.

    PubMed

    Bloch-Zupan, A; Mark, M P; Weber, B; Ruch, J V

    1994-10-01

    The developing dentition is known to express the complete set of retinoic acid (RA) nuclear receptors and cytoplasmic RA-binding proteins (CRABPI and II), and RA is required for in vitro mouse molar morphogenesis, so the role of RA during in vitro mouse incisor development was investigated. Histological procedures, immunocytochemical detection of proliferating cells, immunofluorescence detection of laminin, and in situ hybridization with RNA probes for CRABPI and II were done on the tooth-germ cultures either in the presence or in the absence of RA. RA appeared to control initial morphogenesis, particularly the asymmetrical growth of the cervical loop, and to regulate required differential mitotic activity. RA seemed also to be involved in asymmetrical laminin deposition. The distribution of the CRABP gene transcripts was similar during in vivo and in vitro incisor development. However, CRABPI gene transcript distribution in the labial part of the epithelial loop was detected in vitro only in the presence of RA. A direct role of the CRABPs during tooth development is, however, unlikely because Ch55, a synthetic RA analogue that does not bind to CRABP, had the same effects as RA on in vitro incisor development. PMID:7741659

  11. Morphometric and ultrastructural changes with ageing in mouse peripheral nerve

    PubMed Central

    CEBALLOS, DOLORES; CUADRAS, JORDI; VERDÚ, ENRIQUE; NAVARRO, XAVIER

    1999-01-01

    Qualitative and quantitative information is reported on the morphological changes that occur in nerve fibres and nonneuronal cells of peripheral nerve during the lifetime of the mouse. Tibial nerves of mice aged 6–33 mo were studied. With ageing, collagen accumulates in the perineurium and lipid droplets in the perineurial cells. Macrophages and mast cells increase in number, and onion bulbs and collagen pockets are frequently present. Schwann cells associated with myelinated fibres (MF) slightly decrease in number in parallel with an increase of the internodal length from 6 to 12 mo, but increase in older nerves when demyelination and remyelination are common. The unmyelinated axon to myelinated fibre (UA/MF) ratio was about 2 until 12 mo, decreasing to 1.6 by 27 mo. In older mice, the loss of nerve fibres involves UA (50% loss of 27–33 mo cf. 6 mo) more markedly than MF (35%). In aged nerves wide incisures and infolded or outfolded myelin loops are frequent, resulting in an increased irregularity in the morphology of fibres along the internodes. In the mouse there is an adult time period, 12–20 mo, during which several features of degeneration progressively appear, and an ageing period from 20 mo upwards when the nerve suffers a general disorganisation and marked fibre loss. PMID:10634695

  12. Abnormal tracheal smooth muscle function in the CF mouse

    PubMed Central

    Wallace, Helen L; Southern, Kevin W; Connell, Marilyn G; Wray, Susan; Burdyga, Theodor

    2013-01-01

    Increased airway smooth muscle (ASM) contractility is thought to underlie symptoms of airway hyperresponsiveness (AHR). In the cystic fibrosis (CF) airway, ASM anomalies have been reported, but have not been fully characterized and the underlying mechanisms are largely unknown. We examined ASM in an adult CF mouse tracheal ring preparation, and determined whether changes in contractility were associated with altered ASM morphology. We looked for inherent changes in the cellular pathways involved in contractility, and characterized trachea morphology in the adult trachea and in an embryonic lung culture model during development. Results showed that that there was a reduction in tracheal caliber in CF mice as indicated by a reduction in the number of cartilage rings; proximal cross-sectional areas of cftr−/− tracheas and luminal areas were significantly smaller, but there was no difference in the area or distribution of smooth muscle. Morphological differences observed in adult trachea were not evident in the embryonic lung at 11.5 days gestation or after 72 h in culture. Functional data showed a significant reduction in the amplitude and duration of contraction in response to carbachol (CCh) in Ca-free conditions. The reduction in contraction was agonist specific, and occurred throughout the length of the trachea. These data show that there is a loss in the contractile capacity of the CF mouse trachea due to downregulation of the pathway specific to acetylcholine (ACh) activation. This reduction in contraction is not associated with changes in the area or distribution of ASM. PMID:24400140

  13. TOXICITY OF AMORPHOUS SILICA NANOPARTICLES IN MOUSE KERATINOCYTES

    SciTech Connect

    Yu, Kyung; Wang, Wei; Gu, Baohua; Hussain, Saber

    2009-01-01

    The present study was designed to examine the uptake, localization and the cytotoxic effects of well-dispersed amorphous silica nanoparticles in mouse keratinocytes (HEL-30). Mouse keratinocytes were exposed for 24h to various concentrations of amorphous silica nanoparticles in homogeneous suspensions of average size distribution (30, 48, 118 and 535 nm SiO2) then assessed for uptake and biochemical changes. Results of transmission electron microscopy revealed all sizes of silica were taken up into the cells and localized into the cytoplasm. The lactate dehydrogenase (LDH) assay shows LDH leakage was dose- and size-dependent with exposure to 30 and 48 nm nanoparticles. However, no LDH leakage was observed for either 118 or 535 nm nanoparticles. The mitochondrial viability assay (MTT) showed significant toxicity for 30 and 48 nm at high concentrations (100 g/mL) compare to the 118 and 535 nm particles. Further studies were carried out to investigate if cellular reduced GSH and mitochondria membrane potential are involved in the mechanism of SiO2 toxicity. The redox potential of cells (GSH) was reduced significantly at concentrations of 50, 100 and 200 g/mL at 30 nm nanoparticle exposures. However, silica nanoparticles larger than 30 nm showed no changes in GSH levels. Reactive oxygen species (ROS) formation did not show any significant change between controls and the exposed cells. In summary, amorphous silica nanoparticles below 100 nm induced cytotoxicity suggest size-of the particles is critical to produce biological effects.

  14. The biology of novel animal genes: Mouse APEX gene knockout

    SciTech Connect

    MacInnes, M.; Altherr, M.R.; Ludwig, D.; Pedersen, R.; Mold, C.

    1997-07-01

    This is the final report of a one-year, Laboratory Directed Research and Development (LDRD) project at the Los Alamos National Laboratory (LANL). The controlled breeding of novel genes into mice, including the gene knockout (KO), or conversely by adding back transgenes provide powerful genetic technologies that together suffice to determine in large part the biological role(s) of novel genes. Inbred mouse remains the best understood and most useful mammalian experimental system available for tackling the biology of novel genes. The major mammalian apurinic/apyrimidinic (AP) endonuclease (APE), is involved in a key step in the repair of spontaneous and induced AP sites in DNA. Efficient repair of these lesions is imperative to prevent the stable incorporation of mutations into the cellular genome which may lead to cell death or transformation. Loss or modulation of base excison repair activity in vivo may elevate the spontaneous mutation rate in cells, and may lead to a substantial increase in the incidence of cancer. Despite extensive biochemical analysis, however, the significance of these individual APE functions in vivo has not been elucidated. Mouse embryonic stem (ES) cells heterozygous for a deletion mutation in APE have been generated and whole animals containing the APE mutation have been derived from these ES cells. Animals homozygous for the APE null mutation die early in gestation, underscoring the biological significance of this DNA repair gene.

  15. Axin-1 Regulates Meiotic Spindle Organization in Mouse Oocytes

    PubMed Central

    Liu, Rui; Liu, Yu; Zhang, Fei; Zhang, Zhen; Shen, Yu-Ting; Xu, Lin; Chen, Ming-Huang; Wang, Ya-Long; Xu, Bai-Hui; Yang, Xiang-Jun; Wang, Hai-Long

    2016-01-01

    Axin-1, a negative regulator of Wnt signaling, is a versatile scaffold protein involved in centrosome separation and spindle assembly in mitosis, but its function in mammalian oogenesis remains unknown. Here we examined the localization and function of Axin-1 during meiotic maturation in mouse oocytes. Immunofluorescence analysis showed that Axin-1 was localized around the spindle. Knockdown of the Axin1 gene by microinjection of specific short interfering (si)RNA into the oocyte cytoplasm resulted in severely defective spindles, misaligned chromosomes, failure of first polar body (PB1) extrusion, and impaired pronuclear formation. However, supplementing the culture medium with the Wnt pathway activator LiCl improved spindle morphology and pronuclear formation. Downregulation of Axin1 gene expression also impaired the spindle pole localization of γ-tubulin/Nek9 and resulted in retention of the spindle assembly checkpoint protein BubR1 at kinetochores after 8.5 h of culture. Our results suggest that Axin-1 is critical for spindle organization and cell cycle progression during meiotic maturation in mouse oocytes. PMID:27284927

  16. Susceptibility of mouse macrophage J774 to dengue virus infection.

    PubMed

    Moreno-Altamirano, María M B; Sánchez-García, F Javier; Legorreta-Herrera, Martha; Aguilar-Carmona, Israel

    2007-01-01

    The aim of this study was to investigate whether the J774 mouse macrophage cell line could be used as an in vitro model for dengue virus infection (DENV). After 3 days, infection in J774 cells was assessed by detecting dengue virus non-structural protein 1 (NSP-1) production either by dot blot or indirect immunofluorescence assay (IFA) of saponine-permeabilized J774 cells and then confirmed by RT-PCR (171 bp product, corresponding to the DENV-2 core). Based on the presence of NSP-1 in infected but not in non-infected cells by both IFA and dot blot, as well as the amplification of a 171-bp DENV-2-specific RT-PCR product exclusively in the infected cells, the J774 cell line was found to be permissive for dengue virus infection. As far as we know, this is the first report that the J774 mouse macrophage cell line is infected with dengue virus and, thus, that it can be used as an alternative in vitro model for dengue virus infection studies. This finding could help to further elucidate the mechanisms involved in dengue virus infection and pathogenesis. PMID:17356302

  17. A mouse model for Chlamydia suis genital infection.

    PubMed

    Donati, Manuela; Di Paolo, Maria; Favaroni, Alison; Aldini, Rita; Di Francesco, Antonietta; Ostanello, Fabio; Biondi, Roberta; Cremonini, Eleonora; Ginocchietti, Laura; Cevenini, Roberto

    2015-02-01

    A mouse model for Chlamydia suis genital infection was developed. Ninety-nine mice were randomly divided into three groups and intravaginally inoculated with chlamydia: 45 mice (group 1) received C. suis purified elementary bodies (EBs), 27 (group 2) were inoculated with C. trachomatis genotype E EBs and 27 mice (group 3) with C. trachomatis genotype F EBs. Additionally, 10 mice were used as a negative control. At seven days post-infection (dpi) secretory anti-C. suis IgA were recovered from vaginal swabs of all C. suis inoculated mice. Chlamydia suis was isolated from 93, 84, 71 and 33% vaginal swabs at 3, 5, 7 and 12 dpi. Chlamydia trachomatis genotype E and F were isolated from 100% vaginal swabs up to 7 dpi and from 61 and 72%, respectively, at 12 dpi. Viable C. suis and C. trachomatis organisms were isolated from uterus and tubes up to 16 and 28 dpi, respectively. The results of the present study show the susceptibility of mice to intravaginal inoculation with C. suis. A more rapid course and resolution of C. suis infection, in comparison to C. trachomatis, was highlighted. The mouse model could be useful for comparative investigations involving C. suis and C. trachomatis species.

  18. Autoregulatory loop of Msx1 expression involving its antisense transcripts.

    PubMed

    Petit, Stéphane; Meary, Fleur; Pibouin, Laurence; Jeanny, Jean-Claude; Fernandes, Isabelle; Poliard, Anne; Hotton, Dominique; Berdal, Ariane; Babajko, Sylvie

    2009-08-01

    The Msx1 homeogene plays an important role in epithelial-mesenchymal interactions leading organogenesis. Msx1 gene is submitted to bidirectional transcription generating a long non-coding antisense (AS) RNA potentially involved in Msx1 expression regulation. RT-Q-PCR and RNA-FISH studies indicated that transient overexpression of the Msx1 AS transcript in 705IC5 mouse odontoblasts decreased the abundance of endogenous Msx1 S mRNA at the post-transcriptional level. Conversely, Msx1 overexpression increased the AS RNA level probably by activating AS transcription. In vivo mapping by RT-PCR evidenced both Msx1 RNAs in all adult mouse tissues tested raising the issue of Msx1 function during adulthood. The expression patterns of the two RNAs were similar, confirming the tight S/AS relationship. In particular, both Msx1 mRNAs and Msx1 protein were similarly distributed in eyes, and were found in regions with a common ectodermic origin and in cells potentially involved in regeneration. In conclusion, we report that Msx1 S RNA is negatively controlled by its AS RNA at a post-transcriptional level, and that the AS RNA is retrocontrolled positively by Msx1. The tight link between Msx1 S and AS RNAs constitutes a regulatory loop resulting in a fine-tuned expression of Msx1 which appears to be significant for adult homeostasis.

  19. The morphology of the mouse masticatory musculature

    PubMed Central

    Baverstock, Hester; Jeffery, Nathan S; Cobb, Samuel N

    2013-01-01

    The mouse has been the dominant model organism in studies on the development, genetics and evolution of the mammalian skull and associated soft-tissue for decades. There is the potential to take advantage of this well studied model and the range of mutant, knockin and knockout organisms with diverse craniofacial phenotypes to investigate the functional significance of variation and the role of mechanical forces on the development of the integrated craniofacial skeleton and musculature by using computational mechanical modelling methods (e.g. finite element and multibody dynamic modelling). Currently, there are no detailed published data of the mouse masticatory musculature available. Here, using a combination of micro-dissection and non-invasive segmentation of iodine-enhanced micro-computed tomography, we document the anatomy, architecture and proportions of the mouse masticatory muscles. We report on the superficial masseter (muscle, tendon and pars reflecta), deep masseter, zygomaticomandibularis (anterior, posterior, infraorbital and tendinous parts), temporalis (lateral and medial parts), external and internal pterygoid muscles. Additionally, we report a lateral expansion of the attachment of the temporalis onto the zygomatic arch, which may play a role in stabilising this bone during downwards loading. The data presented in this paper now provide a detailed reference for phenotypic comparison in mouse models and allow the mouse to be used as a model organism in biomechanical and functional modelling and simulation studies of the craniofacial skeleton and particularly the masticatory system. PMID:23692055

  20. Transgenic mouse model of cutaneous adnexal tumors

    PubMed Central

    Kito, Yusuke; Saigo, Chiemi; Atsushi, Kurabayashi; Mutsuo, Furihata; Tamotsu, Takeuchi

    2014-01-01

    TMEM207 was first characterized as being an important molecule for the invasion activity of gastric signet-ring cell carcinoma cells. In order to unravel the pathological properties of TMEM207, we generated several transgenic mouse lines, designated C57BL/6-Tg (ITF-TMEM207), in which murine TMEM207 was ectopically expressed under a truncated (by ~200 bp) proximal promoter of the murine intestinal trefoil factor (ITF) gene (also known as Tff3). Unexpectedly, a C57BL/6-Tg (ITF-TMEM207) mouse line exhibited a high incidence of spontaneous intradermal tumors with histopathological features that resembled those of various human cutaneous adnexal tumors. These tumors were found in ~14% female and 13% of male 6- to 12-month-old mice. TMEM207 immunoreactivity was found in hair follicle bulge cells in non-tumorous skin, as well as in cutaneous adnexal tumors of the transgenic mouse. The ITF-TMEM207 construct in this line appeared to be inserted to a major satellite repeat sequence at chromosome 2, in which no definite coding molecule was found. In addition, we also observed cutaneous adnexal tumors in three other C57BL/6-Tg (ITF-TMEM207) transgenic mouse lines. We believe that the C57BL/6-Tg (ITF-TMEM207) mouse might be a useful model to understand human cutaneous adnexal tumors. PMID:25305140

  1. Children computer mouse use and anthropometry.

    PubMed

    Hughes, Erin E; Johnson, Peter W

    2012-01-01

    Studies have shown that increased computer use among adults in occupational settings is associated with the development of cumulative trauma disorders; however, the need to address how adult-sized mice and keyboards are affecting children is becoming increasingly important as both access to and use of computers is increasing among today's youth. To address the potential mismatch that exists between child stature and computer input device size and activation force, we have applied existing, age-specific, anthropometric data to elements of device design, including mouse size (length, width, height, switch location), and mouse-button activation forces. Trends supported the development of smaller computer input devices with lower activation forces for smaller statured individuals including children. Distinct and consistent trends in size delineations were seen across gender and age groups-trends that correlate well with grades and schooling in the United States education system . Three to four mouse sizes would be recommended: a mouse sized for adult and high school males; one for adult and high school females and junior high males; one for elementary school children, aged 6 to 10 years; and possibly a mouse for the smallest users who are less than six years old. PMID:22316827

  2. Estrogen receptors in the wobbler mouse.

    PubMed

    Siegel, L I; Fox, T O

    1985-12-01

    Recent research has raised the interesting possibility that the neurological mutant mouse, wobbler (wr/wr), possesses an estrogen receptor deficit analogous to the androgen receptor deficiency found in androgen-resistant mice with testicular feminization. In the present report we examined estrogen-binding activity in cytosolic extracts of kidney, liver, and brain from wobbler mice, littermate control animals, and C57BL/6J mice, using DNA-cellulose chromatography. Estrogen binding components exhibiting properties of estrogen receptors were present in all tissues examined. Estrogen receptors adhered to DNA, displayed characteristic elution profiles from DNA-cellulose, and showed high affinity and limited capacity for estradiol, in contrast to non-receptor entities which bind estradiol. The qualitative elution patterns for estrogen receptors did not differ among groups within each tissue studied, and were similar to those reported previously in mouse kidney and brain. While estrogen receptors have been shown in mouse liver by other techniques, this is the first demonstration of putative estrogen receptors in mouse liver by DNA-cellulose chromatography. No consistent deficits in estrogen receptor concentration were found in wobblers compared to littermates. Thus, the data do not support the hypothesis that the wobbler mouse is an estrogen receptor-deficient mutant.

  3. EA-1, a novel adhesion molecule involved in the homing of progenitor T lymphocytes to the thymus

    PubMed Central

    1991-01-01

    The mouse progenitor T lymphocyte (pro-T) cell line FTF1 binds in vitro to thymus blood vessels, the thymic capsule, and liver from newborn mice. A mAb, EA-1, raised against an embryonic mouse endothelial cell line, blocked adhesion. The antibody also interfered with pro-T cell adhesion to a thymus-derived mouse endothelial cell line; it had no effect on the adhesion of mature T lymphocytes and myeloid cells. The antigen recognized by EA-1 is located on the vascular endothelium of various mouse tissues and absent on pro-T cells. EA-1 antibody precipitates molecules with apparent molecular weights of 110,000, 140,000, 160,000, and 200,000. Immunoclearing and binding-inhibition studies with antibodies against known adhesion molecules suggest that the EA-1 antigen is a novel adhesion molecule involved in colonization of the embryonic thymus by T cell progenitors. PMID:1874787

  4. Practical Application of Microelectroporation into Developing Mouse Brain

    NASA Astrophysics Data System (ADS)

    Shimogori, Tomomi; Ogawa, Masaharu

    One key approach toward understanding the genetic mechanisms underlying embryonic development involves the overexpression or misexpression of target genes in specific regions and at specific time points. The mouse gene-knockout system has been used extensively for loss-of-function studies due to the availability of a large number of mutant lines and the technical advantages of this system. In contrast, gain-of-function analyses have been performed through the production of knock-in and transgenic animals and with the use of various viruses (Cornetta 2006; Jakobsson et al., 2003; Hashimoto and Mikoshiba, 2004). However, it is not always possible to express or suppress genes in a spatially and temporally restricted manner, and the generation of genetically modified mice and recombinant viruses is time consuming and labor intensive. With the aim of solving these problems, many attempts have been made to apply the electroporation technique in research on developmental biology. Due to the accessibility of the avian embryo, it has been used as a classic model system for the study of developmental events in vertebrates. A novel technique for successful gene delivery into chick embryos has been established; this technique is known as in ovo electroporation and appears to be an excellent method, permitting quick and direct examination of the function of the delivered genes (Muramatsu et al., 1997; Itasaki et al., 1999; Momose et al., 1999; Nakamura et al., 2000; Yasuda et al., 2000). It seems that this technique can be adapted to the mouse embryo and would permit more rapid functional analysis of genes than is achieved by the generation of knockout or transgenic mouse lines. However, the inaccessibility of embryos in the mammalian uterus renders in utero manipulations targeting precise regions difficult or impossible at most stages of development. Efforts have been undertaken by various researchers to establish an in utero electroporation system, and there have been several

  5. Analysis of Kif5b Expression during Mouse Kidney Development

    PubMed Central

    Xue, Wenqian; Wang, Zai; Lu, Song; Lin, Raozhou; Liu, Mengfei; Zhu, Guixia; Huang, Jian-Dong

    2015-01-01

    Recent studies showed that kidney-specific inactivation of Kif3a produces kidney cysts and renal failure, suggesting that kinesin-mediated intracellular transportation is important for the establishement and maintenance of renal epithelial cell polarity and normal nephron functions. Kif5b, one of the most conserved kinesin heavy chain, is the mouse homologue of the human ubiquitous Kinesin Heavy Chain (uKHC). In order to elucidate the role of Kif5b in kidney development and function, it is essential to establish its expression profile within the organ. Therefore, in this study, we examined the expression pattern of Kif5b in mouse kidney. Kidneys from embryonic (E) 12.5-, 16.5-dpc (days post coitus) mouse fetuses, from postnatal (P) day 0, 10, 20 pups and from adult mice were collected. The distribution of Kif5b was analyzed by immunostaining. The possible involvement of Kif5b in kidney development was investigated in conditional mutant mice by using a Cre-LoxP strategy. This study showed that the distribution of Kif5b displayed spatiotemporal changes during postnatal kidney development. In kidneys of new born mice, Kif5b was strongly expressed in all developing tubules and in the ureteric bud, but not in the glomerulus or in other early-developing structures, such as the cap mesenchyme, the comma-shaped body, and the S-shaped body. In kidneys of postnatal day 20 or of older mice, however, Kif5b was localized selectively in the basolateral domain of epithelial cells of the thick ascending loop of Henle, as well as of the distal convoluted tubule, with little expression being observed in the proximal tubule or in the collecting duct. Conditional knock-down of Kif5b in mouse kidney did not result in detectable morphological defects, but it did lead to a decrease in cell proliferation rate and also to a mislocalization of Na+/K+/-ATPase, indicating that although Kif5b is non-essential for kidney morphogenesis, it is important for nephron maturation. PMID:25885434

  6. OAT3-mediated extrusion of the 99mTc-ECD metabolite in the mouse brain.

    PubMed

    Kikuchi, Tatsuya; Okamura, Toshimitsu; Wakizaka, Hidekatsu; Okada, Maki; Odaka, Kenichi; Yui, Joji; Tsuji, Atsushi B; Fukumura, Toshimitsu; Zhang, Ming-Rong

    2014-04-01

    After administration of the (99m)Tc complex with N,N'-1,2-ethylenediylbis-L-cysteine diethyl ester ((99m)Tc-ECD), a brain perfusion imaging agent, the radioactive metabolite is trapped in primate brain, but not in mouse and rat. Here, we investigate the involvement of metabolite extrusion by organic anion transporter 3 (OAT3), which is highly expressed at the blood-brain barrier in mice, in this species difference. The efflux rate of radioactivity in the cerebrum of Oat3(-/-) mice at later phase was 20% of that of control mice. Thus, organic anion transporters in mouse brain would be involved in the low brain retention of radioactivity after (99m)Tc-ECD administration.

  7. Mouse model of alloimmune-induced vascular rejection and transplant arteriosclerosis.

    PubMed

    Enns, Winnie; von Rossum, Anna; Choy, Jonathan

    2015-05-17

    Vascular rejection that leads to transplant arteriosclerosis (TA) is the leading representation of chronic heart transplant failure. In TA, the immune system of the recipient causes damage of the arterial wall and dysfunction of endothelial cells and smooth muscle cells. This triggers a pathological repair response that is characterized by intimal thickening and luminal occlusion. Understanding the mechanisms by which the immune system causes vasculature rejection and TA may inform the development of novel ways to manage graft failure. Here, we describe a mouse aortic interposition model that can be used to study the pathogenic mechanisms of vascular rejection and TA. The model involves grafting of an aortic segment from a donor animal into an allogeneic recipient. Rejection of the artery segment involves alloimmune reactions and results in arterial changes that resemble vascular rejection. The basic technical approach we describe can be used with different mouse strains and targeted interventions to answer specific questions related to vascular rejection and TA.

  8. Localization of a novel natural killer triggering receptor locus to human chromosome 3p23-p21 and mouse chromosome 9

    SciTech Connect

    Young, H.A.; Jenkins, N.A.; Copeland, N.G.; Simek, S.; Lerman, M.I.; Zbar, B.; Glenn, G.; Ortaldo, J.R.; Anderson, S.K.

    1993-05-01

    A novel gene (NKTR) that is involved in the recognition of tumor cells by large granular lymphocytes (LGLs) has been assigned to the short arm of human chromosome 3 in the region 3p23-p21 by somatic cell hybrid analysis. Interspecific backcross analysis revealed that the murine homologue maps to the distal end of mouse chromosome 9 and is closely linked to the locus coding for cholecystokinin (Cck). This region of mouse 9 shares a region of homology with human 3p. Thus, the placement of NKTR in these regions confirms and extends the relationship between these human and mouse chromosomes. 11 refs., 2 figs.

  9. Operant sensation seeking in the mouse.

    PubMed

    Olsen, Christopher M; Winder, Danny G

    2010-11-10

    Operant methods are powerful behavioral tools for the study of motivated behavior. These 'self-administration' methods have been used extensively in drug addiction research due to their high construct validity. Operant studies provide researchers a tool for preclinical investigation of several aspects of the addiction process. For example, mechanisms of acute reinforcement (both drug and non-drug) can be tested using pharmacological or genetic tools to determine the ability of a molecular target to influence self-administration behavior. Additionally, drug or food seeking behaviors can be studied in the absence of the primary reinforcer, and the ability of pharmacological compounds to disrupt this process is a preclinical model for discovery of molecular targets and compounds that may be useful for the treatment of addiction. One problem with performing intravenous drug self-administration studies in the mouse is the technical difficulty of maintaining catheter patency. Attrition rates in these experiments are high and can reach 40% or higher. Another general problem with drug self-administration is discerning which pharmacologically-induced effects of the reinforcer produce specific behaviors. For example, measurement of the reinforcing and neurological effects of psychostimulants can be confounded by their psychomotor effects. Operant methods using food reinforcement can avoid these pitfalls, although their utility in studying drug addiction is limited by the fact that some manipulations that alter drug self-administration have a minimal impact on food self-administration. For example, mesolimbic dopamine lesion or knockout of the D1 dopamine receptor reduce cocaine self-administration without having a significant impact on food self-administration. Sensory stimuli have been described for their ability to support operant responding as primary reinforcers (i.e. not conditioned reinforcers). Auditory and visual stimuli are self-administered by several species

  10. Parental Involvement and Academic Achievement

    ERIC Educational Resources Information Center

    Goodwin, Sarah Christine

    2015-01-01

    This research study examined the correlation between student achievement and parent's perceptions of their involvement in their child's schooling. Parent participants completed the Parent Involvement Project Parent Questionnaire. Results slightly indicated parents of students with higher level of achievement perceived less demand or invitations…

  11. Parental Involvement in the Classroom

    ERIC Educational Resources Information Center

    Machen, Sandra M.; Wilson, Janell D.; Notar, Charles E.

    2005-01-01

    Improving parental involvement with public schools can improve schools. Parental involvement is highly important for pushing the public school systems to higher standards. Also, research reports that engaging parents in an active role in the school curriculum can open alternative opportunities for children to succeed in academics. This report will…

  12. The Adolescent Drug Involvement Scale.

    ERIC Educational Resources Information Center

    Moberg, D. Paul; Hahn, Lori

    1991-01-01

    Developed Adolescent Drug Involvement Scale (ADIS) to measure level of drug involvement, considered as continuum ranging from no use to severe dependency, in adolescents. Administered ADIS to 453 adolescents referred for treatment. Results indicated acceptable internal consistency and provide preliminary evidence of validity. Scores correlated…

  13. Preparing Teachers for Parent Involvement.

    ERIC Educational Resources Information Center

    Safran, Daniel

    This paper examines the potential impact of parent involvement in the formal education of their children and suggests ways that teacher education can be restructured to prepare teachers to work with parents. This paper attempts to answer five questions: (1) Why should parents be involved in the formal education of their children? (2) Why should…

  14. New Directions in Parent Involvement.

    ERIC Educational Resources Information Center

    Fruchter, Norm; And Others

    This book presents findings of a study that identified and analyzed 18 recently developed programs or reform efforts in the United States that stress effective parental involvement. Chapter 1 provides a review of education literature and research on parent involvement from 1945 to 1985 and situates newly emerging efforts within the current climate…

  15. Teacher Involvement in Curriculum Development.

    ERIC Educational Resources Information Center

    Bowers, Bruce

    1991-01-01

    Four recent journal articles and one meeting paper on teacher involvement in curriculum development are summarized in this research bulletin. Contents include "Motivating Teacher Involvement in Professional Growth Activities," by Ruth Wright; "Teacher Participation in Curriculum Development: What Status Does It Have?" by Jean Young; "The Locus of…

  16. Citrobacter rodentium mouse model of bacterial infection.

    PubMed

    Crepin, Valerie F; Collins, James W; Habibzay, Maryam; Frankel, Gad

    2016-10-01

    Infection of mice with Citrobacter rodentium is a robust model to study bacterial pathogenesis, mucosal immunology, the health benefits of probiotics and the role of the microbiota during infection. C. rodentium was first isolated by Barthold from an outbreak of mouse diarrhea in Yale University in 1972 and was 'rediscovered' by Falkow and Schauer in 1993. Since then the use of the model has proliferated, and it is now the gold standard for studying virulence of the closely related human pathogens enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC, respectively). Here we provide a detailed protocol for various applications of the model, including bacterial growth, site-directed mutagenesis, mouse inoculation (from cultured cells and after cohabitation), monitoring of bacterial colonization, tissue extraction and analysis, immune responses, probiotic treatment and microbiota analysis. The main protocol, from mouse infection to clearance and analysis of tissues and host responses, takes ∼5 weeks to complete. PMID:27606775

  17. Peripheral Neuropathy in Mouse Models of Diabetes.

    PubMed

    Jolivalt, Corinne G; Frizzi, Katie E; Guernsey, Lucie; Marquez, Alex; Ochoa, Joseline; Rodriguez, Maria; Calcutt, Nigel A

    2016-01-01

    Peripheral neuropathy is a frequent complication of chronic diabetes that most commonly presents as a distal degenerative polyneuropathy with sensory loss. Around 20% to 30% of such patients may also experience neuropathic pain. The underlying pathogenic mechanisms are uncertain, and therapeutic options are limited. Rodent models of diabetes have been used for more than 40 years to study neuropathy and evaluate potential therapies. For much of this period, streptozotocin-diabetic rats were the model of choice. The emergence of new technologies that allow relatively cheap and routine manipulations of the mouse genome has prompted increased use of mouse models of diabetes to study neuropathy. In this article, we describe the commonly used mouse models of type 1 and type 2 diabetes, and provide protocols to phenotype the structural, functional, and behavioral indices of peripheral neuropathy, with a particular emphasis on assays pertinent to the human condition. © 2016 by John Wiley & Sons, Inc. PMID:27584552

  18. Mouse behavioural analysis in systems biology

    PubMed Central

    van Meer, Peter; Raber, Jacob

    2005-01-01

    Molecular techniques allowing in vivo modulation of gene expression have provided unique opportunities and challenges for behavioural studies aimed at understanding the function of particular genes or biological systems under physiological or pathological conditions. Although various animal models are available, the laboratory mouse (Mus musculus) has unique features and is therefore a preferred animal model. The mouse shares a remarkable genetic resemblance and aspects of behaviour with humans. In this review, first we describe common mouse models for behavioural analyses. As both genetic and environmental factors influence behavioural performance and need to be carefully evaluated in behavioural experiments, considerations for designing and interpretations of these experiments are subsequently discussed. Finally, common behavioural tests used to assess brain function are reviewed, and it is illustrated how behavioural tests are used to increase our understanding of the role of histaminergic neurotransmission in brain function. PMID:16035954

  19. FISH probes for mouse chromosome identification

    SciTech Connect

    Shi, Yu-Ping; Mohapatra, G.; Hanahan, D.; Miller, J.

    1997-10-01

    P1 clones near the telomeres and centromeres of each mouse chromosome except Y have been selected from a mouse genomic library and mapped using fluorescence in situ hybridization (FISH). Each clone was selected to contain a genetically mapped polymorphic DNA sequence as close as possible to the centromere or telomere of a chromosome. The genetic distance from the various P1 clones to the most distal genetically mapped polymorphic sequence ranged from 0 for about half of the clones to 6.7 cM for the probe at the telomere of chromosome 14. The average distance to the most distal or proximal chromosome marker was 1.5 cM. The use of FISH with these probes for mouse chromosome identification during comparative genomic hybridization is illustrated. 17 refs., 2 figs., 2 tabs.

  20. OCT guided microinjections for mouse embryonic research

    NASA Astrophysics Data System (ADS)

    Larin, Kirill V.; Syed, Saba H.; Coughlin, Andrew J.; Wang, Shang; West, Jennifer L.; Dickinson, Mary E.; Larina, Irina V.

    2013-02-01

    Optical coherence tomography (OCT) is gaining popularity as live imaging tool for embryonic research in animal models. Recently we have demonstrated that OCT can be used for live imaging of cultured early mouse embryos (E7.5-E10) as well as later stage mouse embryos in utero (E12.5 to the end of gestation). Targeted delivery of signaling molecules, drugs, and cells is a powerful approach to study normal and abnormal development, and image guidance is highly important for such manipulations. Here we demonstrate that OCT can be used to guide microinjections of gold nanoshell suspensions in live mouse embryos. This approach can potentially be used for variety of applications such as guided injections of contrast agents, signaling molecules, pharmacological agents, cell transplantation and extraction, as well as other image-guided micromanipulations. Our studies also reveal novel potential for gold nanoshells in embryonic research.

  1. Bloomsbury report on mouse embryo phenotyping: recommendations from the IMPC workshop on embryonic lethal screening

    PubMed Central

    Adams, David; Baldock, Richard; Bhattacharya, Shoumo; Copp, Andrew J.; Dickinson, Mary; Greene, Nicholas D. E.; Henkelman, Mark; Justice, Monica; Mohun, Timothy; Murray, Stephen A.; Pauws, Erwin; Raess, Michael; Rossant, Janet; Weaver, Tom; West, David

    2013-01-01

    Identifying genes that are important for embryo development is a crucial first step towards understanding their many functions in driving the ordered growth, differentiation and organogenesis of embryos. It can also shed light on the origins of developmental disease and congenital abnormalities. Current international efforts to examine gene function in the mouse provide a unique opportunity to pinpoint genes that are involved in embryogenesis, owing to the emergence of embryonic lethal knockout mutants. Through internationally coordinated efforts, the International Knockout Mouse Consortium (IKMC) has generated a public resource of mouse knockout strains and, in April 2012, the International Mouse Phenotyping Consortium (IMPC), supported by the EU InfraCoMP programme, convened a workshop to discuss developing a phenotyping pipeline for the investigation of embryonic lethal knockout lines. This workshop brought together over 100 scientists, from 13 countries, who are working in the academic and commercial research sectors, including experts and opinion leaders in the fields of embryology, animal imaging, data capture, quality control and annotation, high-throughput mouse production, phenotyping, and reporter gene analysis. This article summarises the outcome of the workshop, including (1) the vital scientific importance of phenotyping embryonic lethal mouse strains for basic and translational research; (2) a common framework to harmonise international efforts within this context; (3) the types of phenotyping that are likely to be most appropriate for systematic use, with a focus on 3D embryo imaging; (4) the importance of centralising data in a standardised form to facilitate data mining; and (5) the development of online tools to allow open access to and dissemination of the phenotyping data. PMID:23519032

  2. A Novel Mouse Model of Diffuse Intrinsic Pontine Glioma Initiated in Pax3-Expressing Cells12

    PubMed Central

    Misuraca, Katherine L.; Hu, Guo; Barton, Kelly L.; Chung, Alexander; Becher, Oren J.

    2016-01-01

    Diffuse intrinsic pontine glioma (DIPG) is a rare and incurable brain tumor that arises predominately in children and involves the pons, a structure that along with the midbrain and medulla makes up the brainstem. We have previously developed genetically engineered mouse models of brainstem glioma using the RCAS/Tv-a system by targeting PDGF-B overexpression, p53 loss, and H3.3K27M mutation to Nestin-expressing brainstem progenitor cells of the neonatal mouse. Here we describe a novel mouse model targeting these same genetic alterations to Pax3-expressing cells, which in the neonatal mouse pons consist of a Pax3 +/Nestin +/Sox2 + population lining the fourth ventricle and a Pax3 +/NeuN + parenchymal population. Injection of RCAS-PDGF-B into the brainstem of Pax3-Tv-a mice at postnatal day 3 results in 40% of mice developing asymptomatic low-grade glioma. A mixture of low- and high-grade glioma results from injection of Pax3-Tv-a;p53fl/fl mice with RCAS-PDGF-B and RCAS-Cre, with or without RCAS-H3.3K27M. These tumors are Ki67 +, Nestin +, Olig2 +, and largely GFAP − and can arise anywhere within the brainstem, including the classic DIPG location of the ventral pons. Expression of the H3.3K27M mutation reduces overall H3K27me3 as compared with tumors without the mutation, similar to what has been previously shown in human and mouse tumors. Thus, we have generated a novel genetically engineered mouse model of DIPG, which faithfully recapitulates the human disease and represents a novel platform with which to study the biology and treatment of this deadly disease. PMID:26806352

  3. Generation of a neuro-specific microarray reveals novel differentially expressed noncoding RNAs in mouse models for neurodegenerative diseases.

    PubMed

    Gstir, Ronald; Schafferer, Simon; Scheideler, Marcel; Misslinger, Matthias; Griehl, Matthias; Daschil, Nina; Humpel, Christian; Obermair, Gerald J; Schmuckermair, Claudia; Striessnig, Joerg; Flucher, Bernhard E; Hüttenhofer, Alexander

    2014-12-01

    We have generated a novel, neuro-specific ncRNA microarray, covering 1472 ncRNA species, to investigate their expression in different mouse models for central nervous system diseases. Thereby, we analyzed ncRNA expression in two mouse models with impaired calcium channel activity, implicated in Epilepsy or Parkinson's disease, respectively, as well as in a mouse model mimicking pathophysiological aspects of Alzheimer's disease. We identified well over a hundred differentially expressed ncRNAs, either from known classes of ncRNAs, such as miRNAs or snoRNAs or which represented entirely novel ncRNA species. Several differentially expressed ncRNAs in the calcium channel mouse models were assigned as miRNAs and target genes involved in calcium signaling, thus suggesting feedback regulation of miRNAs by calcium signaling. In the Alzheimer mouse model, we identified two snoRNAs, whose expression was deregulated prior to amyloid plaque formation. Interestingly, the presence of snoRNAs could be detected in cerebral spine fluid samples in humans, thus potentially serving as early diagnostic markers for Alzheimer's disease. In addition to known ncRNAs species, we also identified 63 differentially expressed, entirely novel ncRNA candidates, located in intronic or intergenic regions of the mouse genome, genomic locations, which previously have been shown to harbor the majority of functional ncRNAs.

  4. Generation of a neuro-specific microarray reveals novel differentially expressed noncoding RNAs in mouse models for neurodegenerative diseases

    PubMed Central

    Gstir, Ronald; Schafferer, Simon; Scheideler, Marcel; Misslinger, Matthias; Griehl, Matthias; Daschil, Nina; Humpel, Christian; Obermair, Gerald J.; Schmuckermair, Claudia; Striessnig, Joerg; Flucher, Bernhard E.

    2014-01-01

    We have generated a novel, neuro-specific ncRNA microarray, covering 1472 ncRNA species, to investigate their expression in different mouse models for central nervous system diseases. Thereby, we analyzed ncRNA expression in two mouse models with impaired calcium channel activity, implicated in Epilepsy or Parkinson's disease, respectively, as well as in a mouse model mimicking pathophysiological aspects of Alzheimer's disease. We identified well over a hundred differentially expressed ncRNAs, either from known classes of ncRNAs, such as miRNAs or snoRNAs or which represented entirely novel ncRNA species. Several differentially expressed ncRNAs in the calcium channel mouse models were assigned as miRNAs and target genes involved in calcium signaling, thus suggesting feedback regulation of miRNAs by calcium signaling. In the Alzheimer mouse model, we identified two snoRNAs, whose expression was deregulated prior to amyloid plaque formation. Interestingly, the presence of snoRNAs could be detected in cerebral spine fluid samples in humans, thus potentially serving as early diagnostic markers for Alzheimer's disease. In addition to known ncRNAs species, we also identified 63 differentially expressed, entirely novel ncRNA candidates, located in intronic or intergenic regions of the mouse genome, genomic locations, which previously have been shown to harbor the majority of functional ncRNAs. PMID:25344396

  5. cDNA sequence and mapping of the mouse Copb gene encoding the beta subunit of the COPI coatomer complex.

    PubMed

    LI, W; Elliott, R W; Novak, E K; Swank, R T

    1999-05-01

    COPI-coated vesicles are involved in retrograde-directed selective transport of proteins from the Golgi complex to the endoplasmic reticulum (ER) as well as mediate anterograde transport of cargo proteins within the Golgi or in endosomal trafficking. The COPI protein complex contains an ADP-ribosylation factor (ARF1) and seven coatamer subunits (alpha, beta, beta', gamma, delta, epsilon, zeta-COP). The localization and function of human beta subunit of coatamer (COPB) suggests it is likely a candidate gene of ruby-eye-2 (ru2), which is a mouse model of human Hermansky-Pudlak syndrome characterized by the dysfunction of several subcellular organelles. In this study, we determined the entire coding sequence of mouse (Copb) cDNA by combining an overlapping mouse EST contig with EST walking. beta-COP was found highly conserved in mouse, rat, and human, and it is ubiquitously expressed in mouse. The Copb gene was mapped to mouse Chr 7 at a position of 53.3 cM by radiation hybrid mapping. Our RH mapping data, sequencing of RT-PCR products, and Western blotting exclude the Copb gene as a candidate for ru2.

  6. Employee involvement: motivation or manipulation?

    PubMed

    McConnell, C R

    1998-03-01

    Employee involvement is subject to a great deal of verbal tribute; there is hardly a manager at work today who will not praise the value of employee input. However, many employee involvement efforts leave employees feeling more manipulated than motivated. This occurs because supervisors and managers, while expecting employees to change the way they work, are themselves either unwilling to change or remain unconscious of the need to change. The result is that, although employee input is regularly solicited in a number of forms, it is often discounted, ignored, or altered to fit the manager's preconceptions. Often the employee is left feeling manipulated. Since the opportunity for involvement can be a strong motivator, it becomes the manager's task to learn how to provide involvement opportunity in manipulative fashion. This can be accomplished by providing involvement opportunity accompanied by clear outcome expectations and allowing employees the freedom to pursue those outcomes in their own way.

  7. An anaphylaxis inhibitory factor present in mouse serum.

    PubMed

    Astorquiza, M I; Droguett, M A

    1996-01-01

    We have previously described that normal mouse serum inhibits the PCA reaction mediated by IgE. The present study attempts to characterize this PCA inhibitory factor from the biologically active fraction of the serum. The physicochemical properties of this glycoprotein are the following: it is inactivated at 55 degrees C; it has a molecular weight between 182 kD and 240 kD, determined by gel filtration; it shows affinity to concanavalin A and lentil lectin but not to peanut agglutin; it demonstrates affinity to IgE and, apparently, its carbohydrate moiety is not required for its biological activity. Two bands corresponding to 64.5 kD and 48. 1 kD, which are likely to constitute the biologically active molecule, are observed by SDS-PAGE. These properties are different from those found in factors with IgE affinity involved in IgE synthesis.

  8. Protoporphyrin IX binding and transport by recombinant mouse PBR.

    PubMed

    Wendler, Gregor; Lindemann, Peter; Lacapère, Jean-Jacques; Papadopoulos, Vassilios

    2003-11-28

    The mitochondrial 18kDa peripheral-type benzodiazepine receptor (PBR), a high affinity cholesterol binding protein, has been shown to interact with protoporphyrin IX (PPIX) and this interaction was assumed to be involved in the regulation of heme biosynthesis and porphyrin-based photodynamic therapy in cancer. In order to test this hypothesis recombinant mouse PBR was expressed in Escherichia coli. The recombinant gene product showed in E. coli protoplasts specific affinity for PPIX binding. PPIX could displace PK 11195 binding. Moreover, induced PBR protein expression in E. coli protoplasts caused an uptake of PPIX that could be completely inhibited by cholesterol and to a lesser extent inhibited by PK 11195 and Ro5-4864. These results suggest that PBR, in addition to its role in cholesterol and coproporphyrinogen III transport, is also directing the mitochondrial PPIX import, a function that can be ascribed to the 18kDa PBR protein alone.

  9. Thymic necrosis following oral inoculation of mouse thymic virus.

    PubMed

    Morse, S S

    1989-11-01

    Mouse thymic virus (MTLV;ICTV designation murid herpesvirus 3) infects developing T lymphocytes of neonatal mice, causing thymic necrosis and acute immunosuppression. Infected animals shed virus indefinitely. However, although transmission in nature is presumably by contact and is likely to involve the oral-nasal route, virtually all experimental studies with MTLV have used systemic (intraperitoneal) inoculation. In order to determine whether systemic inoculation causes artifacts in pathogenesis of the infection, effects of intraperitoneal and oral-nasal inoculation were compared in newborn mice. Thymic necrosis occurred with either route of inoculation, although rate of infection was lower with oral inoculation, varying from about 20% to 67%. There were no gross differences in pathogenesis. Orally infected animals seroconverted and shed virus. These data indicate that the apparent lymphotropism of thymic virus, and induction of thymic necrosis, are not dependent on route of inoculation.

  10. Targeted disruption of the mouse Lipoma Preferred Partner gene

    SciTech Connect

    Vervenne, Hilke B.V.K.; Crombez, Koen R.M.O.; Delvaux, Els L.; Janssens, Veerle; Ven, Wim J.M. van de Petit, Marleen M.R.

    2009-02-06

    LPP (Lipoma Preferred Partner) is a zyxin-related cell adhesion protein that is involved in the regulation of cell migration. We generated mice with a targeted disruption of the Lpp gene and analysed the importance of Lpp for embryonic development and adult functions. Aberrant Mendelian inheritance in heterozygous crosses suggested partial embryonic lethality of Lpp{sup -/-} females. Fertility of Lpp{sup -/-} males was proven to be normal, however, females from Lpp{sup -/-} x Lpp{sup -/-} crosses produced a strongly reduced number of offspring, probably due to a combination of female embryonic lethality and aberrant pregnancies. Apart from these developmental and reproductive abnormalities, Lpp{sup -/-} mice that were born reached adulthood without displaying any additional macroscopic defects. On the other hand, Lpp{sup -/-} mouse embryonic fibroblasts exhibited reduced migration capacity, reduced viability, and reduced expression of some Lpp interaction partners. Finally, we discovered a short nuclear form of Lpp, expressed mainly in testis via an alternative promoter.

  11. Serotonin competence of mouse beta cells during pregnancy.

    PubMed

    Goyvaerts, Lotte; Schraenen, Anica; Schuit, Frans

    2016-07-01

    Pregnancy is a key mammalian reproductive event in which growth and differentiation of the fetus imposes extra metabolic and hormonal demands on the mother. Its successful outcome depends on major changes in maternal blood circulation, metabolism and endocrine function. One example is the endocrine pancreas, where beta cells undergo a number of changes in pregnancy that result in enhanced functional beta cell mass in order to compensate for the rising metabolic needs for maternal insulin. During the last 5 years, a series of studies have increased our understanding of the molecular events involved in this functional adaptation. In the mouse, a prominent functional change during pregnancy is the capacity of some beta cells to produce serotonin. In this review we will discuss the mechanism and potential effects of pregnancy-related serotonin production in beta cells, considering functional consequences at the local intra-islet and systemic level. PMID:27056372

  12. Endpoints for Mouse Abdominal Tumor Models: Refinement of Current Criteria

    PubMed Central

    Paster, Eden V; Villines, Kimberly A; Hickman, Debra L

    2009-01-01

    Accurate, rapid, and noninvasive health assessments are required to establish more appropriate endpoints in mouse cancer models where tumor size is not easily measured. We evaluated potential endpoints in mice with experimentally induced peritoneal lymphoma, an abdominal tumor model, by comparing body weight, body condition, and behavior with those of a control group of mice not developing lymphoma. Our hypothesis was that body weight would increase or plateau, whereas body condition and behavioral scores would decrease, as disease progressed. Results indicated that body weight did not differ significantly between the control and experimental groups, but the experimental group experienced significant decreases in both body condition and behavioral scores. Our results support the use of body condition and behavioral scoring as adjunctive assessment methods for mice involved in abdominal lymphoma tumor studies in which health may decline despite an increase or plateau in body weight. PMID:19619413

  13. Endoplasmic reticulum stress in mouse decidua during early pregnancy.

    PubMed

    Gu, Xiao-Wei; Yan, Jia-Qi; Dou, Hai-Ting; Liu, Jie; Liu, Li; Zhao, Meng-Long; Liang, Xiao-Huan; Yang, Zeng-Ming

    2016-10-15

    Unfolded or misfolded protein accumulation in the endoplasmic reticulum lumen leads to endoplasmic reticulum stress (ER stress). Although it is known that ER stress is crucial for mammalian reproduction, little is known about its physiological significance and underlying mechanism during decidualization. Here we show that Ire-Xbp1 signal transduction pathway of unfolded protein response (UPR) is activated in decidual cells. The process of decidualization is compromised by ER stress inhibitor tauroursodeoxycholic acid sodium (TUDCA) and Ire specific inhibitor STF-083010 both in vivo and in vitro. A high concentration of ER stress inducer tunicamycin (TM) suppresses stromal cells proliferation and decidualization, while a lower concentration is beneficial. We further show that ER stress induces DNA damage and polyploidization in stromal cells. In conclusion, our data suggest that the GRP78/Ire1/Xbp1 signaling pathway of ER stress-UPR is activated and involved in mouse decidualization.

  14. Modifiers of epigenetic reprogramming show paternal effects in the mouse

    PubMed Central

    Chong, Suyinn; Vickaryous, Nicola; Ashe, Alyson; Zamudio, Natasha; Youngson, Neil; Hemley, Sarah; Stopka, Tomas; Skoultchi, Arthur; Matthews, Jacqui; Scott, Hamish S; de Kretser, David; O’Bryan, Moira; Blewitt, Marnie; Whitelaw, Emma

    2011-01-01

    There is increasing evidence that epigenetic information can be inherited across generations in mammals, despite extensive reprogramming both in the gametes and in the early developing embryo. One corollary to this is that disrupting the establishment of epigenetic state in the gametes of a parent, as a result of heterozygosity for mutations in genes involved in reprogramming, could affect the phenotype of offspring that do not inherit the mutant allele. Here we show that such effects do occur following paternal inheritance in the mouse. We detected changes to transcription and chromosome ploidy in adult animals. Paternal effects of this type have not been reported previously in mammals and suggest that the untransmitted genotype of male parents can influence the phenotype of their offspring. PMID:17450140

  15. Noninvasive Imaging of Natural Killer Cell-Mediated Apoptosis in a Mouse Tumor Model.

    PubMed

    Singh, Thoudam Debraj; Lee, Jaetae; Jeon, Yong Hyun

    2016-01-01

    Natural killer (NK) cells are cytotoxic lymphocytes that induce apoptosis in cancer cells infected with viruses and bacteria through a caspase-3-dependent pathway. Effective NK cell-based immunotherapy requires highly sensitive imaging tools for in vivo monitoring of the dynamic events involved in apoptosis. Here, we describe a noninvasive bioluminescence imaging approach to determine the antitumor effects of NK cell-based therapy by serial imaging of caspase-3-dependent apoptosis in a mouse model of human glioma. PMID:27177676

  16. Sphingolipid metabolism in organotypic mouse keratinocyte cultures

    SciTech Connect

    Madison, K.C.; Swartzendruber, D.C.; Wertz, P.W.; Downing, D.T. )

    1990-12-01

    Ceramides are the dominant component of the stratum corneum intercellular lipid lamellae, which constitute the epidermal permeability barrier. Only pig and human epidermal ceramides have been extensively characterized and the structures of the ceramides of cultured keratinocytes have not been previously investigated. In the present studies, we have characterized the ceramides synthesized by organotypic lifted mouse keratinocyte cultures for the first time and compared them to the ceramides of intact mouse epidermis. Both mouse epidermis and cultures contained five ceramides, ceramide 1 being the least polar and ceramide 5 the most polar. Ceramide 1 was a group of acylceramides, i.e., very-long-chain omega-hydroxyceramides with an ester-linked nonhydroxy fatty acid. Ceramide 2 contained medium-length saturated nonhydroxy fatty acids. (In culture, the ceramide 2 band was split into two parts with the slightly more polar ceramide 2' containing short-chain saturated nonhydroxy fatty acids.) Ceramide 5 contained short-chain alpha-hydroxy fatty acids. The structures of ceramides 1, 2, and 5 were analagous to those of pig and human epidermis. Mouse epidermal ceramide 3 was quite unusual, containing beta-hydroxy fatty acids, a structure not previously identified among mammalian ceramides. In contrast, culture ceramide 3 was composed of omega-hydroxy fatty acids with a chain-length distribution similar to that of ceramide 1. Mouse ceramide 4 was composed of fatty acids with chromatographic mobility similar to hydroxy fatty acids but with different chemical reactivity; it remains only partially characterized. Culture ceramide 4 was present in quantities too small for analysis. All ceramides in mouse epidermis and cultures contained only sphingosine bases, whereas pig and human ceramides also contain phytosphingosine.

  17. Cancer mouse models: past, present and future.

    PubMed

    Khaled, Walid T; Liu, Pentao

    2014-03-01

    The development and advances in gene targeting technology over the past three decades has facilitated the generation of cancer mouse models that recapitulate features of human malignancies. These models have been and still remain instrumental in revealing the complexities of human cancer biology. However, they will need to evolve in the post-genomic era of cancer research. In this review we will highlight some of the key developments over the past decades and will discuss the new possibilities of cancer mouse models in the light of emerging powerful gene manipulating tools.

  18. Development and characterization of mouse hybridomas.

    PubMed

    Mechetner, Eugene

    2007-01-01

    Cell fusion protocols that were developed by Kohler and Milstein in the mid-1970s and aimed at producing and characterization of mouse monoclonal antibodies (MAbs) remain the gold standard of hybridoma development. Despite tremendous progress in using MAbs in multiple research, diagnostic, and therapeutic areas, major experimental flaws in designing and carrying out hybridoma experimentation often result in the production of hybridomas exhibiting poor growth parameters and secreting low-specificity and low-affinity antibodies. This methodology chapter is built around the conventional hybridoma protocol, with a special emphasis on tissue culture and biochemical techniques aimed at producing truly monospecific and highly active mouse MAbs.

  19. Islet Insulin Secretion Measurements in the Mouse.

    PubMed

    Hugill, Alison; Shimomura, Kenju; Cox, Roger D

    2016-01-01

    This article describes detailed protocols for in vitro measurements of insulin function and secretion in isolated mouse islets for the analysis of glucose homeostasis. We specify a method of enzyme digestion and hand picking to isolate and release the greatest number of high quality islets from the pancreas of the mouse. We describe an effective method for generating dynamic measurements of insulin secretion using a perifusion assay including a detailed protocol for constructing a peristaltic pump and tubing assembly. In addition we describe an alternative and simple technique for measuring insulin secretion using static incubation of isolated islets. © 2016 by John Wiley & Sons, Inc. PMID:27584553

  20. The Riken mouse genome encyclopedia project.

    PubMed

    Hayashizaki, Yoshihide

    2003-01-01

    The Riken mouse genome encyclopedia a comprehensive full-length cDNA collection and sequence database. High-level functional annotation is based on sequence homology search, expression profiling, mapping and protein-protein interactions. More than 1000000 clones prepared from 163 tissues were end-sequenced and classified into 128000 clusters, and 60000 representative clones were fully sequenced representing 24000 clear protein-encoding genes. The application of the mouse genome database for positional cloning and gene network regulation analysis is reported.

  1. Identification of Disease Specific Pathways Using in Vivo SILAC Proteomics in Dystrophin Deficient mdx Mouse*

    PubMed Central

    Rayavarapu, Sree; Coley, William; Cakir, Erdinc; Jahnke, Vanessa; Takeda, Shin'ichi; Aoki, Yoshitsugu; Grodish-Dressman, Heather; Jaiswal, Jyoti K.; Hoffman, Eric P.; Brown, Kristy J.; Hathout, Yetrib; Nagaraju, Kanneboyina

    2013-01-01

    Duchenne muscular dystrophy (DMD) is an X-linked neuromuscular disorder caused by a mutation in the dystrophin gene. DMD is characterized by progressive weakness of skeletal, cardiac, and respiratory muscles. The molecular mechanisms underlying dystrophy-associated muscle weakness and damage are not well understood. Quantitative proteomics techniques could help to identify disease-specific pathways. Recent advances in the in vivo labeling strategies such as stable isotope labeling in mouse (SILAC mouse) with 13C6-lysine or stable isotope labeling in mammals (SILAM) with 15N have enabled accurate quantitative analysis of the proteomes of whole organs and tissues as a function of disease. Here we describe the use of the SILAC mouse strategy to define the underlying pathological mechanisms in dystrophin-deficient skeletal muscle. Differential SILAC proteome profiling was performed on the gastrocnemius muscles of 3-week-old (early stage) dystrophin-deficient mdx mice and wild-type (normal) mice. The generated data were further confirmed in an independent set of mdx and normal mice using a SILAC spike-in strategy. A total of 789 proteins were quantified; of these, 73 were found to be significantly altered between mdx and normal mice (p < 0.05). Bioinformatics analyses using Ingenuity Pathway software established that the integrin-linked kinase pathway, actin cytoskeleton signaling, mitochondrial energy metabolism, and calcium homeostasis are the pathways initially affected in dystrophin-deficient muscle at early stages of pathogenesis. The key proteins involved in these pathways were validated by means of immunoblotting and immunohistochemistry in independent sets of mdx mice and in human DMD muscle biopsies. The specific involvement of these molecular networks early in dystrophic pathology makes them potential therapeutic targets. In sum, our findings indicate that SILAC mouse strategy has uncovered previously unidentified pathological pathways in mouse models of human

  2. Identification of disease specific pathways using in vivo SILAC proteomics in dystrophin deficient mdx mouse.

    PubMed

    Rayavarapu, Sree; Coley, William; Cakir, Erdinc; Jahnke, Vanessa; Takeda, Shin'ichi; Aoki, Yoshitsugu; Grodish-Dressman, Heather; Jaiswal, Jyoti K; Hoffman, Eric P; Brown, Kristy J; Hathout, Yetrib; Nagaraju, Kanneboyina

    2013-05-01

    Duchenne muscular dystrophy (DMD) is an X-linked neuromuscular disorder caused by a mutation in the dystrophin gene. DMD is characterized by progressive weakness of skeletal, cardiac, and respiratory muscles. The molecular mechanisms underlying dystrophy-associated muscle weakness and damage are not well understood. Quantitative proteomics techniques could help to identify disease-specific pathways. Recent advances in the in vivo labeling strategies such as stable isotope labeling in mouse (SILAC mouse) with (13)C6-lysine or stable isotope labeling in mammals (SILAM) with (15)N have enabled accurate quantitative analysis of the proteomes of whole organs and tissues as a function of disease. Here we describe the use of the SILAC mouse strategy to define the underlying pathological mechanisms in dystrophin-deficient skeletal muscle. Differential SILAC proteome profiling was performed on the gastrocnemius muscles of 3-week-old (early stage) dystrophin-deficient mdx mice and wild-type (normal) mice. The generated data were further confirmed in an independent set of mdx and normal mice using a SILAC spike-in strategy. A total of 789 proteins were quantified; of these, 73 were found to be significantly altered between mdx and normal mice (p < 0.05). Bioinformatics analyses using Ingenuity Pathway software established that the integrin-linked kinase pathway, actin cytoskeleton signaling, mitochondrial energy metabolism, and calcium homeostasis are the pathways initially affected in dystrophin-deficient muscle at early stages of pathogenesis. The key proteins involved in these pathways were validated by means of immunoblotting and immunohistochemistry in independent sets of mdx mice and in human DMD muscle biopsies. The specific involvement of these molecular networks early in dystrophic pathology makes them potential therapeutic targets. In sum, our findings indicate that SILAC mouse strategy has uncovered previously unidentified pathological pathways in mouse models of

  3. Epigenetic Basis of Regeneration: Analysis of Genomic DNA Methylation Profiles in the MRL/MpJ Mouse

    PubMed Central

    Górnikiewicz, Bartosz; Ronowicz, Anna; Podolak, Justyna; Madanecki, Piotr; Stanisławska-Sachadyn, Anna; Sachadyn, PaweŁ

    2013-01-01

    Epigenetic regulation plays essential role in cell differentiation and dedifferentiation, which are the intrinsic processes involved in regeneration. To investigate the epigenetic basis of regeneration capacity, we choose DNA methylation as one of the most important epigenetic mechanisms and the MRL/MpJ mouse as a model of mammalian regeneration known to exhibit enhanced regeneration response in different organs. We report the comparative analysis of genomic DNA methylation profiles of the MRL/MpJ and the control C57BL/6J mouse. Methylated DNA immunoprecipitation followed by microarray analysis using the Nimblegen ‘3 × 720 K CpG Island Plus RefSeq Promoter’ platform was applied in order to carry out genome-wide DNA methylation profiling covering 20 404 promoter regions. We identified hundreds of hypo- and hypermethylated genes and CpG islands in the heart, liver, and spleen, and 37 of them in the three tissues. Decreased inter-tissue diversification and the shift of DNA methylation balance upstream the genes distinguish the genomic methylation patterns of the MRL/MpJ mouse from the C57BL/6J. Homeobox genes and a number of other genes involved in embryonic morphogenesis are significantly overrepresented among the genes hypomethylated in the MRL/MpJ mouse. These findings indicate that epigenetic patterning might be a likely molecular basis of regeneration capability in the MRL/MpJ mouse. PMID:23929942

  4. Mouse models of liver fibrosis mimic human liver fibrosis of different etiologies

    PubMed Central

    Martínez, Allyson K.; Maroni, Luca; Marzioni, Marco; Ahmed, Syed T.; Milad, Mena; Ray, Debolina; Alpini, Gianfranco; Glaser, Shannon S.

    2014-01-01

    The liver has the amazing capacity to repair itself after injury; however, the same processes that are involved in liver regeneration after acute injury can cause serious consequences during chronic liver injury. In an effort to repair damage, activated hepatic stellate cells trigger a cascade of events that lead to deposition and accumulation of extracellular matrix components causing the progressive replacement of the liver parenchyma by scar tissue, thus resulting in fibrosis. Although fibrosis occurs as a result of many chronic liver diseases, the molecular mechanisms involved depend on the underlying etiology. Since studying liver fibrosis in human subjects is complicated by many factors, mouse models of liver fibrosis that mimic the human conditions fill this void. This review summarizes the general mouse models of liver fibrosis and mouse models that mimic specific human disease conditions that result in liver fibrosis. Additionally, recent progress that has been made in understanding the molecular mechanisms involved in the fibrogenic processes of each of the human disease conditions is highlighted. PMID:25396098

  5. Mouse Genome Database: From sequence to phenotypes and disease models.

    PubMed

    Eppig, Janan T; Richardson, Joel E; Kadin, James A; Smith, Cynthia L; Blake, Judith A; Bult, Carol J

    2015-08-01

    The Mouse Genome Database (MGD, www.informatics.jax.org) is the international scientific database for genetic, genomic, and biological data on the laboratory mouse to support the research requirements of the biomedical community. To accomplish this goal, MGD provides broad data coverage, serves as the authoritative standard for mouse nomenclature for genes, mutants, and strains, and curates and integrates many types of data from literature and electronic sources. Among the key data sets MGD supports are: the complete catalog of mouse genes and genome features, comparative homology data for mouse and vertebrate genes, the authoritative set of Gene Ontology (GO) annotations for mouse gene functions, a comprehensive catalog of mouse mutations and their phenotypes, and a curated compendium of mouse models of human diseases. Here, we describe the data acquisition process, specifics about MGD's key data areas, methods to access and query MGD data, and outreach and user help facilities. PMID:26150326

  6. Genes and translocations involved in POF.

    PubMed

    Schlessinger, David; Herrera, Luisa; Crisponi, Laura; Mumm, Steven; Percesepe, Antonio; Pellegrini, Massimo; Pilia, Giuseppe; Forabosco, Antonino

    2002-08-15

    Changes at a single autosomal locus and many X-linked loci have been implicated in women with gonadal dysgenesis [premature ovarian failure (POF) with deficits in ovarian follicles]. For the chromosome 3 locus, a forkhead transcription factor gene (FOXL2) has been identified, in which lesions result in decreased follicles by haploinsufficiency. In contrast, sporadic X; autosomal translocations are distributed at many points on the X, but concentrate in a critical region on Xq. The association of the breakpoints with genes involved in ovarian function is thus far weak (in four analyzed cases) and has not been related to pathology in other POF patients. While many more translocations can be analyzed in detail as the human genome sequence is refined, it remains possible that translocations like X monosomy (Turner syndrome) lead to POF not by interrupting specific genes important in ovarian development, but by causing aberrations in pairing or X-inactivation during folliculogenesis. It is noted that the critical region has unusual features, neighboring the X-inactivation center and including an 18 Mb region of very low recombination. These suggest that chromosome dynamics in the region may be sensitive to structural changes, and when modified by translocations might provoke apoptosis at meiotic checkpoints. Choices among models for the etiology of POF should be feasible based on studies of ovarian follicle development and attrition in mouse models. Studies would prominently include gene expression profiling of developmental-specific pathways in nascent ovaries with controlled levels of Foxl2 and interacting proteins, or with defined changes in the X chromosome.

  7. Pulmonary involvement in rheumatoid arthritis.

    PubMed

    Bilgici, Ayhan; Ulusoy, H; Kuru, O; Celenk, C; Unsal, M; Danaci, M

    2005-08-01

    The primary objective of this investigation was to assess the relationships between clinical characteristics, lung involvement, and frequency of pulmonary involvement in rheumatoid arthritis (RA). Using high-resolution computed tomography (HRCT) and pulmonary function tests (PFT), we prospectively evaluated 52 patients with RA (eight males and 44 females, mean age 53.6 years). The HRCT was abnormal in 35 patients (67.3%), the most frequent abnormalities being reticulonodular patterns, which were found in 22 patients (62.9%), ground-glass attenuation (20%), and bronchiectasis (17%). In this group of patients, PFT results were normal in 13 patients (37%). Titers of rheumatoid factor and erythrocyte sedimentation rate were significantly higher in abnormal HRCT presence. Higher Larsen's score, advanced age, and severe disease were significant risk factors for lung involvement (p<0.001, p<0.01, and p<0.01, respectively) and are suggested by our data to be statistically significant predictors of lung involvement in RA.

  8. The susceptibility of the hamster to mouse encephalomyelitis virus.

    PubMed

    DEAN, D J; DALLDORF, G

    1948-12-01

    The OT strain of mouse encephalomyelitis virus induces an inapparent infection in suckling hamsters associated with lesions of the central nervous system and skeletal muscles. The virus increases in pathogenicity after alternating mouse-hamster transfers and then induces both paralysis and encephalitis. Pathogenicity is lost through serial hamster passages but is restored by a single mouse transfer.

  9. 9 CFR 113.33 - Mouse safety tests.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Mouse safety tests. 113.33 Section 113... Procedures § 113.33 Mouse safety tests. One of the mouse safety tests provided in this section shall be... or more ingredients makes the biological product lethal or toxic for mice but not lethal or toxic...

  10. 9 CFR 113.33 - Mouse safety tests.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Mouse safety tests. 113.33 Section 113... Procedures § 113.33 Mouse safety tests. One of the mouse safety tests provided in this section shall be... or more ingredients makes the biological product lethal or toxic for mice but not lethal or toxic...

  11. Mouse Driven Window Graphics for Network Teaching.

    ERIC Educational Resources Information Center

    Makinson, G. J.; And Others

    Computer enhanced teaching of computational mathematics on a network system driving graphics terminals is being redeveloped for a mouse-driven, high resolution, windowed environment of a UNIX work station. Preservation of the features of networked access by heterogeneous terminals is provided by the use of the X Window environment. A dmonstrator…

  12. Somatic Cell Nuclear Transfer in the Mouse

    NASA Astrophysics Data System (ADS)

    Kishigami, Satoshi; Wakayama, Teruhiko

    Somatic cell nuclear transfer (SCNT) has become a unique and powerful tool for epigenetic reprogramming research and gene manipulation in animals since “Dolly,” the first animal cloned from an adult cell was reported in 1997. Although the success rates of somatic cloning have been inefficient and the mechanism of reprogramming is still largely unknown, this technique has been proven to work in more than 10 mammalian species. Among them, the mouse provides the best model for both basic and applied research of somatic cloning because of its abounding genetic resources, rapid sexual maturity and propagation, minimal requirements for housing, etc. This chapter describes a basic protocol for mouse cloning using cumulus cells, the most popular cell type for NT, in which donor nuclei are directly injected into the oocyte using a piezo-actuated micromanipulator. In particular, we focus on a new, more efficient mouse cloning protocol using trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, which increases both in vitro and in vivo developmental rates from twofold to fivefold. This new method including TSA will be helpful to establish mouse cloning in many laboratories.

  13. Large-scale mouse knockouts and phenotypes.

    PubMed

    Ramírez-Solis, Ramiro; Ryder, Edward; Houghton, Richard; White, Jacqueline K; Bottomley, Joanna

    2012-01-01

    Standardized phenotypic analysis of mutant forms of every gene in the mouse genome will provide fundamental insights into mammalian gene function and advance human and animal health. The availability of the human and mouse genome sequences, the development of embryonic stem cell mutagenesis technology, the standardization of phenotypic analysis pipelines, and the paradigm-shifting industrialization of these processes have made this a realistic and achievable goal. The size of this enterprise will require global coordination to ensure economies of scale in both the generation and primary phenotypic analysis of the mutant strains, and to minimize unnecessary duplication of effort. To provide more depth to the functional annotation of the genome, effective mechanisms will also need to be developed to disseminate the information and resources produced to the wider community. Better models of disease, potential new drug targets with novel mechanisms of action, and completely unsuspected genotype-phenotype relationships covering broad aspects of biology will become apparent. To reach these goals, solutions to challenges in mouse production and distribution, as well as development of novel, ever more powerful phenotypic analysis modalities will be necessary. It is a challenging and exciting time to work in mouse genetics.

  14. Translating Mouse Vocalizations: Prosody and Frequency Modulation

    PubMed Central

    Lahvis, Garet P.; Alleva, Enrico; Scattoni, Maria Luisa

    2010-01-01

    Mental illness can include impaired abilities to express emotions or respond to the emotions of others. Speech provides a mechanism for expressing emotions, by both what words are spoken and by the melody or intonation of speech (prosody). Through the perception of variations in prosody, an individual can detect changes in another's emotional state. Prosodic features of mouse ultrasonic vocalizations (USVs), indicated by changes in frequency and amplitude, also convey information. Dams retrieve pups that emit separation calls, females approach males emitting solicitous calls, and mice can become fearful of a cue associated with the vocalizations of a distressed conspecific. Since acoustic features of mouse USVs respond to drugs and genetic manipulations that influence reward circuits, USV analysis can be employed to examine how genes influence social motivation, affect regulation, and communication. The purpose of this review is to discuss how genetic and developmental factors influence aspects of the mouse vocal repertoire and how mice respond to the vocalizations of their conspecifics. To generate falsifiable hypotheses about the emotional content of particular calls, this review addresses USV analysis within the framework of affective neuroscience (e.g. measures of motivated behavior such as conditioned place preference tests, brain activity, and systemic physiology). Suggested future studies include employment of an expanded array of physiological and statistical approaches to identify the salient acoustic features of mouse vocalizations. We are particularly interested in rearing environments that incorporate sufficient spatial and temporal complexity to familiarize developing mice with a broader array of affective states. PMID:20497235

  15. Having Fun with a Cordless Mouse

    ERIC Educational Resources Information Center

    Nunn, John

    2016-01-01

    A cordless mouse with an added reed switch is used as a wireless data logger to record every time the wheel of a trolley completes a revolution. The limitations of the system in terms of maximum clicking rate and spatial resolution are considered and data obtained from the descent of a trolley down a ramp at various different angles is analysed in…

  16. MMP responses in the MRL mouse

    PubMed Central

    Heber-Katz, Ellen; Gourevitch, Dmitri

    2013-01-01

    The matrix metalloproteinases (MMPs) have been implicated the regenerative response in amphibians and various mammalian models of regeneration. The neutrophil response is known to bring MMPs and other proteases to the wound to promote bacterial elimination and tissue remodeling. These issues in relation to what is occurring in the MRL mouse model of regeneration/wound healing is discussed. PMID:19735244

  17. Immunohistochemistry of Paraffin Sections from Mouse Ovaries.

    PubMed

    Akkoyunlu, Gokhan; Tepekoy, Filiz

    2016-01-01

    Immunohistochemistry (IHC) is an efficient technique to detect cellular localizations of the proteins in paraffin-embedded tissues. It allows specific proteins to be visualized by the interaction of antibodies with an enzyme-substrate-chromogen system. Here, we describe indirect immunohistochemistry method for paraffin-embedded mouse ovaries fixed with Bouin's Fixative. PMID:27557588

  18. Inhibitor(s) of the classical complement pathway in mouse serum limit the utility of mice as experimental models of neuromyelitis optica.

    PubMed

    Ratelade, Julien; Verkman, A S

    2014-11-01

    Neuromyelitis optica (NMO) is an inflammatory demyelinating disease of the central nervous system in which anti-aquaporin-4 (AQP4) autoantibodies (AQP4-IgG) cause damage to astrocytes by complement-dependent cytotoxicity (CDC). Various approaches have been attempted to produce NMO lesions in rodents, some involving genetically modified mice with altered immune cell function. Here, we found that mouse serum strongly inhibits complement from multiple species, preventing AQP4-IgG-dependent CDC. Effects of mouse serum on complement activation were tested in CDC assays in which AQP4-expressing cells were incubated with AQP4-IgG and complement from different species. Biochemical assays and mass spectrometry were used to characterize complement inhibitor(s) in mouse serum. Sera from different strains of mice produced almost no AQP4-IgG-dependent CDC compared with human, rat and guinea pig sera. Remarkably, addition of mouse serum prevented AQP4-IgG-dependent CDC caused by human, rat or guinea pig serum, with 50% inhibition at <5% mouse serum. Hemolysis assays indicated that the inhibitor(s) in mouse serum target the classical and not the alternative complement pathway. We found that the complement inhibitor(s) in mouse serum were contained in a serum fraction purified with protein-A resin; however, the inhibitor was not IgG as determined using serum from IgG-deficient mice. Mass spectrometry on the protein A-purified fraction produced several inhibitor candidates. The low intrinsic complement activity of mouse serum and the presence of complement inhibitor(s) limit the utility of mouse models to study disorders, such as NMO, involving the classical complement pathway.

  19. Inhibitor(s) of the classical complement pathway in mouse serum limit the utility of mice as experimental models of neuromyelitis optica

    PubMed Central

    Ratelade, Julien; Verkman, A.S.

    2015-01-01

    Neuromyelitis optica (NMO) is an inflammatory demyelinating disease of the central nervous system in which anti-aquaporin-4 (AQP4) autoantibodies (AQP4-IgG) cause damage to astrocytes by complement-dependent cytotoxicity (CDC). Various approaches have been attempted to produce NMO lesions in rodents, some involving genetically modified mice with altered immune cell function. Here, we found that mouse serum strongly inhibits complement from multiple species, preventing AQP4-IgG-dependent CDC. Effects of mouse serum on complement activation were tested in CDC assays in which AQP4-expressing cells were incubated with AQP4-IgG and complement from different species. Biochemical assays and mass spectrometry were used to characterize complement inhibitor(s) in mouse serum. Sera from different strains of mice produced almost no AQP4-IgG-dependent CDC compared with human, rat and guinea pig sera. Remarkably, addition of mouse serum prevented AQP4-IgG-dependent CDC caused by human, rat or guinea pig serum, with 50% inhibition at <5% mouse serum. Hemolysis assays indicated that the inhibitor(s) in mouse serum target the classical and not the alternative complement pathway. We found that the complement inhibitor(s) in mouse serum were contained in a serum fraction purified with protein-A resin; however, the inhibitor was not IgG as determined using serum from IgG-deficient mice. Mass spectrometry on the protein A-purified fraction produced several inhibitor candidates. The low intrinsic complement activity of mouse serum and the presence of complement inhibitor(s) limit the utility of mouse models to study disorders, such as NMO, involving the classical complement pathway. PMID:24980869

  20. Different cysteine proteinases involved in bone resorption and osteoclast formation.

    PubMed

    Brage, M; Abrahamson, M; Lindström, V; Grubb, A; Lerner, U H

    2005-06-01

    Cysteine proteinases, especially cathepsin K, play an important role in osteoclastic degradation of bone matrix proteins and the process can, consequently, be significantly inhibited by cysteine proteinase inhibitors. We have recently reported that cystatin C and other cysteine proteinase inhibitors also reduce osteoclast formation. However, it is not known which cysteine proteinase(s) are involved in osteoclast differentiation. In the present study, we compared the relative potencies of cystatins C and D as inhibitors of bone resorption in cultured mouse calvariae, osteoclastogenesis in mouse bone marrow cultures, and cathepsin K activity. Inhibition of cathepsin K activity was assessed by determining equilibrium constants for inhibitor complexes in fluorogenic substrate assays. The data demonstrate that whereas human cystatins C and D are equipotent as inhibitors of bone resorption, cystatin D is 10-fold less potent as an inhibitor of osteoclastogenesis and 200-fold less potent as an inhibitor of cathepsin K activity. A recombinant human cystatin C variant with Gly substitutions for residues Arg8, Leu9, Val10, and Trp106 did not inhibit bone resorption, had 1,000-fold decreased inhibitory effect on cathepsin K activity compared to wildtype cystatin C, but was equipotent with wildtype cystatin C as an inhibitor of osteoclastogenesis. It is concluded that (i) different cysteine proteinases are likely to be involved in bone resorption and osteoclast formation, (ii) cathepsin K may not be an exclusive target enzyme in any of the two systems, and (iii) the enzyme(s) involved in osteoclastogenesis might not be a typical papain-like cysteine proteinase.

  1. Association of mitogen-activated protein kinases with microtubules in mouse macrophages

    PubMed Central

    1996-01-01

    Taxol, a microtubule-binding diterpene, mimics many effects of lipopolysaccharide (LPS) on mouse macrophages. The LPS-mimetic effects of taxol appear to be under the same genetic control as responses to LPS itself. Thus we have postulated a role for microtubule-associated proteins (MAP) in the response of macrophages to LPS. Stimulation of macrophages by LPS quickly induces the activation of mitogen-activated protein kinases (MAPK). MAPK are generally considered cytosolic enzymes. Herein we report that much of the LPS-activatable pool of MAPK in primary mouse peritoneal macrophages is microtubule associated. By immunofluorescence, MAPK were localized to colchicine- and nocodazole- disruptible filaments. From both mouse brain and RAW 264.7 macrophages, MAPK could be coisolated with polymerized tubulin. Fractionation of primary macrophages into cytosol-, microfilament-, microtubule-, and intermediated filament-rich extracts revealed that approximately 10% of MAPK but none of MAPK kinase (MEK1A and MEK2) was microtubule bound. Exposure of macrophages to LPS did not change the proportion of MAPK bound to microtubules, but preferentially activated the microtubule- associated pool. These findings confirm the prediction that LPS activates a kinase bound to microtubules. Together with LPS-mimetic actions of taxol and the shared genetic control of responses to LPS and taxol, these results support the hypothesis that a major LPS-signaling pathway in mouse macrophages may involve activation of one or more microtubule-associated kinases. PMID:8666946

  2. Diversity of Transgenic Mouse Models for Selective Targeting of Midbrain Dopamine Neurons

    PubMed Central

    Lammel, Stephan; Steinberg, Elizabeth E.; Földy, Csaba; Wall, Nicholas R.; Beier, Kevin; Luo, Liqun; Malenka, Robert C.

    2015-01-01

    Ventral tegmental area (VTA) dopamine (DA) neurons have been implicated in reward, aversion, salience, cognition, and several neuropsychiatric disorders. Optogenetic approaches involving transgenic Cre-driver mouse lines provide powerful tools for dissecting DA-specific functions. However, the emerging complexity of VTA circuits requires Cre-driver mouse lines that restrict transgene expression to a precisely defined cell population. Because of recent work reporting that VTA DA neurons projecting to the lateral habenula release GABA, but not DA, we performed an extensive anatomical, molecular, and functional characterization of prominent DA transgenic mouse driver lines. We find that transgenes under control of the tyrosine hydroxylase, but not the dopamine transporter, promoter exhibit dramatic non-DA cell-specific expression patterns within and around VTA nuclei. Our results demonstrate how Cre expression in unintentionally targeted cells in transgenic mouse lines can confound the interpretation of supposedly cell-type-specific experiments. This Matters Arising paper is in response to Stamatakis et al. (2013), published in Neuron. See also the Matters Arising Response paper by Stuber et al. (2015), published concurrently with this Matters Arising in Neuron. PMID:25611513

  3. Macrophage-mediated inflammatory response decreases mycobacterial survival in mouse MSCs by augmenting NO production

    PubMed Central

    Yang, Kun; Wu, Yongjian; Xie, Heping; Li, Miao; Ming, Siqi; Li, Liyan; Li, Meiyu; Wu, Minhao; Gong, Sitang; Huang, Xi

    2016-01-01

    Mycobacterium tuberculosis (MTB) is a hard-to-eradicate intracellular microbe, which escapes host immune attack during latent infection. Recent studies reveal that mesenchymal stem cells (MSCs) provide a protective niche for MTB to maintain latency. However, the regulation of mycobacterial residency in MSCs in the infectious microenvironment remains largely unknown. Here, we found that macrophage-mediated inflammatory response during MTB infection facilitated the clearance of bacilli residing in mouse MSCs. Higher inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production were observed in mouse MSCs under macrophage-mediated inflammatory circumstance. Blocking NO production in MSCs increased the survival of intracellular mycobacteria, indicating NO-mediated antimycobacterial activity. Moreover, both nuclear factor κB (NF-κB) and Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathways were involved in iNOS expression and NO production in inflammatory microenvironment. Furthermore, pro-inflammatory cytokine interleukin-1β could trigger NO production in MSCs and exert anti-mycobacterial activity via NF-κB signaling pathway. Neutralization of interleukin-1β in macrophage-mediated inflammatory microenvironment dampened the ability of mouse MSCs to produce NO. Together, our findings demonstrated that macrophage-mediated inflammatory response during mycobacterial infection promotes the clearance of bacilli in mouse MSCs by increasing NO production, which may provide a better understanding of latent MTB infection. PMID:27251437

  4. Abnormalities in the tricarboxylic Acid cycle in Huntington disease and in a Huntington disease mouse model.

    PubMed

    Naseri, Nima N; Xu, Hui; Bonica, Joseph; Vonsattel, Jean Paul G; Cortes, Etty P; Park, Larry C; Arjomand, Jamshid; Gibson, Gary E

    2015-06-01

    Glucose metabolism is reduced in the brains of patients with Huntington disease (HD). The mechanisms underlying this deficit, its link to the pathology of the disease, and the vulnerability of the striatum in HD remain unknown. Abnormalities in some of the key mitochondrial enzymes involved in glucose metabolism, including the pyruvate dehydrogenase complex (PDHC) and the tricarboxylic acid (TCA) cycle, may contribute to these deficits. Here, activities for these enzymes and select protein levels were measured in human postmortem cortex and in striatum and cortex of an HD mouse model (Q175); mRNA levels encoding for these enzymes were also measured in the Q175 mouse cortex. The activities of PDHC and nearly all of the TCA cycle enzymes were dramatically lower (-50% to 90%) in humans than in mice. The activity of succinate dehydrogenase increased with HD in human (35%) and mouse (23%) cortex. No other changes were detected in the human HD cortex or mouse striatum. In Q175 cortex, there were increased activities of PDHC (+12%) and aconitase (+32%). Increased mRNA levels for succinyl thiokinase (+88%) and isocitrate dehydrogenase (+64%) suggested an upregulation of the TCA cycle. These patterns of change differ from those reported in other diseases, which may offer unique metabolic therapeutic opportunities for HD patients.

  5. Macrophage-mediated inflammatory response decreases mycobacterial survival in mouse MSCs by augmenting NO production.

    PubMed

    Yang, Kun; Wu, Yongjian; Xie, Heping; Li, Miao; Ming, Siqi; Li, Liyan; Li, Meiyu; Wu, Minhao; Gong, Sitang; Huang, Xi

    2016-01-01

    Mycobacterium tuberculosis (MTB) is a hard-to-eradicate intracellular microbe, which escapes host immune attack during latent infection. Recent studies reveal that mesenchymal stem cells (MSCs) provide a protective niche for MTB to maintain latency. However, the regulation of mycobacterial residency in MSCs in the infectious microenvironment remains largely unknown. Here, we found that macrophage-mediated inflammatory response during MTB infection facilitated the clearance of bacilli residing in mouse MSCs. Higher inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production were observed in mouse MSCs under macrophage-mediated inflammatory circumstance. Blocking NO production in MSCs increased the survival of intracellular mycobacteria, indicating NO-mediated antimycobacterial activity. Moreover, both nuclear factor κB (NF-κB) and Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathways were involved in iNOS expression and NO production in inflammatory microenvironment. Furthermore, pro-inflammatory cytokine interleukin-1β could trigger NO production in MSCs and exert anti-mycobacterial activity via NF-κB signaling pathway. Neutralization of interleukin-1β in macrophage-mediated inflammatory microenvironment dampened the ability of mouse MSCs to produce NO. Together, our findings demonstrated that macrophage-mediated inflammatory response during mycobacterial infection promotes the clearance of bacilli in mouse MSCs by increasing NO production, which may provide a better understanding of latent MTB infection.

  6. Chromosomal localization and genomic characterization of the mouse melastatin gene (Mlsn1).

    PubMed

    Hunter, J J; Shao, J; Smutko, J S; Dussault, B J; Nagle, D L; Woolf, E A; Holmgren, L M; Moore, K J; Shyjan, A W

    1998-11-15

    We recently described a novel gene, melastatin, whose expression is inversely correlated with melanoma aggressiveness. Chromosomal localization of this gene places it on mouse chromosome 7 and in the 15q13-q14 region of the human genome. Although expression patterns and chromosomal localization in the mouse are consistent with involvement of melastatin mutations in the mouse ruby-eye-2 defect, congenic analysis showed genetic segregation of the two loci. Cloning of the full-length human cDNA revealed a much larger transcript than we had previously identified, corresponding to a 1533-amino-acid protein product with homology to members of the transient receptor potential (Trp) family of calcium channels. The mouse melastatin gene contains 27 exons and spans at least 58 kb of genomic DNA. The promoter region of Mlsn1 contains four potential microphthalmia binding sites including an M box, a transcriptional regulatory element unique to genes with a restricted melanocytic expression pattern. A 1-kb PvuII fragment from this region was capable of driving high levels of luciferase expression in B16 melanoma cells. PMID:9806836

  7. In vivo analysis of mouse gastrin gene regulation in enhanced GFP-BAC transgenic mice.

    PubMed

    Takaishi, Shigeo; Shibata, Wataru; Tomita, Hiroyuki; Jin, Guangchun; Yang, Xiangdong; Ericksen, Russell; Dubeykovskaya, Zinaida; Asfaha, Samuel; Quante, Michael; Betz, Kelly S; Shulkes, Arthur; Wang, Timothy C

    2011-02-01

    Gastrin is secreted from a subset of neuroendocrine cells residing in the gastric antrum known as G cells, but low levels are also expressed in fetal pancreas and intestine and in many solid malignancies. Although past studies have suggested that antral gastrin is transcriptionally regulated by inflammation, gastric pH, somatostatin, and neoplastic transformation, the transcriptional regulation of gastrin has not previously been demonstrated in vivo. Here, we describe the creation of an enhanced green fluorescent protein reporter (mGAS-EGFP) mouse using a bacterial artificial chromosome that contains the entire mouse gastrin gene. Three founder lines expressed GFP signals in the gastric antrum and the transitional zone to the corpus. In addition, GFP(+) cells could be detected in the fetal pancreatic islets and small intestinal villi, but not in these organs of the adult mice. The administration of acid-suppressive reagents such as proton pump inhibitor omeprazole and gastrin/CCK-2 receptor antagonist YF476 significantly increased GFP signal intensity and GFP(+) cell numbers in the antrum, whereas these parameters were decreased by overnight fasting, octreotide (long-lasting somatostatin ortholog) infusion, and Helicobacter felis infection. GFP(+) cells were also detected in the anterior lobe of the pituitary gland and importantly in the colonic tumor cells induced by administration with azoxymethane and dextran sulfate sodium salt. This transgenic mouse provides a useful tool to study the regulation of mouse gastrin gene in vivo, thus contributing to our understanding of the mechanisms involved in transcriptional control of the gastrin gene.

  8. Zinc Chloride Transiently Maintains Mouse Embryonic Stem Cell Pluripotency by Activating Stat3 Signaling

    PubMed Central

    Hu, Jing; Yang, Zhiyong; Wang, Jinbo; Yu, Jia; Guo, Jing; Liu, Shiying; Qian, Chunmei; Song, Liwen; Wu, Yi; Cheng, Jiajing

    2016-01-01

    An improved understanding of the pluripotency maintenance of embryonic stem (ES) cells is important for investigations of early embryo development and for cell replacement therapy, but the mechanism behind pluripotency is still incompletely understood. Recent findings show that zinc, an essential trace element in humans, is critically involved in regulating various signaling pathways and genes expression. However, its role in ES cell fate determination remains to be further explored. Here we showed that 2μM zinc chloride (ZnCl2) transiently maintained mouse ES cell pluripotency in vitro. The cultured mouse ES cells remained undifferentiated under 2μM ZnCl2 treatment in leukemia inhibitory factor (LIF) withdrawal, retinoic acid (RA) or embryoid bodies (EBs) differentiation assays. In addition, ZnCl2 increased pluripotency genes expression and inhibited differentiation genes expression. Further mechanistic studies revealed that ZnCl2 transiently activated signal transducers and activators of transcription 3 (Stat3) signaling through promoting Stat3 phosphorylation. Inhibition of Stat3 signaling abrogated the effects of ZnCl2 on mouse ES cell pluripotency. Taken together, this study demonstrated a critical role of zinc in the pluripotency maintenance of mouse ES cells, as well as an important regulator of Stat3 signaling. PMID:26910359

  9. DAPK3 Suppresses Acini Morphogenesis and is Required for Mouse Development

    PubMed Central

    Kocher, Brandon A.; White, Lynn S.; Piwnica-Worms, David

    2014-01-01

    Death-associated protein kinase (DAPK3) is a serine/threonine kinase involved in various signaling pathways important to tissue homeostasis and mammalian biology. Considered to be a putative tumor suppressor, the molecular mechanism by which DAPK3 exerts its suppressive function is not fully understood and the field lacks an appropriate mouse model. To address these gaps, an in vitro 3D tumorigenesis model was used and a constitutive DAPK3 knockout mouse was generated. In the 3D morphogenesis model, loss of DAPK3 through lentiviral-mediated knockdown enlarged acinar size by accelerated acini proliferation and apoptosis while maintaining acini polarity. Depletion of DAPK3 enhanced growth factor-dependent mTOR activation and, furthermore, enlarged DAPK3 acini structures were uniquely sensitive to low doses of rapamycin. Simultaneous knockdown of RAPTOR, a key mTORC1 component, reversed the augmented acinar size in DAPK3-depleted structures indicating an epistatic interaction. Utilizing a validated gene trap strategy to generate a constitutive DAPK3 knockout mouse, it was demonstrated that DAPK3 is vital for early mouse development. The Dapk3 promoter exhibits spatio-temporal activity in developing mice and is actively expressed in normal breast epithelia of adult mice. Importantly, reduction of DAPK3 expression correlates with the development of DCIS and more aggressive breast cancer as observed in the Oncomine database of clinical breast cancer specimens. PMID:25304685

  10. Cellular functions of genetically imprinted genes in human and mouse as annotated in the gene ontology.

    PubMed

    Hamed, Mohamed; Ismael, Siba; Paulsen, Martina; Helms, Volkhard

    2012-01-01

    By analyzing the cellular functions of genetically imprinted genes as annotated in the Gene Ontology for human and mouse, we found that imprinted genes are often involved in developmental, transport and regulatory processes. In the human, paternally expressed genes are enriched in GO terms related to the development of organs and of anatomical structures. In the mouse, maternally expressed genes regulate cation transport as well as G-protein signaling processes. Furthermore, we investigated if imprinted genes are regulated by common transcription factors. We identified 25 TF families that showed an enrichment of binding sites in the set of imprinted genes in human and 40 TF families in mouse. In general, maternally and paternally expressed genes are not regulated by different transcription factors. The genes Nnat, Klf14, Blcap, Gnas and Ube3a contribute most to the enrichment of TF families. In the mouse, genes that are maternally expressed in placenta are enriched for AP1 binding sites. In the human, we found that these genes possessed binding sites for both, AP1 and SP1. PMID:23226257

  11. Mouse Activity across Time Scales: Fractal Scenarios

    PubMed Central

    Lima, G. Z. dos Santos; Lobão-Soares, B.; do Nascimento, G. C.; França, Arthur S. C.; Muratori, L.; Ribeiro, S.; Corso, G.

    2014-01-01

    In this work we devise a classification of mouse activity patterns based on accelerometer data using Detrended Fluctuation Analysis. We use two characteristic mouse behavioural states as benchmarks in this study: waking in free activity and slow-wave sleep (SWS). In both situations we find roughly the same pattern: for short time intervals we observe high correlation in activity - a typical 1/f complex pattern - while for large time intervals there is anti-correlation. High correlation of short intervals ( to : waking state and to : SWS) is related to highly coordinated muscle activity. In the waking state we associate high correlation both to muscle activity and to mouse stereotyped movements (grooming, waking, etc.). On the other side, the observed anti-correlation over large time scales ( to : waking state and to : SWS) during SWS appears related to a feedback autonomic response. The transition from correlated regime at short scales to an anti-correlated regime at large scales during SWS is given by the respiratory cycle interval, while during the waking state this transition occurs at the time scale corresponding to the duration of the stereotyped mouse movements. Furthermore, we find that the waking state is characterized by longer time scales than SWS and by a softer transition from correlation to anti-correlation. Moreover, this soft transition in the waking state encompass a behavioural time scale window that gives rise to a multifractal pattern. We believe that the observed multifractality in mouse activity is formed by the integration of several stereotyped movements each one with a characteristic time correlation. Finally, we compare scaling properties of body acceleration fluctuation time series during sleep and wake periods for healthy mice. Interestingly, differences between sleep and wake in the scaling exponents are comparable to previous works regarding human heartbeat. Complementarily, the nature of these sleep-wake dynamics could lead to a better

  12. New routes for transgenesis of the mouse.

    PubMed

    Belizário, José E; Akamini, Priscilla; Wolf, Philip; Strauss, Bryan; Xavier-Neto, José

    2012-08-01

    Transgenesis refers to the molecular genetic techniques for directing specific insertions, deletions and point mutations in the genome of germ cells in order to create genetically modified organisms (GMO). Genetic modification is becoming more practicable, efficient and predictable with the development and use of a variety of cell and molecular biology tools and DNA sequencing technologies. A collection of plasmidial and viral vectors, cell-type specific promoters, positive and negative selectable markers, reporter genes, drug-inducible Cre-loxP and Flp/FRT recombinase systems are available which ensure efficient transgenesis in the mouse. The technologies for the insertion and removal of genes by homologous-directed recombination in embryonic stem cells (ES) and generation of targeted gain- and loss-of function alleles have allowed the creation of thousands of mouse models of a variety of diseases. The engineered zinc finger nucleases (ZFNs) and small hairpin RNA-expressing constructs are novel tools with useful properties for gene knockout free of ES manipulation. In this review we briefly outline the different approaches and technologies for transgenesis as well as their advantages and disadvantages. We also present an overview on how the novel integrative mouse and human genomic databases and bioinformatics approaches have been used to understand genotype-phenotype relationships of hundreds of mutated and candidate disease genes in mouse models. The updating and continued improvements of the genomic technologies will eventually help us to unraveling the biological and pathological processes in such a way that they can be translated more efficiently from mouse to human and vise-versa.

  13. The Mouse Genome Database: integration of and access to knowledge about the laboratory mouse.

    PubMed

    Blake, Judith A; Bult, Carol J; Eppig, Janan T; Kadin, James A; Richardson, Joel E

    2014-01-01

    The Mouse Genome Database (MGD) (http://www.informatics.jax.org) is the community model organism database resource for the laboratory mouse, a premier animal model for the study of genetic and genomic systems relevant to human biology and disease. MGD maintains a comprehensive catalog of genes, functional RNAs and other genome features as well as heritable phenotypes and quantitative trait loci. The genome feature catalog is generated by the integration of computational and manual genome annotations generated by NCBI, Ensembl and Vega/HAVANA. MGD curates and maintains the comprehensive listing of functional annotations for mouse genes using the Gene Ontology, and MGD curates and integrates comprehensive phenotype annotations including associations of mouse models with human diseases. Recent improvements include integration of the latest mouse genome build (GRCm38), improved access to comparative and functional annotations for mouse genes with expanded representation of comparative vertebrate genomes and new loads of phenotype data from high-throughput phenotyping projects. All MGD resources are freely available to the research community.

  14. Multiple myeloma involving the orbit.

    PubMed

    Fay, A M; Leib, M L; Fountain, K S

    1998-01-01

    Multiple myeloma is a plasma cell malignancy often associated with destructive skeletal lesions. Orbital involvement in multiple myeloma is rare. Risk factors for orbital involvement have not been established, although risk may vary with immunoglobulin subtype. Early detection of orbital plasmacytoma may affect treatment and clinical course. A case is reported of multiple myeloma without elevated serum immunoglobulins that involves the orbit, and the implications of early detection are discussed. The patient was first examined by an ophthalmologist 13 months after multiple myeloma was diagnosed and 5 months after the external appearance of an orbital tumor. Urine protein electrophoresis demonstrated kappa light chains. Hypergammaglobulinemia was not detected. Plain-film roentgenography showed orbital involvement at the time of initial diagnosis. An impressive clinical response to external beam radiation therapy was seen. Attention to immunoprotein characteristics in multiple myeloma may help to identify risk factors for orbital involvement. Early detection may permit safer and equally effective treatment. All patients with multiple myeloma should undergo thorough ophthalmic examination at the time of initial diagnosis.

  15. Fibrosis and inflammation are greater in muscles of beta-sarcoglycan-null mouse than mdx mouse.

    PubMed

    Gibertini, Sara; Zanotti, Simona; Savadori, Paolo; Curcio, Maurizio; Saredi, Simona; Salerno, Franco; Andreetta, Francesca; Bernasconi, Pia; Mantegazza, Renato; Mora, Marina

    2014-05-01

    The Sgcb-null mouse, with knocked-down β-sarcoglycan, develops severe muscular dystrophy as in type 2E human limb girdle muscular dystrophy. The mdx mouse, lacking dystrophin, is the most used model for Duchenne muscular dystrophy (DMD). Unlike DMD, the mdx mouse has mild clinical features and shows little fibrosis in limb muscles. To characterize ECM protein deposition and the progression of muscle fibrosis, we evaluated protein and transcript levels of collagens I, III and VI, decorin, and TGF-β1, in quadriceps and diaphragm, at 2, 4, 8, 12, 26, and 52 weeks in Sgcb-null mice, and protein levels at 12, 26, and 52 weeks in mdx mice. In Sgcb-null mice, severe morphological disruption was present from 4 weeks in both quadriceps and diaphragm, and included conspicuous deposition of extracellular matrix components. Histopathological features of Sgcb-null mouse muscles were similar to those of age-matched mdx muscles at all ages examined, but, in the Sgcb-null mouse, the extent of connective tissue deposition was generally greater than mdx. Furthermore, in the Sgcb-null mouse, the amount of all three collagen isoforms increased steadily, while, in the mdx, they remained stable. We also found that, at 12 weeks, macrophages were significantly more numerous in mildly inflamed areas of Sgcb-null quadriceps compared to mdx quadriceps (but not in highly inflamed regions), while, in the diaphragm, macrophages did not differ significantly between the two models, in either region. Osteopontin mRNA was also significantly greater at 12 weeks in laser-dissected highly inflamed areas of the Sgcb-null quadriceps compared to the mdx quadriceps. TGF-β1 was present in areas of degeneration-regeneration, but levels were highly variable and in general did not differ significantly between the two models and controls. The roles of the various subtypes of macrophages in muscle repair and fibrosis in the two models require further study. The Sgcb-null mouse, which develops early fibrosis

  16. Lupus pernio without systemic involvement

    PubMed Central

    Anjaneyan, Gopikrishnan; Vora, Rita

    2013-01-01

    Sarcoidosis is a multisystem, granulomatous disease of unknown etiology that can affect the pulmonary, reticulo-endothelial, skin, gastrointestinal, cardiac, musculo – skeletal, endocrine or central nervous system. Exclusive cutaneous involvement is very rare in sarcoidosis. Lupus pernio is a variant of cutaneous sarcoidosis presenting with erythematous to violaceous nodules and plaques located symmetrically over the nose, cheeks, ears and digits. We present a case of lupus pernio which showed rapid improvement with topical steroids and has yet not developed any systemic involvement even after 6 years of regular follow up. PMID:24350015

  17. Multisystem involvement in neuromyelitis optica

    PubMed Central

    Langille, Megan M.; Desai, Jay

    2015-01-01

    We describe a case of pediatric neuromyelitis optica (NMO) with muscle and lung involvement in addition to central nervous system disease. Our patient initially presented with features of area postrema syndrome, then subsequently with optic neuritis. The patient also had recurrent hyperCKemia that responded to corticosteroids. Finally, axillary and hilar adenopathy with pulmonary consolidation were noted as well and responded to immunomodulation. Our case highlights multisystem involvement in NMO including non-infectious pulmonary findings which have not been described in the pediatric population previously. PMID:26538850

  18. MouseMine: a new data warehouse for MGI.

    PubMed

    Motenko, H; Neuhauser, S B; O'Keefe, M; Richardson, J E

    2015-08-01

    MouseMine (www.mousemine.org) is a new data warehouse for accessing mouse data from Mouse Genome Informatics (MGI). Based on the InterMine software framework, MouseMine supports powerful query, reporting, and analysis capabilities, the ability to save and combine results from different queries, easy integration into larger workflows, and a comprehensive Web Services layer. Through MouseMine, users can access a significant portion of MGI data in new and useful ways. Importantly, MouseMine is also a member of a growing community of online data resources based on InterMine, including those established by other model organism databases. Adopting common interfaces and collaborating on data representation standards are critical to fostering cross-species data analysis. This paper presents a general introduction to MouseMine, presents examples of its use, and discusses the potential for further integration into the MGI interface. PMID:26092688

  19. Genetically Determined Susceptibility to Tuberculosis in Mice Causally Involves Accelerated and Enhanced Recruitment of Granulocytes

    PubMed Central

    Keller, Christine; Hoffmann, Reinhard; Lang, Roland; Brandau, Sven; Hermann, Corinna; Ehlers, Stefan

    2006-01-01

    Classical twin studies and recent linkage analyses of African populations have revealed a potential involvement of host genetic factors in susceptibility or resistance to Mycobacterium tuberculosis infection. In order to identify the candidate genes involved and test their causal implication, we capitalized on the mouse model of tuberculosis, since inbred mouse strains also differ substantially in their susceptibility to infection. Two susceptible and two resistant mouse strains were aerogenically infected with 1,000 CFU of M. tuberculosis, and the regulation of gene expression was examined by Affymetrix GeneChip U74A array with total lung RNA 2 and 4 weeks postinfection. Four weeks after infection, 96 genes, many of which are involved in inflammatory cell recruitment and activation, were regulated in common. One hundred seven genes were differentially regulated in susceptible mouse strains, whereas 43 genes were differentially expressed only in resistant mice. Data mining revealed a bias towards the expression of genes involved in granulocyte pathophysiology in susceptible mice, such as an upregulation of those for the neutrophil chemoattractant LIX (CXCL5), interleukin 17 receptor, phosphoinositide kinase 3 delta, or gamma interferon-inducible protein 10. Following M. tuberculosis challenge in both airways or peritoneum, granulocytes were recruited significantly faster and at higher numbers in susceptible than in resistant mice. When granulocytes were efficiently depleted by either of two regimens at the onset of infection, only susceptible mice survived aerosol challenge with M. tuberculosis significantly longer than control mice. We conclude that initially enhanced recruitment of granulocytes contributes to susceptibility to tuberculosis. PMID:16790804

  20. Molecular mechanism of extinction of liver-specific functions in mouse hepatoma x rat fibroblast hybrids: extinction of the albumin gene

    SciTech Connect

    Papaconstantinou, J.; Wong, E.; Ratrie, H.; Szpirer, C.; Szpirer, J.

    1982-01-01

    Hybrids formed by the fusion of mouse hepatoma (BWTG3) and rat fibroblast (JF1) cells exhibit the extinction of mouse albumin and ..cap alpha..-fetoprotein synthesis. Karyotype analyses suggest that all parental chromosomes are present in the hybrids. The extinction, therefore, of mouse hepatocyte genes is attributed to the inhibitory action of the rat genome. In these studies, we show that these hybrids possess and express the mouse ..beta..-glucyronidase gene (which is encoded on the same chromosome as the mouse albumin and ..cap alpha..-fetoprotein gene), and we present data of Southern blot analysis which demonstrate that such hybrids have indeed retained both mouse and rat albumin DNA sequences. In addition, using mouse albumin cDNA, we have shown by cDNA-RNA reassociation kinetics that albumin mRNA is virtually absent in these hybrids. We conclude from these studies that the extinction of albumin synthesis involves a mechanism which results in the loss of cytoplasmic albumin mRNA.

  1. Imaging Ca2+ dynamics in cone photoreceptor axon terminals of the mouse retina.

    PubMed

    Kulkarni, Manoj; Schubert, Timm; Baden, Tom; Wissinger, Bernd; Euler, Thomas; Paquet-Durand, Francois

    2015-01-01

    Retinal cone photoreceptors (cones) serve daylight vision and are the basis of color discrimination. They are subject to degeneration, often leading to blindness in many retinal diseases. Calcium (Ca(2+)), a key second messenger in photoreceptor signaling and metabolism, has been proposed to be indirectly linked with photoreceptor degeneration in various animal models. Systematically studying these aspects of cone physiology and pathophysiology has been hampered by the difficulties of electrically recording from these small cells, in particular in the mouse where the retina is dominated by rod photoreceptors. To circumvent this issue, we established a two-photon Ca(2+) imaging protocol using a transgenic mouse line that expresses the genetically encoded Ca(2+) biosensor TN-XL exclusively in cones and can be crossbred with mouse models for photoreceptor degeneration. The protocol described here involves preparing vertical sections ("slices") of retinas from mice and optical imaging of light stimulus-evoked changes in cone Ca(2+) level. The protocol also allows "in-slice measurement" of absolute Ca(2+) concentrations; as the recordings can be followed by calibration. This protocol enables studies into functional cone properties and is expected to contribute to the understanding of cone Ca(2+) signaling as well as the potential involvement of Ca(2+) in photoreceptor death and retinal degeneration. PMID:25993489

  2. Recent advances in mouse models of obesity- and nonalcoholic steatohepatitis-associated hepatocarcinogenesis

    PubMed Central

    Nakagawa, Hayato

    2015-01-01

    Hepatocellular carcinoma (HCC) is the fifth most common cancer, and obesity has been established as a risk factor for HCC development. Nonalcoholic steatohepatitis (NASH) is apparently the key link between obesity and hepatocarcinogenesis, and obesity also accelerates HCC development synergistically with other risk factors, such as hepatitis virus infection and alcohol consumption. As an explanation for the pathogenesis of NASH, the so-called “two-hit” theory has been widely accepted, but recently, a better model, the so-called “multiple-hits hypothesis” was proposed, which states that many disease-promoting factors may occur in parallel, rather than consecutively. However, the overall mechanism remains largely unknown. Various cell-cell and organ-organ interactions are involved in the pathogenesis of NASH, and thus appropriate in vivo disease models are essential for a deeper understanding. However, replicating the full spectrum of human NASH has been difficult, as NASH involves obesity, insulin resistance, steatohepatitis, fibrosis, and ultimately HCC, and the lack of an appropriate mouse model has been a considerable barrier to determining the missing links among obesity, NASH, and HCC. In recent years, several innovative mouse models presenting obesity- and NASH-associated HCC have been established by modified diets, chemotoxic agents, genetic manipulation, or a combination of these factors, shedding some light on this complex network and providing new therapeutic strategies. Thus, in this paper, I review the mouse models of obesity- and NASH-associated HCC, especially focusing on recent advances and their clinical relevance. PMID:26301053

  3. Imaging Ca2+ dynamics in cone photoreceptor axon terminals of the mouse retina.

    PubMed

    Kulkarni, Manoj; Schubert, Timm; Baden, Tom; Wissinger, Bernd; Euler, Thomas; Paquet-Durand, Francois

    2015-05-06

    Retinal cone photoreceptors (cones) serve daylight vision and are the basis of color discrimination. They are subject to degeneration, often leading to blindness in many retinal diseases. Calcium (Ca(2+)), a key second messenger in photoreceptor signaling and metabolism, has been proposed to be indirectly linked with photoreceptor degeneration in various animal models. Systematically studying these aspects of cone physiology and pathophysiology has been hampered by the difficulties of electrically recording from these small cells, in particular in the mouse where the retina is dominated by rod photoreceptors. To circumvent this issue, we established a two-photon Ca(2+) imaging protocol using a transgenic mouse line that expresses the genetically encoded Ca(2+) biosensor TN-XL exclusively in cones and can be crossbred with mouse models for photoreceptor degeneration. The protocol described here involves preparing vertical sections ("slices") of retinas from mice and optical imaging of light stimulus-evoked changes in cone Ca(2+) level. The protocol also allows "in-slice measurement" of absolute Ca(2+) concentrations; as the recordings can be followed by calibration. This protocol enables studies into functional cone properties and is expected to contribute to the understanding of cone Ca(2+) signaling as well as the potential involvement of Ca(2+) in photoreceptor death and retinal degeneration.

  4. Imaging Ca2+ Dynamics in Cone Photoreceptor Axon Terminals of the Mouse Retina

    PubMed Central

    Kulkarni, Manoj; Schubert, Timm; Baden, Tom; Wissinger, Bernd; Euler, Thomas; Paquet-Durand, Francois

    2015-01-01

    Retinal cone photoreceptors (cones) serve daylight vision and are the basis of color discrimination. They are subject to degeneration, often leading to blindness in many retinal diseases. Calcium (Ca2+), a key second messenger in photoreceptor signaling and metabolism, has been proposed to be indirectly linked with photoreceptor degeneration in various animal models. Systematically studying these aspects of cone physiology and pathophysiology has been hampered by the difficulties of electrically recording from these small cells, in particular in the mouse where the retina is dominated by rod photoreceptors. To circumvent this issue, we established a two-photon Ca2+ imaging protocol using a transgenic mouse line that expresses the genetically encoded Ca2+ biosensor TN-XL exclusively in cones and can be crossbred with mouse models for photoreceptor degeneration. The protocol described here involves preparing vertical sections (“slices”) of retinas from mice and optical imaging of light stimulus-evoked changes in cone Ca2+ level. The protocol also allows “in-slice measurement” of absolute Ca2+ concentrations; as the recordings can be followed by calibration. This protocol enables studies into functional cone properties and is expected to contribute to the understanding of cone Ca2+ signaling as well as the potential involvement of Ca2+ in photoreceptor death and retinal degeneration. PMID:25993489

  5. Methylation-Associated Gene Silencing of RARB in Areca Carcinogens Induced Mouse Oral Squamous Cell Carcinoma

    PubMed Central

    Tsou, Yung-An; Fan, Shin-Ru; Tsai, Ming-Hsui; Chen, Hsiao-Ling; Chang, Nai-Wen; Cheng, Ju-Chien

    2014-01-01

    Regarding oral squamous cell carcinoma (OSCC) development, chewing areca is known to be a strong risk factor in many Asian cultures. Therefore, we established an OSCC induced mouse model by 4-nitroquinoline-1-oxide (4-NQO), or arecoline, or both treatments, respectively. These are the main two components of the areca nut that could increase the occurrence of OSCC. We examined the effects with the noncommercial MCGI (mouse CpG islands) microarray for genome-wide screening the DNA methylation aberrant in induced OSCC mice. The microarray results showed 34 hypermethylated genes in 4-NQO plus arecoline induced OSCC mice tongue tissues. The examinations also used methylation-specific polymerase chain reaction (MS-PCR) and bisulfite sequencing to realize the methylation pattern in collected mouse tongue tissues and human OSCC cell lines of different grades, respectively. These results showed that retinoic acid receptor β (RARB) was indicated in hypermethylation at the promoter region and the loss of expression during cancer development. According to the results of real-time PCR, it was shown that de novo DNA methyltransferases were involved in gene epigenetic alternations of OSCC. Collectively, our results showed that RARB hypermethylation was involved in the areca-associated oral carcinogenesis. PMID:25197641

  6. Proteomic analysis of the mouse brain after repetitive exposure to hypoxia.

    PubMed

    Cui, Can; Zhou, Tao; Li, Jingyi; Wang, Hong; Li, Xiaorong; Xiong, Jie; Xu, Pingxiang; Xue, Ming

    2015-07-01

    Hypoxic preconditioning (HPC) is known to have a protective effect against hypoxic damage; however, the precise mechanisms involved remain unknown. In this study, an acute and repetitive hypoxia mouse model, two-dimensional fluorescence difference gel electrophoresis (2D-DIGE) coupled with matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF/TOF-MS), and Western blot experiments were used to identify the differential expression of key proteins in the mouse brain during HPC. Approximately 2100 2D-DIGE spots were observed following gel imaging and spot detection. Significant differences (p < 0.05) in the expression of 66 proteins were observed between the 3× HPC treatment group and the control group, 45 proteins were observed between the 6× HPC treatment group and the control group, and 70 proteins were observed between the 3× HPC treatment group and the 6× HPC group. Consistent results among Western blot, 2D-DIGE and MS methods were observed for the proteins, ATP synthase subunit alpha, malate dehydrogenase, guanine nucleotide-binding protein subunit beta-1 and proteasome subunit alpha type-2. The proteins associated with ATP synthesis and the citric acid cycle were down-regulated, while those linked to glycolysis and oxygen-binding were up-regulated. This proteomic analysis of the mouse brain after HPC furthers understanding of the molecular pathways involved in the protective effect of HPC and these findings provide new insight into the mechanisms of hypoxia and HPC. PMID:25937538

  7. Analysis of Gene and Protein Expression in Atherosclerotic Mouse Aorta by Western Blot and Quantitative Real-Time PCR.

    PubMed

    Rivera-Torres, José

    2015-01-01

    Atherosclerosis involves changes in gene and protein expression patterns in affected arteries. Quantification of these alterations is essential for understanding the molecular mechanisms underlying this pathology. Western blot and real-time PCR-used to quantify protein and messenger RNA levels, respectively-are invaluable molecular biology tools, particularly when material is limited. The availability of many genetically modified mouse models of atherosclerosis makes the mouse aorta an ideal tissue in which to carry out these expression pattern analyses. In this chapter, protocols are presented for mRNA and protein extraction from mouse aorta and for the accurate quantification of mRNA expression by RT-PCR and of proteins by western blot.

  8. Topoisomerase I identified by scleroderma 70 antisera: enrichment of topoisomerase I at the centromere in mouse mitotic cells before anaphase.

    PubMed Central

    Maul, G G; French, B T; van Venrooij, W J; Jimenez, S A

    1986-01-01

    Antibodies derived from scleroderma patients positive for the extractable 70-kDa antigen were shown to react with topoisomerase I. Topoisomerase I was identified by molecular size and by antibody inhibition of the topoisomerase I-specific relaxation of supercoiled plasmid DNA. By using in situ localization by indirect fluorescence, we found topoisomerase I preferentially located in the centromeric regions of the mouse G2-phase cells and chromosomes, while the distribution in human cells is much more dispersed. Moreover, comparison of a published consensus sequence for topoisomerase I binding with mouse satellite DNA revealed a high degree of homology. The localization of topoisomerase I in the centromeres of mouse cells in the later part of the cell cycle and prior to anaphase suggests functional involvement in mitosis. Images PMID:3014535

  9. Separation of mouse embryonic facial ectoderm and mesenchyme.

    PubMed

    Li, Hong; Williams, Trevor

    2013-04-12

    Orofacial clefts are the most frequent craniofacial defects, which affect 1.5 in 1,000 newborns worldwide. Orofacial clefting is caused by abnormal facial development. In human and mouse, initial growth and patterning of the face relies on several small buds of tissue, the facial prominences. The face is derived from six main prominences: paired frontal nasal processes (FNP), maxillary prominences (MxP) and mandibular prominences (MdP). These prominences consist of swellings of mesenchyme that are encased in an overlying epithelium. Studies in multiple species have shown that signaling crosstalk between facial ectoderm and mesenchyme is critical for shaping the face. Yet, mechanistic details concerning the genes involved in these signaling relays are lacking. One way to gain a comprehensive understanding of gene expression, transcription factor binding, and chromatin marks associated with the developing facial ectoderm and mesenchyme is to isolate and characterize the separated tissue compartments. Here we present a method for separating facial ectoderm and mesenchyme at embryonic day (E) 10.5, a critical developmental stage in mouse facial formation that precedes fusion of the prominences. Our method is adapted from the approach we have previously used for dissecting facial prominences. In this earlier study we had employed inbred C57BL/6 mice as this strain has become a standard for genetics, genomics and facial morphology. Here, though, due to the more limited quantities of tissue available, we have utilized the outbred CD-1 strain that is cheaper to purchase, more robust for husbandry, and tending to produce more embryos (12-18) per litter than any inbred mouse strain. Following embryo isolation, neutral protease Dispase II was used to treat the whole embryo. Then, the facial prominences were dissected out, and the facial ectoderm was separated from the mesenchyme. This method keeps both the facial ectoderm and mesenchyme intact. The samples obtained using this

  10. Predictors of Residence Hall Involvement

    ERIC Educational Resources Information Center

    Arboleda, Ana; Wang, Yongyi; Shelley, Mack C., II; Whalen, Donald F.

    2003-01-01

    Residence hall students' (N = 1,186, 52% male, 90% White, 66% freshmen) involvement in their living community is influenced significantly by precollege student characteristics (gender, ethnicity), classification, attitudes (toward hall director, house cabinet, academic comfort, social environment, group study), and environmental variables (noise,…

  11. Corporate Involvement in C AI

    ERIC Educational Resources Information Center

    Baker, Justine C.

    1978-01-01

    Historic perspective of computer manufacturers and their contribution to CAI. Corporate CAI products and services are mentioned, as is a forecast for educational involvement by computer corporations. A chart of major computer corporations shows gross sales, net earnings, products and services offered, and other corporate information. (RAO)

  12. Promoting Active Involvement in Classrooms

    ERIC Educational Resources Information Center

    Conderman, Greg; Bresnahan, Val; Hedin, Laura

    2012-01-01

    This article presents a rationale for using active involvement techniques, describes large- and small-group methods based on their documented effectiveness and applicability to K-12 classrooms, and illustrates their use. These approaches include ways of engaging students in large groups (e.g., unison responses, response cards, dry-erase boards,…

  13. Parent Involvement as Ritualized Practice

    ERIC Educational Resources Information Center

    Doucet, Fabienne

    2011-01-01

    This article examines parent involvement (PI) as a ritual system using Turner's concept of root paradigms. Through a twofold analysis, I argue that the highly ritualized nature of PI practices creates a group identity among mainstream parents and schools that marginalizes diverse families. First, I point out three root paradigms in the ritual…

  14. Managing Parent Involvement during Crisis

    ERIC Educational Resources Information Center

    Merriman, Lynette S.

    2008-01-01

    In the wake of 9/11, Hurricane Katrina, and the Virginia Tech shooting tragedy, it is no surprise that concern for students' safety is the primary reason attributed to parents' increased involvement. Parents and university administrators share in their commitment to student safety. However, college and university staff who assume responsibility…

  15. Veterinary involvement in poultry production.

    PubMed

    Parker, Daniel

    2016-01-16

    The worldwide poultry sector is expected to grow substantially over the next few decades, as the world looks to feed a rapidly expanding population. In a further article in Veterinary Record's series looking at the state of different sectors of the veterinary profession, Daniel Parker looks at veterinary involvement in the poultry sector.

  16. Drug Involvement and Academic Striving.

    ERIC Educational Resources Information Center

    Kahn, Malcolm; Holroyd, Kenneth

    This study attempted to clarify the relationship between drug involvement and academic accomplishments. Unlike other studies, it was controlled for aptitude and sex. In a structured interview, the College Behavior Questionnaire (CBQ) was administered to 77 male and 67 female student subjects. Based on the CBQ results three groups were identified:…

  17. Veterinary involvement in poultry production.

    PubMed

    Parker, Daniel

    2016-01-16

    The worldwide poultry sector is expected to grow substantially over the next few decades, as the world looks to feed a rapidly expanding population. In a further article in Veterinary Record's series looking at the state of different sectors of the veterinary profession, Daniel Parker looks at veterinary involvement in the poultry sector. PMID:26769809

  18. Parotid involvement by desmoplastic melanoma.

    PubMed

    Jennings, T A; Okby, N T; Schroer, K R; Wolf, B C; Mihm, M C

    1996-08-01

    Desmoplastic malignant melanoma often arises in sun damaged skin of the head and neck and shows frequent neurotropism. Although metastatic melanoma frequently involve the parotid, direct spread to the parotid has been rarely reported. We evaluated five cases of desmoplastic malignant melanoma involving the parotid gland with clinical and pathological evidence of precursor cutaneous lesions in four of the five cases. The parotid involvement in four cases was tumoural, and three of these were not clinically suspected to be melanoma. The histological appearance in all five cases was that of a sarcomatoid tumour. Immunohistochemistry and electronmicroscopy performed on three of the cases showed only evidence of schwannian differentiation: the tumour cells were positive for S-100 protein and vimentin, and negative for cytokeratin and HMB-45. Electronmicroscopy showed no evidence of melanogenesis. All five tumours showed histological evidence of prominent neurotropism with one case demonstrating extension from overlying skin along cutaneous nerves to the superficial parotid. Thus, desmoplastic malignant melanoma may involve the parotid by neurotropic spread and can be pathologically indistinguishable from malignant schwannoma, a diagnosis which may be made erroneously in the absence of clinical information. PMID:8872151

  19. Parental Involvement in Norwegian Schools

    ERIC Educational Resources Information Center

    Paulsen, Jan Merok

    2012-01-01

    This article examines findings on key challenges of school-parent relations in Norway. The review is based on recent large-scale studies on several issues, including formalized school-parent cooperation, parental involvement in the pedagogical discourse, and teacher perspectives on the parents' role in the school community. Findings suggest a…

  20. Orbital involvement in multifocal fibrosclerosis.

    PubMed Central

    Aylward, G W; Sullivan, T J; Garner, A; Moseley, I; Wright, J E

    1995-01-01

    Multifocal fibrosclerosis is a condition of unknown aetiology, characterised by fibrous lesions occurring at a variety of sites. Clinical variants include retroperitoneal fibrosis, Riedel's thyroiditis, sclerosing cholangitis, and mediastinal fibrosis. Orbital pseudotumour has been reported as a manifestation of this condition. Three patients with multifocal fibrosclerosis in whom orbital involvement was the dominant feature are described. Images PMID:7703203