Measurement of the concentration of serum soluble interleukin-2 receptor alpha chain in dogs with lymphoma.

Soluble interleukin-2 receptor (sIL-2r) is released directly from the surface of lymphocytes expressing interleukin-2 receptor alpha chain (CD25), and its serum concentration has been found to reflect the prognosis of human lymphoproliferative malignancies. In this study, we demonstrated the presence of sIL-2r in canine serum and developed a specific sandwich enzyme-linked immunosorbent assay (ELISA) to quantify the concentration of canine serum sIL-2r. In the immunoprecipitation (IP) assay, CD25 protein weighing approximately 45 kDa was detected in canine serum, smaller than the membrane-bound CD25 (approximately 55 kDa). To measure the concentration of serum sIL-2r in dogs, an ELISA system was developed. Serum sIL-2r levels were significantly higher in dogs with multicentric high-grade B-cell lymphoma before therapy than that in healthy dogs. Serum sIL-2r concentration was also found to be elevated in a proportion of dogs with other types of lymphoma. Changes in serum sIL-2r levels generally paralleled the changes in mass and lymph node size in dogs with high-grade B-cell lymphoma. This study demonstrated that serum sIL-2r level would be a marker to monitor tumour growth and regression in canine lymphoma.

Early pregnancy loss in 15-hydroxyprostaglandin dehydrogenase knockout (15-HPGD-/-) mice due to requirement for embryo 15-HPGD activity.

Prostaglandins (PGs) have critical signaling functions in a variety of processes including the establishment and maintenance of pregnancy, and the initiation of labor. Most PGs are non-enzymatically degraded, however, the two PGs most prominently implicated in the termination of pregnancy, including the initiation of labor, prostaglandin E2 (PGE2) and prostaglandin F2α (PGF2α), are enzymatically degraded by 15-hydroxyprostaglandin dehydrogenase (15-HPGD). The role of PG metabolism by 15-HPGD in the maintenance of pregnancy remains largely unknown, as direct functional studies are lacking. To test the hypothesis that 15-PGDH-mediated PG metabolism is essential for pregnancy maintenance and normal labor timing, we generated and analyzed pregnancy in 15-HPGD knockout mice (Hpgd-/-). We report here that pregnancies resulting from matings between 15-HPGD KO mice (Hpgd-/- X Hpgd-/-KO mating) are terminated at mid gestation due to a requirement for embryo derived 15-HPGD. Aside from altered implantation site spacing, pregnancies from KO matings look grossly and histologically normal at days post coitum (dpc) 6.5 and 7.5 of pregnancy. However, virtually all of these pregnancies are resorbed by dpc 8.5. This resorption is preceded by elevation of PGF2∝ but is not preceded by a decrease in circulating progesterone, suggesting that pregnancy loss is a local inflammatory phenomenon rather than a centrally mediated phenomena. This pregnancy loss can be temporarily deferred by indomethacin treatment, but treated pregnancies are not maintained to term and indomethacin treatment increases maternal mortality. We conclude that PG metabolism to inactive products by embryo derived 15-HPGD is essential for pregnancy maintenance in mice, and may serve a similar function during human pregnancy.

Measurement of Three-Dimensional Force Applied to Elastic Suture Training Pads for Laparoscopic Suturing.

Laparoscopic surgery requires highly trained skills to manipulate the laparoscopic instruments. The effectiveness and efficiency of manipulation training are expected to increase by quantitatively evaluating how a series of trainee's manipulations are different from those conducted by skilled surgeons and providing feedback. However, such detailed feed- back is not available because adequate measurement systems have not been proposed to measure the precise forces applied to suture training pads without disturbing the delicate manipulations. Therefore, we proposed a sensorized suture training pad satisfying the above requirements. Three surgical residents participated in an experiment to measure time sequences of three-dimensional forces applied to the pad when executing a transfixion suture. As a result, the differences in manipulation patterns among the residents were found by dividing the transfixion operation based on the sequential force data and recorded videos.

Preterm birth without progesterone withdrawal in 15-hydroxyprostaglandin dehydrogenase hypomorphic mice.

Parturition is a complex mammalian physiological process whose fundamental determinants have remained elusive. The increasing incidence of human preterm birth, a leading cause of infant mortality, highlights the importance of further understanding mechanisms regulating the timing of birth. Parturition is initiated in most nonprimate mammals, including mice, through a decrease in circulating progesterone caused by elevated prostaglandins. In humans, other higher primates, and guinea pigs, no consistent decrease in circulating progesterone occurs before the onset of labor. The divergence in endocrine control of labor initiation between most mammals compared with the great apes and guinea pigs gives rise to the question: how could a mechanism for the initiation of labor not requiring the withdrawal of progesterone evolve? Here, we genetically modulate prostaglandin signaling to determine the role of prostaglandin catabolism in the timing of birth. We find spontaneous preterm labor in the absence of progesterone withdrawal in 15-hydroxyprostaglandin dehydrogenase hypomorphic mice. The onset of labor in these hypomorphic mice is preceded by prematurely increased concentrations of prostaglandin E(2) and F(2alpha). Moreover, genetic crosses demonstrate a role for fetal genotype in birth timing. Together, these findings demonstrate a 15-hydroxyprostaglandin dehydrogenase-dependent shift in the physiology of murine parturition to one resembling the physiology of higher primates. Thus, endocrine control of labor has the capacity to plastically adapt to changes in genetically determined prostaglandin signals.

Involvement of neural cell adhesion molecule signaling in glial cell line-derived neurotrophic factor-induced analgesia in a rat model of neuropathic pain.

Since neuropathic pain is resistant to conventional analgesics such as opiates and non-steroidal anti-inflammatory drugs, the development of new types of drugs for its treatment has been awaited. Several key molecules associated with nociception have been suggested as potential targets for new analgesics. Glial cell line-derived neurotrophic factor (GDNF) has a variety of functions affecting the survival and development of specified neural cell populations, mediated via transmission of intracellular signals through binding to its high-affinity receptor, GFR*1, and subsequent activation of a tyrosine receptor kinase, RET, neural cell adhesion molecule (NCAM), or other signaling molecules. GDNF also exhibits analgesic effects in rodent models of neuropathic pain, although the underlying mechanisms are still largely unknown, including the intracellular signal transduction involved. We report here that NCAM signaling plays a role in mediating the analgesic effect of GDNF in rats with chronic constrictive injury (CCI). We found that NCAM was expressed in intrinsic neurons in the spinal dorsal horn and in dorsal root ganglion neurons with small cell bodies. Reduction of NCAM expression by NCAM antisense oligodeoxynucleotide administration to CCI rats abolished the analgesic effect of GDNF without affecting RET signaling activation. An NCAM mimetic peptide, C3d, partially reduced the chronic pain induced by CCI. These findings suggest that NCAM signaling plays a critical role in the analgesic effect of GDNF and that development of new drugs activating GDNF-NCAM signaling may represent a new strategy for the relief of intractable pain.

Combined treatment of leukemia cells with vitamin K2 and 1alpha,25-dihydroxy vitamin D3 enhances monocytic differentiation along with becoming resistant to apoptosis by induction of cytoplasmic p21CIP1.

Vitamin K2 (VK2) effectively induces apoptosis in leukemia cell lines, including HL-60 and U937. However, combined treatment of cells with VK2 plus 1alpha,25-dihydroxy vitamin D3 (VD3) resulted in suppression of VK2-inducing apoptosis and pronounced induction of monocytic differentiation as compared with that by VD3 alone. After achieving monocytic differentiation by pre-exposure to VK2 and VD3, the cells became resistant to various apoptotic stimuli including VK2- and H2O2-treatment and serum deprivation. Accumulation of cytoplasm p21CIP1 along with disappearance of nuclear p21CIP1 was detected in cells in response to 96-h treatment with VK2 plus VD3. A stable transfectant, U937-deltaNLS-p21CIP1, which lacked the nuclear localization signal of p21CIP1 and showed overexpression of cytoplasm p21CIP1 without monocytic differentiation, was resistant to apoptosis. These data suggest that a change of intracellular distribution of p21CIP1 from nucleus to cytoplasm along with differentiation appears to be anti-apoptotic. Clinical benefits of using VK2 for treatment of patients with leukemia and myelodysplastic syndrome (MDS) have been reported. Our data suggest that VK2 plus VD3 may be an effective combination for differentiation-based therapy for leukemia and also MDS whose cytopenias are mediated though apoptosis.

Brap2 functions as a cytoplasmic retention protein for p21 during monocyte differentiation.

The cell cycle inhibitor p21 plays an important role in monocytic cell differentiation, during which it translocates from the nucleus to cytoplasm. This process involves the negative regulation of the p21 nuclear localization signal (NLS). Here, we sought to determine the relationship between the cytoplasmic translocation of p21 and another molecule, Brap2, a cytoplasmic protein which binds the NLS of BRCA1 and was recently reported to inactivate KSR in the Ras-activating signal pathway under the name of IMP. We report that p21 and Brap2 directly interact, both in vitro and in vivo, in a manner requiring the NLS of p21 and the C-terminal portion of Brap2. When it is cotransfected with Brap2, p21 is expressed in the cytoplasm. Monocytic differentiation of the promyelomonocytic cell lines U937 and HL60 is associated with the upregulation of Brap2 expression concomitantly with the upregulation and cytoplasmic relocalization of p21. Our results underscore the role played by Brap2 in the process of cytoplasmic translocation of p21 during monocyte differentiation.

Pancytopenia presenting with monosomy 7 which disappeared after immunosuppressive therapy.

Monosomy 7 syndrome in infant is considered as pre-leukemic condition of poor prognosis. However, it seems controversial recently, because some cases of monosomy 7 syndrome showed spontaneous remission. We report 2-year-old girl with severe pancytopenia, who presented with monosomy 7. Morphologically, there was little dysplasia in the trilineage hematopoiesis. Monosomy 7 clone of CD34 positive cells, bone marrow mononuclear cells (BMMNC), and peripheral nuclear cells was 4.0, 40, and 3.8%, respectively. Immunosuppressive therapy was effective along with the disappearance of monosomy 7 clone. WT1 mRNA expression was not increased in monosomy 7 clone. Pathogenesis of monosomy 7 and its relation to aplastic anemia is discussed.

Microarray analysis of uterine gene expression in mouse and human pregnancy.

Improved care of infants born prematurely has increased their survival. However, the incidence of preterm labor has not changed. To understand the processes involved in preterm labor, we used oligonucleotide microarrays to study gene expression in murine and human uterus during pregnancy. The induction of enzymes for prostaglandin synthesis was used as a marker for important changes during pregnancy because prostaglandins strongly contribute to both human and murine labor. We identified 504 genes that changed at least 2-fold between d 13.5 and 19.0 in the gravid mouse uterus. In the pregnant human myometrium, we found 478 genes that changed at least 2-fold in either term or preterm labor compared with preterm nonlabor specimens and 77 genes that significantly varied in both preterm and term labor. Patterns of gene regulation within functional groups comparing human preterm and term labor were similar, although the magnitude of change often varied. Surprisingly, few genes that changed significantly throughout pregnancy were the same in the mouse and human. These data suggest that functional progesterone withdrawal in human myometrium may not be the primary mechanism for labor induction, may implicate similar mechanisms for idiopathic preterm and term labor in humans, and may identify novel targets for further study.

Insulin-like growth factor I receptor is expressed at normal levels in Nijmegen breakage syndrome cells.

Nijmegen breakage syndrome (NBS) is an autosomal recessive disorder sharing a pleiotropic phenotype with ataxia-telangiectasia (A-T), including increased radiosensitivity and cancer disposition. Insulin-like growth factor I receptor (IGF-IR) expression is reportedly decreased in A-T cells, which is thought to contribute to its increased radiosensitivity. In this study, we investigated whether the same mechanism underlies the radiosensitivity of NBS cells. GM7166VA7 cells lacking NBS1 protein displayed a phenotype of increased radiosensitivity, while the introduction of NBS1 cDNA conferred radioresistance comparable to normal cells. IGF-IR expression levels were essentially the same among normal, NBS, and NBS1-complemented NBS cells. There was no significant difference between NBS and NBS1-complemented cells in activation of major downstream pathways of IGF-IR upon IGF-I stimulation, including phosphatidylinositol-3(') kinase (PI3-K) and mitogen-activated protein kinase (MAPK). Collectively, IGF-IR-related events are unlikely to be disrupted in NBS cells, and therefore, defects in IGF-IR signaling do not explain the increased radiosensitivity of NBS cells.