IGF-I Overexpression causes fetal loss during placentation in mice

To understand the role of IGF-I in murine pregnancy, we studied the reproductive performance of IGF-I overexpressed mice. Fetal loss occurred only in the transfected uterine horn during day 10-15 of pregnancy. The placenta appeared healthy until Day 10 of pregnancy. From day 12, the decidua basalis of the transfected horn increased in thickness. The vascular lumen was expanded, and most of embryos were dead. Uterine natural killer cells did not undergo apoptosis from day 10 to day 15 when they usually go through apoptosis. Thus, it is likely that IGF-I plays a role in the decidual formation through regulation of uNK cells. This is the first report to demonstrate that IGF-I overexpression can cause fetal loss during murine placentation.     

Epizootic hemorrhagic disease virus infection of type I interferon receptor deficient mice

Type I interferon receptor deficient (IFNAR(-/-)) mice were infected with an Israeli isolate of serotype 7 of epizootic hemorrhagic disease virus (EHDV; Orbivirus, Reoviridae). Two out of two mice that received 5×10(5) 50% tissue culture infectious doses (TCID(50)) by intraperitoneal injection died or were euthanized in a moribund state on day 5 after infection. One mouse out of three that had been inoculated with 5×10(2) TCID(50) died on day 7 while the remaining mice did not show any clinical signs and survived until the end of the experiment. Spleens of all dead mice were highly positive in an EHDV real-time RT-PCR (quantification cycle values ≤15) and contained ≥10(5.8) TCID(50) of virus per ml of homogenate. The viral RNA content and virus titer in the spleens of the two surviving mice, on the other hand, were over 100-fold lower. Different from data reported for BTV, the outcome of EHDV infection of IFNAR(-/-) mice is dose-dependent and subclinical infections can occur.     

ConA induced suppressor cells in scrapie-infected mice

Lymphocyte suspensions prepared from the spleens of mice (LACG/COM) clinically affected with scrapie (ME7) were cultured for 48 hrs in the presence of Concanavalin (ConA) in order to induce suppressor cells. These cells behaved exactly as similarly prepared cells from normal age-matched mice in suppressing the response of fresh normal splenic lymphocytes to phytohaemagglutinin (PHA).     

Pulmonary lesions induced by Pasteurella haemolytica in neutrophil sufficient and neutrophil deficient calves.

The role of neutrophils in the development of peracute lung lesions of bovine pneumonic pasteurellosis was investigated. Eight calves were divided into two groups of four calves each. Group I was treated with intravenous phosphate-buffered saline and served as the neutrophil sufficient calves. Group II was treated with intravenous hydroxyurea which produced a state of neutropenia. When peripheral blood neutrophil numbers dropped below 300 cells/microL in group II, all calves were challenged with an intrabronchial bolus of Pasteurella haemolytica in the log phase of growth. An acute inflammatory process occurred in both groups of calves indicated by a rise in body temperature. While pulmonary lesions occurred in both groups by six hours postinoculation, they varied in pathological characteristics. Pulmonary lesions in the neutrophil sufficient calves consisted of fibrinopurulent alveolitis-bronchiolitis with associated alveolar septal necrosis, interlobular edema, and intravascular thrombi. The neutrophil deficient calves had extensive intra-alveolar edema, interlobular edema, intraalveolar hemorrhage, atelectasis, and focal areas of alveolar septal necrosis. These results show that P. haemolytica can induce severe pulmonary tissue damage through both neutrophil dependent and neutrophil independent mechanisms.     

Susceptibility of liver proliferative lesions in heterozygous p53 deficient CBA mice to various carcinogens

To investigate the liver tumorigenic sensitivity to various carcinogens in heterozygous p53 deficient [p53 (+/-)] CBA mice and their wild-type littermates [p53 (+/+) mice], 71 p53 (+/-) and 74 p53 (+/+) CBA mice (male, 6-12 weeks of age) were given diet containing 4,000 or 0 ppm flumequine (FL) for 26 weeks or a single intraperitoneal injection of 5 mg/kg body weights dimethylnitrosamine (DMN) at start of the study in Exp. 1, diet containing 6,000 or 0 ppm di(2-ethylhexyl)-phthalate (DEHP) for 26 weeks in Exp. 2, or diet containing 12,000, 6,000 or 0 ppm phenolphthalein (PhP) for 26 weeks in Exp. 3. All surviving animals of these groups were killed after completion of treatment of the test substances for 26 weeks. In the FL groups, the incidences of hepatocellular altered foci in p53 (+/-) mice, the multiplicities of those in p53 (+/-) and p53 (+/+) mice were significantly increased as compared to the corresponding control groups. The incidences and multiplicities of altered foci in the DMN groups were higher than those in the corresponding control groups in p53 (+/-) and p53 (+/+) mice, but no significant differences were indicated between the groups. There were no significant differences in the incidences, multiplicities and proliferating cell nuclear antigen labeling indices of altered foci in the FL or DMN groups between p53 (+/-) and p53 (+/+) mice. There were no significant differences in the incidences and multiplicities of altered foci between the DEHP or PhP and control groups. The present results suggest that p53 gene knocked out heterozygously does not enhance the chemical hepatocarcinogenesis in CBA mice.     

Antibody complement-dependent bacteriolysis in experimentally induced pasteurellosis in mice

Affinity-purified bovine immunoglobulin isotypes were bacteriolytic for Pasteurella haemolytica biotype A, serotype 1 (PHA-1). This bacteriolysis was specific and complement-dependent. The IgM and IgG1 were the most active isotypes in the classic complement cascade. These isotypes also induced bacteriolysis through the alternative complement cascade. The comparative bacteriolytic activities of IgG1 and IgM were equal within each cascade; however, the bacteriolytic activities of IgG1 and IgM were lower in the alternative cascade than in the classical cascade. The IgG2 was more bacteriolytic than IgA in the classic and alternative complement pathways. Bovine immunoglobulins passively protected C57BL/6 mice from experimentally induced pasteurellosis. There were no major differences in the protection among hyperimmune sera, purified IgM, or purified IgG. Mice were protected from PHA-1 by approximately 1.9 micrograms of IgG and 1.2 or 0.1 micrograms of IgM. Elimination of murine complement with cobra venom factor 3 reduced PHA-1 clearance in passively immunized C57BL/6 mice. The protective effect of IgM mediated resistance was highly dependent on an intact complement system. The intact complement cascade was associated with enhanced clearance of PHA-1 from the liver. Although PHA-1 was susceptible to antibody complement-mediated bacteriolysis in vitro, the dependence on an intact complement cascade was not absolute in experimentally induced murine septicemic pasteurellosis.     

LPS诱导小鼠子宫内膜炎模型的建立

为了建立小鼠子宫内膜炎模型,为子宫内膜炎的发病机理及其防治方法的研究提供合适的动物疾病模型,在小鼠子宫内灌注不同浓度的脂多糖(LPS),分别于灌注后12、24、36 h和48 h收集子宫,观察子宫组织形态学变化,检测子宫组织髓过氧化物酶(MPO)和一氧化氮(NO)的含量。结果在灌注LPS后可见小鼠子宫上皮细胞脱落、水肿、充血、出血和炎性细胞浸润。灌注1.25、2.5 mg/m L和5 mg/m L的LPS作用12 h,小鼠子宫组织MPO和NO含量极显著高于对照组(P0.01);灌注2.5 mg/m L和5 mg/m L的LPS作用24 h,小鼠子宫组织MPO和NO含量极显著高于对照组(P0.01)。表明采用子宫灌注LPS法成功建立小鼠子宫内膜炎模型,且每只小鼠灌注20μL浓度为2.5~5.0 mg/m L的LPS作用12~24 h为最佳的建模条件。     

Fate of conceptus and corpus luteum after induced embryonic loss in heifers

Ultrasonography was performed to determine the fate of the conceptus and the corpus luteum after induced embryonic loss in heifers. When embryonic loss was induced by administration of prostaglandin F2 alpha (25 mg, IM) on day 28 (n = 10) or day 42 (n = 10), embryonic death occurred 2.3 +/- 0.2 (mean +/- SEM) and 2.6 +/- 0.2 days, respectively, after treatment, and subsequent ovulation occurred 2.4 +/- 0.3 and 2.7 +/- 0.2 days, respectively, after embryonic death. Administration of a mitotic inhibitor (1 mg of colchicine, intrauterine; n = 4) on day 42 resulted in embryonic death in 1.2 +/- 0.2 days and ovulation in 20.8 +/- 3.1 days after embryonic death. Rupture of the amnion on day 42 (n = 3) resulted in immediate death of the embryo and ovulation in 35 +/- 3.5 days. In prostaglandin-treated heifers, area of the corpus luteum decreased (P less than 0.01) between days -2 and -1, relative to day of embryonic death. Volume of uterine contents remained approximately constant for 2 or 3 days after treatment, then decreased greater than 75% between examinations on consecutive days. Embryonic death apparently resulted from luteolysis and was characterized by rapid loss of the conceptus with minimal conceptus degeneration; an intact conceptus was recovered from the vagina of 3 heifers treated on day 42. In heifers treated with colchicine or in which the amnion was ruptured, embryonic death was associated with maintenance of the corpus luteum and prolonged retention of the conceptus, which underwent extensive degeneration.(ABSTRACT TRUNCATED AT 250 WORDS)     

Fetal loss and hyposulfataemia in pregnant NaS1 transporter null mice

Sulfate is important for growth and development, and is supplied from mother to fetus throughout pregnancy. We used NaS1 sulfate transporter null (Nas1(-/-)) mice to investigate the role of NaS1 in maintaining sulfate homeostasis during pregnancy and to determine the physiological consequences of maternal hyposulfataemia on fetal, placental and postnatal growth. We show that maternal serum (≤0.5 mM), fetal serum (<0.1 mM) and amniotic fluid (≤0.5 mM) sulfate levels were significantly lower in pregnant Nas1(-/-) mice when compared with maternal serum (?2.0 mM), fetal serum (?1.5 mM) and amniotic fluid (?1.7 mM) sulfate levels in pregnant Nas1(+/+) mice. After 12 days of pregnancy, fetal reabsorptions led to markedly reduced (by ≥50%) fetal numbers in Nas1(-/-) mice. Placental labyrinth and spongiotrophoblast layers were increased (by ?140%) in pregnant Nas1(-/-) mice when compared to pregnant Nas1(+/+) mice. Birth weights of progeny from female Nas1(-/-) mice were increased (by ?7%) when compared to progeny of Nas1(+/+) mice. These findings show that NaS1 is essential to maintain high maternal and fetal sulfate levels, which is important for maintaining pregnancy, placental development and normal birth weight.     

Nutritionally induced body weight loss and ovarian quiescence in Shiba goats

Four female Shiba goats were used to determine the influence of body weight loss by dietary restriction on estrous cyclicity. The dietary restriction was started on the day following ovulation. The goats were fed hay cube and straw at an amount of 30% of energy requirement based on weekly body weight measurement. The ovaries were monitored daily by transrectal ultrasonography and blood samples were collected daily by jugular venipuncture for ovarian steroids analysis. After the start of food restriction, all animals lost body weight and entered ovarian quiescence. Intervals to the onset of ovarian quiescence tended to depend on the body weight of each animal at the start of food restriction. The mean concentration of progesterone during the mid-luteal phase (from 7 to 13 days after ovulation) in the last estrous cycle before ovarian quiescence was significantly lower than that in normal estrous cycle of the control period (19.7 +/- 2.8 vs 12.3 +/- 2.2 ng/ml, P<0.05), whereas there was no significant difference in the length of the luteal phase, determined as the period when corpora lutea existed and concentrations of progesterone were equal to or greater than 1 ng/ml (15.8 +/- 1.5 vs 15.0 +/- 2.8 days, P>0.1). A rise of estradiol concentration and follicular growth in the follicular phase following a decline of progesterone level after luteal regression tended to be suppressed at the onset of ovarian quiescence. It seems that the present results are consistent with previous findings that nutritionally induced body weight loss influences the secretion of ovarian steroids and eventually induces ovarian quiescence.     

GeneScanning analysis of Ig/TCR gene rearrangements to detect clonality in canine lymphomas

The diagnosis of canine lymphoma is achieved using morphological and immunological methods. In a certain percentage of cases, difficulties in making a definitive diagnosis of lymphoproliferative disorders may occur despite extensive immunophenotyping. Therefore, additional diagnostics, such as molecular assessment of Ig/TCR gene rearrangements clonality, may confirm the final diagnosis. Polyacrylamide gel electrophoresis and heteroduplex analysis have already been proven to be suitable for detecting clonality but are cumbersome and labor-intensive. In the present study, GeneScanning analysis of PCR products originating from different primer sets targeting different regions of Ig and TCR was validated in improving sensitivity as well as in reducing the turnaround time of gene rearrangement assays. GeneScanning exploits 5' fluorescently labelled primers for the automated and fast analysis of PCR products either as singleplex or multiplex runs. Initially, the assay was set up using DNA purified from normal tissues (n=6), hyperplastic/reactive tissues (n=10) and a small set of immunophenotyped lymphoma samples (n=12). The optimized methods were then used in a large set of 96 canine lymphoma samples. Normal and hyperplastic/reactive lymphoid tissues showed typically polyclonal or, occasionally, oligoclonal PCR products. Lymphoma samples showed monoclonal peaks arranged as a single or, occasionally, a double narrow base peak sometimes embedded in a polyclonal background. In all immunophenotyped cases, an Ig or TCR clonal finding corresponded to B- and T-cell lymphomas, respectively. Overall, 94/96 (97.9%) samples showed clonal Ig/TCR clonal rearrangements among which clonal Ig was found in 61/96 (63.5%) of samples and clonal TCR in 33/35 Ig negative samples (34.4% of all cases). In one out of ten randomly chosen cases, both Ig and TCR clonal gene rearrangements were found. Among the factors affecting assay accuracy, DNA quality has been shown to be critical and the amplification of DNA controls of different size are recommended to evaluate DNA integrity. Frozen material such as that which remained inside the hub of the needle used for diagnostic procedures is optimal for the analysis herein described. In conclusion, GeneScanning represents a versatile tool for routinely assessing Ig/TCR clonal rearrangements and supporting the diagnostic protocol of canine lymphomas.     

Immunosuppression in deer mice with experimentally induced trypanosomiasis.

Light and electron microscopic examinations of deer mice (Peromyscus maniculatus) chronically infected with Trypanosoma equiperdum revealed hyperplasia of germinal center lymphocytes (germanocytes) in the lymph follicles of spleen and lymph nodes and infiltration of the splenic red pulp cords and nodal medullary cords with plasma cells. Proliferation and infiltration of plasma cells caused disruption of the B- and T-lymphocyte areas in these organs. Stimulation of splenic lymphocytes in vitro by phytohemagglutinin and concanavalin A revealed marked depression in T-lymphocyte response; stimulation with lipopolysaccharide and pokeweed mitogens showed depression of B-cell response. Deer mice infected with virulent trypanosomes had decreased immunologic response to injection of sheep red blood cells, whereas deer mice given radioattenuated trypanosomes had normal to enhanced immunologic response to injection of sheep red blood cells.     

Engraftment of severe combined immune deficient/beige mice with bovine foetal lymphoid tissues.

To develop a model of bovine thymus and lymph node growth in vivo, we have implanted bovine foetal tissues (16-23 weeks gestation) under the renal capsule of severe combined immune deficient (SCID)/beige (BG) mice and assayed for graft growth and characteristics 2-18 weeks after engraftment. Bovine foetal thymus and lymph node grew considerably following engraftment of SCID/BG mice. Growth was optimal if bovine foetal tissues were used before gestation Week 17. Bovine-mouse chimerism was confirmed using glucose phosphate isomerase analysis. Bovine thymus grew during the entire 18 weeks of study. Growth of bovine lymph node was initially rapid, reaching a maximum at 2 weeks after transplantation followed by a progressive decrease in size. Transplanted bovine lymph node and thymus were morphologically similar to age-matched bovine foetal tissue for a limited time period. Fibrosis, degeneration and depletion of lymphocytes were evident 6 weeks after engraftment; changes were more severe in lymph node than in thymus whereas increases in lymphocytes, lymphopoiesis and follicle formation were evident in age-matched bovine foetal tissue. Despite growth and morphological similarities of the transplanted tissue, blood counts suggested there was no peripheralization of bovine leucocytes. Bovine immunoglobulins (IgG1 and IgG2) were detected in serum of some SCID/BG chimeric mice for a limited time. The appearance of bovine immunoglobulins at 2 weeks in SCID/BG chimeric mice depended on the age of the foetal donor (> 18 weeks) and coincided with the appearance of morphologically mature lymphocytes in the donor foetus lymph nodes. The ability to produce bovine immunoglobulins decreased 8 weeks after engraftment, coinciding with the depletion of lymphocytes in the engrafted lymph node. Lymphocyte depletion and loss of function of engrafted tissues appear the result of a lack of lymphoid progenitors normally derived from hematopoietic stem cells in the bone marrow.     

Effects of novel vaccines on weight loss in diet-induced-obese (DIO) mice

The purpose of the study was to test the therapeutic effects of novel vaccines for reducing weight gain and increasing weight loss in diet induced obesity (DIO) model. Male C57BL/6 J mice, fed a 60% Kcal fat diet for 8 weeks prior to the start of the study, were vaccinated via the intraperitoneal route with two formulations (JH17 & JH18) of chimeric-somatostatin vaccines at 1 and 22 days of the study. Control mice were injected with PBS. All mice continued to be feed the 60% Kcal fat diet for the 6 week study. Body weights were measured two times a week and food intake was measured weekly. At week 6, mice were euthanized and a terminal bleed was made and antibody levels to somatostatin and levels of insulin-like growth factor 1 (IGF-1) were determined. Vaccination with both vaccine formulations induced a statistically significant body weight change over the study period, as compared with PBS controls. Percentage of baseline body weight was also significantly affected by vaccination during the study period. Vaccinates finished the study at 104% and 107% of baseline weight, JH17 & JH18 respectively, while untreated controls reached 115% of baseline weight. Food intake per mouse was similar in all mouse groups during the entire study. Control mice did not demonstrate any antibody titers to somatostatin, while all vaccinated mice had measurable antibody responses (> 1:500,000 titer). IGF-1 levels were not statistically significant among the groups, but were elevated in the JH18 vaccinates (mean 440.4 ng/mL) when compared with PBS controls (mean 365.6 ng/mL). Vaccination with either JH17 or JH18 chimeric –somatostatin vaccines produced a statistically significant weight loss as compared with PBS controls (P < 0.0001), even though the DIO mice with continually fed a 60% Kcal fat diet. The weight loss/lower weight gain observations were even more significant, as all mice consumed similar amounts of food for the entire study. The presence of high levels of anti-somatostatin antibodies at 6 weeks was correlative with the weight observations and confirmed the success of vaccination.     

Ultrasonographic study of early embryonic loss induced by Actinomyces pyogenes in cattle.

Four cows were inoculated into the uterus with Actinomyces pyogenes between 30 and 41 days of gestation. Gross morphological changes were monitored by rectal palpation and with a realtime B-mode two-dimensional scanner with a 7.5 MHz transrectal linear transducer, shortly before infection and afterwards at three to 12 hours intervals. Two control groups of cows 27 to 50 days pregnant were used: two cows were inoculated with 6 ml of sterile saline into the uterine lumen and four cows were treated with cloprostenol (prostaglandin F2 alpha analogue). There was a change in the uterine fluid from a black, non-echogenic image before infection to a grey or cloudy echogenic image as early as 21 hours after infection. There was an increase in the thickness of the endometrium of the horns and body of the uterus. The embryonic membranes thickened and separated from the endometrium as early as four hours after infection, followed by cessation of the embryonic heart beat, opening of the cervix and abortion. Abortion was followed by an increase in the amount of echogenic intrauterine fluid leading to an increase in the size of the uterus, and the cervix remained open for at least eight days. The area of the corpus luteum remained greater than 2 cm2 throughout the whole period. Intrauterine inoculation with sterile saline had no effects, but the administration of cloprostenol was followed by the death of the embryo and abortion within 72 hours, and the regression of the corpus luteum from greater than 2 cm2 at treatment to 0.4 cm2, 24 hours after abortion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of novel vaccines on weight loss in diet-induced-obese （DIO） mice

The purpose of the study was to test the therapeutic effects of novel vaccines for reducing weight gain and increasing weight loss in diet induced obesity (DIO) model. Male C57BL/6 J mice, fed a 60% Kcal fat diet for 8 weeks prior to the start of the study, were vaccinated via the intraperitoneal route with two formulations (JH17 & JH18) of chimeric-somatostatin vaccines at 1 and 22 days of the study. Control mice were injected with PBS. All mice continued to be feed the 60% Kcal fat diet for the 6 week study. Body weights were measured two times a week and food intake was measured weekly. At week 6, mice were euthanized and a terminal bleed was made and antibody levels to somatostatin and levels of insulin-like growth factor 1 (IGF-1) were determined. Vaccination with both vaccine formulations induced a statistically significant body weight change over the study period, as compared with PBS controls. Percentage of baseline body weight was also significantly affected by vaccination during the study period. Vaccinates finished the study at 104% and 107% of baseline weight, JH17 & JH18 respectively, while untreated controls reached 115% of baseline weight. Food intake per mouse was similar in all mouse groups during the entire study. Control mice did not demonstrate any antibody titers to somatostatin, while all vaccinated mice had measurable antibody responses (> 1:500,000 titer). IGF-1 levels were not statistically significant among the groups, but were elevated in the JH18 vaccinates (mean 440.4 ng/mL) when compared with PBS controls (mean 365.6 ng/mL). Vaccination with either JH17 or JH18 chimeric-somatostatin vaccines produced a statistically significant weight loss as compared with PBS controls (P < 0.0001), even though the DIO mice with continually fed a 60% Kcal fat diet. The weight loss/lower weight gain observations were even more significant, as all mice consumed similar amounts of food for the entire study. The presence of high levels of anti-somatostatin antibodies at 6 weeks was correlative with the weight observations and confirmed the success of vaccination.     

Uncoupling proteins and energy expenditure in mice divergently selected for heat loss

The objective of this study was to determine whether variation in energy expenditure created by selection on heat loss is mediated by uncoupling protein-1 (UCP1) in brown adipose tissue. Divergent selection for heat loss developed lines of mice with high (MH) and low (ML) maintenance energy expenditure. Concentration of UCP1 mRNA in brown adipose tissue (BAT) was 93% greater in ML than in MH mice (P < 0.02). Two new lines of mice, KH and KL, were bred by backcrossing a UCP1 knockout gene into the MH and ML lines, respectively; KH and KL with both knock-out (-/-) and wild type (+/+) UCP1 genotypes were generated. At 13 wk of age, KH mice exhibited greater heat loss (166 kcal x kg(0.75) x d(-1)) than KL mice (126.4 kcalkg(0.75) x d(-1)) regardless of the UCP1 knockout (P < 0.0001). Concentration of UCP2 mRNA in BAT was 74% greater in UCP1 knockout mice (-/-) than in wild type (+/+; P = 0.0001). We conclude that response to selection for increased energy expenditure was not mediated by increased expression or function of UCP1.     

Genetic analysis of litter size in mice

We performed quantitative trait locus (QTL) mapping analysis for litter size (total number of pups born and/or number of pups born alive) in 255 backcross mice derived from C57BL/6J and RR/Sgn inbred mice. We identified one significant QTL on chromosome 7 and 4 suggestive QTLs on chromosomes 3, 5, 10 and 13. In addition, two suggestive QTLs were identified on chromosomes 1 and 4 for the number of stillbirth. These results suggested that both litter size and number of stillbirth were heritable traits, although they were controlled by distinct genes. The RR allele was associated with reduced litter size and increased stillbirth at all QTLs. Therefore, RR mothers were observed to have reduced prolificacy in this particular genetic cross.