The aim of this study was to evaluate the pattern of protein expression of the steroid receptor isoforms of nuclear progesterone receptors (PGR) A and B, and estrogen receptors (ESR1 and 2) in utero-placental compartments during early pregnancy. Utero-placental tissues were collected from days 14-30 (n = 4 ewes/day), and uterine tissues were collected from non-pregnant ewes on day 10 after estrus (n = 4). Cross sections of formalin-fixed and paraffin embedded tissues were immunofluorescently stained to detect PGRAB, PGRB, ESR1 and ESR2, followed by image generation of entire cross-sections of uterine and utero-placental tissues, confocal imaging of individual uterine and utero-placental compartments, and image and statistical analyses. PGRAB, PGRB, ESR1 and ESR2 were detected in several compartments of uterine and utero-placental tissues. Quantitative image analysis of staining intensity demonstrated that compared to non-pregnant controls 1) expression of PGRAB and PGRB was less in luminal epithelium and endometrial glands from day 14-16 till 30; 2) PGRAB expression tended to be greater in endometrial and myometrial blood vessels on days 28 and/or 30; 3) PGRB expression in myometrum was lower on days 16 and 28; 4) ESR1 in endometrial stroma was lower in all days of pregnancy; 5) ESR2 expression was similar in all compartments and not affected by pregnancy stage; and 6) in FM, expression of steroid receptors was similar. Thus, we have demonstrated spatial and temporal expression of nuclear PGR and ESR isoforms in utero-placental compartments during early pregnancy.
Bacillus thuringiensis (Bt) being an eco-friendly bioinsecticide is effectively used in pest management strategies and, therefore, isolation and identification of new strains effective against a broad range of target pests is important. In the present study, new indigenous B. thuringiensis strains were isolated and investigated so that these could be used as an alternative and/or support the current commercial strains/cry proteins in use. For this, 159 samples including soil, leaf and spider webs were collected from ten districts of Kashmir valley (India). Of 1447 bacterial strains screened, 68 Bt strains were identified with 4 types of crystalline inclusions. Crystal morphology ranking among the Bt strains was spherical (69.11%) [ spore attached (8.82%) [ rod (5.88%) = bipyramidal (5.88%) [ spherical plus rod (4.41%) [ spherical plus bipyramidal (2.94%) = cuboidal (2.94%). SDS-PAGE investigation of the spore-crystal mixture demonstrated Bt strains contained proteins of various molecular weights ranging from 150 to 28 kDa. Insecticidal activity of the 68 indigenous Bt strains against Spodoptera litura neonates showed that Bt strain SWK1 strain had the highest mortality. Lepidopteron active genes (cry1, cry2Ab, cry2Ab) were present in six Bt strains. Further, analysis of a full-length cry2A gene (*1.9 kb) by PCR-RFLP in strain SWK1 revealed that it was a new cry2A gene in Bt strain SWK1 and was named as cry2Al1 (GenBank Accession No. KJ149819.1) using the Bt toxin nomenclature (http://www.btnomenclature. info). Insect bioassays with neonate larvae of S. litura and H. armigera showed that the purified Cry2Al1 is toxic to S. litura with LC 50 2.448 lg/ml and H. armigera with LC 50 3.374 lg/ml, respectively. However, it did not produce any mortality in third instar larvae of Aedes aegypti, Culex quinquefasciatus and Anopheles stephensi larvae/pupae insects (100 lg/ml) at 28 ± 2°C and 75 to 85% relative humidity under a photoperiod of 14L:10D.
BackgroundObjectives were to examine the effects of selenium (Se) supply and maternal nutritional plane during gestation on mammary gland growth, cellular proliferation, and vascularity at parturition and d 20 of lactation. Rambouillet primiparous ewes (n = 84) were allocated to treatments in a 2 x 3 factorial. Factors were dietary Se (adequate Se [ASe, 11.5 μg/kg BW] or high Se [HSe, 77.0 μg/kg BW]) and nutritional plane (60% [RES], 100% [CON], or 140% [EXC]). At parturition, lambs were removed and 42 ewes (7/treatment) were necropsied. Remaining ewes were fed a common diet meeting requirements for lactation and mechanically milked twice daily until necropsy on d 20. At both necropsy periods, mammary glands were dissected and tissues harvested. Samples were analyzed for RNA, DNA, and protein content, cell proliferation, and vascularity. Where interactions were present (P ≤ 0.05), least squares means from the highest-order interaction are presented.ResultsFinal body weight of ewes was least (P ≤ 0.002) in RES, intermediate for CON, and greatest for EXC, regardless of stage of the ewe at necropsy (parturition or d 20 of lactation). In ewes necropsied at parturition, mammary glands were heavier (P = 0.02) in EXC compared to RES, with CON intermediate. Concentration of RNA (mg/g) was decreased (P = 0.01) in EXC compared to CON at parturition. There was a tendency (P = 0.07) for a Se by nutrition interaction in percentage of cells proliferating where ASe-EXC ewes had greater (P ≤ 0.02) number of proliferating cells then all other treatments. Mammary vascular area tended (P = 0.08) to be affected by a Se by nutrition interaction where ASe-CON had less (P = 0.007) vascular area than HSe-CON ewes. In ewes necropsied at d 20 of lactation, the number of alveoli per area was decreased (P ≤ 0.05) in RES compared to CON and EXC-fed ewes.ConclusionsResults of this study indicate that proper maternal nutritional plane during gestation is important for mammary gland development, even out to d 20 of lactation.
Sex steroid hormones are major regulators of uterine and placental growth and functions, as well as many other biological processes. To examine the mRNA expression of nuclear estrogen (ESR1 and 2) and progesterone (PGRAB and B) receptors in different compartments of the uterus and placenta, tissues were collected in experiment 1, on days 16, 20 and 28 after natural mating (NAT) and on day 10 after estrus (non-pregnant controls [NP]); and in experiment 2, on day 22 of NAT, and pregnancies established after transfer of embryos generated through mating of FSH-treated ewes (NAT-ET), in vitro fertilization (IVF), or in vitro activation (IVA; parthenotes). In experiment 1, ESR1 expression in endometrial stroma (ES), endometrial glands (EG) and myometrial blood vessels (MBV), ESR2 in endometrial blood vessels (EBV), PGRAB in ES, and PGRB in ES, EG and MBV was greater in pregnant than NP depending on day of pregnancy. Day of pregnancy affected expression of ESR1 in MBV, ESR2 in EBV and MBV, and PGRAB in ES. In experiment 2, ESR1, PGRAB and PGRB in EG, but not in other compartments, was greater in NAT-ET than NAT, and PGRB was greater in NAT-ET than the IVF. These data demonstrated that the ESR and PGR expression was different in pregnant vs. NP ewes in selected compartments, and was affected by pregnancy stage or embryo origin in selected utero-placental compartments. Thus, sex steroid hormone mRNA expression is differentially regulated in a spatio-temporal manner in uterus and placenta, and is affected by application of assisted reproductive technology in sheep.
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