Immunosuppressive activity of uterine luminal protein from steroid-treated ovariectomized heifers

Uterine luminal protein (ULP) collected from ovariectomized steroid-treated crossbred heifers was tested for immunosuppressive activity in vitro. Heifers were allotted to treatment groups and for 16 d received daily injections of the following steroids or vehicle: Control (C, corn oil only, n=10); estradiol-17beta (E(2), 1.1 mug/kg body wt, n=10); progesterone (P(4), 2.2 mg/kg body wt, n=10); and E(2)+P(4) (1.1 mug + 2.2 mg/kg body wt, n=9). On Day 17, uterine flushings were collected, concentrated and quantitated for total ULP. ULP was tested for suppression of lymphocyte blastogenesis. For each experiment, 5 x 10(5) bovine lymphocytes were incubated with 0.4 mug of phytohemagglutinin (PHA) and ULP (25 to 400 mug ULP/ml) using standard culture conditions. At 48 h, 0.5 muCi of (3H) thymidine was added to cultures with cells harvested at 60 +/- 1 h by automation. Incorporated thymidine was measured by scintillation chromatography. Mean total ULP values for C-, E(2)-, P(4)- and E(2)+P(4)-treated groups were 4.7, 8.4, 13.6, and 25.5 mg, respectively (E(2)+P(4)>C and E(2), P<0.05). ULP from all treatment groups suppressed (P<0.0001) lymphocyte blastogenesis (thymidine incorporation) to PHA; however, suppression was greater (P<0.0001) for ULP from E(2)- and P(4)-than C-treated heifers at 100 and 200 mug ULP/ml. In conclusion, E(2) and P(4) injections enhanced immunosuppressive activity of ULP secretions.     

Temporal patterns of secretion of porcine uterine suppressor and stimulatory macromolecules

Temporal secretory patterns of porcine uterine suppressor (>/=230 kD) and stimulatory (29 kD) macromolecules were evaluated within uterine luminal protein (ULP) secretions recovered during early pregnancy. The ULP was recovered by uterine flushing from four Landrace gilts each on Days 9, 12, 15 and 18 of pregnancy. Unfractionated and fractionated ULP (using Sephacryl S-200) was tested for suppression or stimulation of phytohemagglutinin-induced peripheral blood T-lymphocyte proliferation. For all days of pregnancy, unfractionated ULP suppressed (P<0.002 to 0.0001) lymphocyte proliferative responses, with the greatest (P<0.05) activity observed for ULP collected on Day 9 of pregnancy. Suppressor activity resulted from the >/=230 kD component, in which the activity was greater (P<0.05) for ULP from gilts on Day 9 than Days 12, 15 and 18 of pregnancy. The 29 kD component failed (P>0.05) to stimulate lymphocyte proliferation, although there was a nonsignificant stimulatory trend for 2 of 4 gilts each at Days 12 and 15 of pregnancy. These findings demonstrate a temporal secretory pattern for the >/=230 kD lymphocyte suppressor component, which may be requisite for the immunological survival of the conceptus during early pregnancy. The inconsistent appearance of the lymphocyte stimulatory factor (29 kD component) tends to minimize its biological significance relative to the immunology of pregnancy.     

Integrating Ecology and Economics for Restoration: Using Ecological Indicators in Valuation of Ecosystem Services

Because it can uniquely furnish insights into nonuse values for ecosystem services, survey‐based Stated Preference (SP) valuation is widely used to estimate the benefits of ecological restoration. SP surveys ask respondents to select among restoration options yielding different ecological outcomes. This review examines the representation of ecological outcomes in SP studies seeking to quantify values for restoration of aquatic ecosystems. To promote the validity of ecological indicators used in SP valuation, we identified four standards: indicators should be measurable, interpretable, applicable, and comprehensive. We reviewed recent SP studies estimating the value of aquatic ecosystem services to assess whether ecological indicators in current use had these desirable properties. More than half of the 54 indicators reviewed were measurable, meaning referable to potentially precise quantification. About one‐third were interpretable, that is, presented in a way that facilitates understanding the effects of restoration. About three quarters of the indicators were applicable; SP valuation practitioners typically consult with natural scientists to ensure that indicators represent the effect of stressors on ecological systems and with focus groups to ensure that indicators have a connection with ecosystem services that contribute to public well‐being. While most of the SP studies employed diverse and potentially comprehensive indicators that could capture direct and indirect effects of restoration, and 6 of 20 studies used indicators that met all standards, shortcomings in the indicators were common. These problems can be rectified with attention to how natural scientists measure change and to relationships between restoration outcomes and characteristics of fully restored reference ecosystems.     

The Arabidopsis thaliana ortholog of a purported maize cholinesterase gene encodes a GDSL-lipase

Acetylcholinesterase is an enzyme that is intimately associated with regulation of synaptic transmission in the cholinergic nervous system and in neuromuscular junctions of animals. However the presence of cholinesterase activity has been described also in non-metazoan organisms such as slime molds, fungi and plants. More recently, a gene purportedly encoding for acetylcholinesterase was cloned from maize. We have cloned the Arabidopsis thaliana homolog of the Zea mays gene, At3g26430, and studied its biochemical properties. Our results indicate that the protein encoded by the gene exhibited lipase activity with preference to long chain substrates but did not hydrolyze choline esters. The At3g26430 protein belongs to the SGNH clan of serine hydrolases, and more specifically to the GDS(L) lipase family.     

Serum and testis selenium concentrations and testis morphology in ram lambs of varying ages

Serum and testis selenium (Se) concentrations, body and testes weights, seminiferous tubule height and width measurements and percent of tubules containing luminal spermatozoa were determined in Se-treated (SSe) and control (NSe) crossbred ram lambs at 60, 90, 120, 150 and 180 days of age. With IM injections, SSe lambs received 3 mg of Se as selenite and NSe lambs received 0.9% saline at 30-day intervals throughout the study. For each age group, lambs were weighed, jugular vein blood collected and testes removed at the designated age. Serum and testis tissue samples for each lamb were assayed for Se, and testis tissue was also evaluated for histological parameters. For all parameters, only serum Se concentrations were affected (P<0.0001) by Se treatment; however, all other parameters were affected (P<0.0001) by age. For combined groups, mean testis Se concentration (0.33 ppm), testes weights, seminiferous tubule measurements and percent of tubules (82.2) containing luminal spermatozoa were greatest (P<0.05) at 180 days of age, and mean testis Se concentrations were significantly correlated with these testicular parameters. These data lend support to the hypothesis that the increase in concentration of testicular Se to adult concentrations (>0.3 ppm) around the time of puberty is associated with rapid testicular development and production of spermatozoa.     

Interaction of bovine uterine luminal protein with interleukin-2 and the interleukin-2 receptor of T lymphocytes

Bovine uterine luminal proteins (ULP) collected on Day 17 of pregnancy were tested for inhibition of binding of interleukin-2 (IL-2) to the IL-2 receptor (IL-2R) of bovine (CLC) and human (HLC) T lymphocytes and for binding to IL-2. Additional experiments assessed IL-2 binding to the p55 alpha chain (Tac protein) of the IL-2R of HLC. High- and low-molecular weight (Mr) ULP components (H-ULP greater than 248,000 Mr and L-ULP 21,000 Mr, respectively) inhibited (p less than 0.05 and 0.01, respectively) the binding of 125I-IL-2 to the IL-2R of CLC, whereas only H-ULP inhibited (p less than 0.05) binding to the IL-2R (presumably, the p75 beta chain) of HLC. H-ULP failed (p greater than 0.05) to bind to the p55 alpha chain of the IL-2R of HLC. For IL-2 binding, L-ULP failed (p greater than 0.05) to bind 125I-IL-2 in short (2 h)-term and long (45 h)-term experiments, whereas binding was evident (p less than 0.05) for H-ULP at 2 h of incubation. For H-ULP, mean (+/- SEM) percentages for bound and unbound 125I-IL-2 were 70.1 +/- 11.4 and 29.9 +/- 11.4, respectively. Further purification of H-ULP yielded a component (1.76 x 10(6) Mr) that bound 11.7% of 125I-IL-2 and inhibited (p less than 0.01) thymidine uptake and binding of 125I-IL-2 to the IL-2R of CLC. H-ULP-mediated suppression of lymphocyte proliferation may result from blocking IL-2R recognition of IL-2 as well as binding to IL-2, whereas suppression by L-ULP may predominantly result from blocking IL-2R.     

Selenium status and phytohemagglutinin-stimulated T lymphocytes from beef calves

The effect of selenium (Se), supplemented by subcutaneous injections, upon in vitro phytohemagglutinin (PHA)-stimulated T lymphocyte blastogenesis was evaluated in crossbred calves at 45, 124, and 270 d of age. Peripheral blood lymphocytes (5×10⁵) obtained from control (n=4) and Se-treated (n=4) calves were cultured with 0, 0.4, and 0.8 μg PHA and 0.5 μCi [³H]-thymidine in a complete culture medium. At 60h, cultures were harvested for the quantitation of thymidine incorporation. The degree of lymphocyte blastogenesis or stimulation to PHA, expressed as the stimulation index (SI), was assessed for treatment and age effects. At the onset and end of the experimental period, plasma Se and blood glutathione peroxidase concentrations revealed that control calves [0.045 ppm and 48.4 units/g hemoglobin (Hb), respectively] were marginally deficient and treated calves (0.075 ppm and 143.0 units/g Hb, respectively) were adequate in Se status. The SI values were not influenced (P>0.01) by Se treatment, but were affected (P<0.003) by age. For combined treatments and PHA dosages, mean SI values were greater (P<0.05), when lymphocytes were tested at 45 (SI=25.9) than at 124 and 270 (SI=13.7 and 15.5, respectively) d of age. Lymphocyte viability and number were not affected by Se treatment, but lymphocyte number tended (P<0.1) to increase with age. In conclusion, T lymphocyte responsiveness to mitogen stimulation was normal in beef calves residing in a geographical area considered deficient in Se.     

Characterization of immunosuppressive substances in the basic protein fraction of uterine secretions from pregnant ewes

The basic protein fraction of ovine uterine secretions collected late in pregnancy (Days 125-140) contains a substance capable of inhibiting in vitro blastogenic responses of lymphocytes to phytohemagglutinin (PHA) or mixed lymphocyte reactions. In this study, the immunosuppressive substance in the basic protein fraction of uterine secretions was further defined by gel filtration. The immunosuppressive activity resided in a group of high molecular weight proteins eluting at the void volume of Sephacryl S-200 and Sepharose CL-6B columns. For example, incorporation of thymidine by PHA-stimulated lymphocytes incubated with 20, 40, 80, and 120 micrograms/ml of protein from the void volume of Sepharose CL-6B was 65, 28, 2, and 0 percent of control lymphocytes, respectively. Based on polyacrylamide-gel electrophoresis in the presence of sodium dodeylsulfate (SDS), the immunosuppressive fraction from Sepharose CL-6B chromatography contained aggregates of uterine milk proteins (UTM-proteins) and a pair of proteins running at the top of a 5% (w/v) polyacrylamide gel. Other protein peaks resolved by Sephacryl S-200 and Sepharose CL-6B contained aggregates of UTM-proteins but were not immunosuppressive. The substance inhibiting in vitro lymphocyte function was not of conceptus origin, because it was found in fluid from the ligated uterine horn of unilaterally pregnant ewes and from the uterus of an ovariectomized ewe treated for 60 days with progesterone and estrone.