Soil legacy effects of plants and drought on aboveground insects in native and range-expanding plant communities

Soils contain biotic and abiotic legacies of previous conditions that may influence plant community biomass and associated aboveground biodiversity. However, little is known about the relative strengths and interactions of the various belowground legacies on aboveground plant–insect interactions. We used an outdoor mesocosm experiment to investigate the belowground legacy effects of range-expanding versus native plants, extreme drought and their interactions on plants, aphids and pollinators. We show that plant biomass was influenced more strongly by the previous plant community than by the previous summer drought. Plant communities consisted of four congeneric pairs of natives and range expanders, and their responses were not unanimous. Legacy effects affected the abundance of aphids more strongly than pollinators. We conclude that legacies can be contained as soil ‘memories’ that influence aboveground plant community interactions in the next growing season. These soil-borne ‘memories’ can be altered by climate warming-induced plant range shifts and extreme drought.     

Notch1 mediates uterine stromal differentiation and is critical for complete decidualization in the mouse

Uterine receptivity implies a dialogue between the hormonally primed maternal endometrium and the free-floating blastocyst. Endometrial stromal cells proliferate, avert apoptosis, and undergo decidualization in preparation for implantation; however, the molecular mechanisms that underlie differentiation into the decidual phenotype remain largely undefined. The Notch family of transmembrane receptors transduce extracellular signals responsible for cell survival, cell-to-cell communication, and differentiation, all fundamental processes for decidualization and pregnancy. Using a murine artificial decidualization model, pharmacological inhibition of Notch signaling by γ-secretase inhibition resulted in a significantly decreased deciduoma. Furthermore, a progesterone receptor (PR)-Cre Notch1 bigenic (Notch1(d/d)) confirmed a Notch1-dependent hypomorphic decidual phenotype. Microarray and pathway analysis, following Notch1 ablation, demonstrated significantly altered signaling repertoire. Concomitantly, hierarchical clustering demonstrated Notch1-dependent differences in gene expression. Uteri deprived of Notch1 signaling demonstrated decreased cellular proliferation; namely, reduced proliferation-specific antigen, Ki67, altered p21, cdk6, and cyclinD activity and an increased apoptotic-profile, cleaved caspase-3, Bad, and attenuated Bcl2. The results demonstrate that the preimplantation uterus relies on Notch signaling to inhibit apoptosis of stromal fibroblasts and regulate cell cycle progression, which together promotes successful decidualization. In summary, Notch1 signaling modulates multiple signaling mechanisms crucial for decidualization and these studies provide additional perspectives to the coordination of multiple signaling modalities required during decidualization.     

Inhibition of fibroblast growth by Notch1 signaling is mediated by induction of Wnt11-dependent WISP-1

Fibroblasts are an integral component of stroma and important source of growth factors and extracellular matrix (ECM). They play a prominent role in maintaining tissue homeostasis and in wound healing and tumor growth. Notch signaling regulates biological function in a variety of cells. To elucidate the physiological function of Notch signaling in fibroblasts, we ablated Notch1 in mouse (Notch1(Flox/Flox)) embryonic fibroblasts (MEFs). Notch1-deficient (Notch1(-/-)) MEFs displayed faster growth and motility rate compared to Notch1(Flox/Flox) MEFs. Such phenotypic changes, however, were reversible by reconstitution of Notch1 activation via overexpression of the intracellular domain of Notch1 (NICD1) in Notch1-deficient MEFs. In contrast, constitutive activation of Notch1 signaling by introducing NICD1 into primary human dermal fibroblasts (FF2441), which caused pan-Notch activation, inhibited cell growth and motility, whereas cellular inhibition was relievable when the Notch activation was countered with dominant-negative mutant of Master-mind like 1 (DN-MAML-1). Functionally, "Notch-activated" stromal fibroblasts could inhibit tumor cell growth/invasion. Moreover, Notch activation induced expression of Wnt-induced secreted proteins-1 (WISP-1/CCN4) in FF2441 cells while deletion of Notch1 in MEFs resulted in an opposite effect. Notably, WISP-1 suppressed fibroblast proliferation, and was responsible for mediating Notch1's inhibitory effect since siRNA-mediated blockade of WISP-1 expression could relieve cell growth inhibition. Notch1-induced WISP-1 expression appeared to be Wnt11-dependent, but Wnt1-independent. Blockade of Wnt11 expression resulted in decreased WISP-1 expression and liberated Notch-induced cell growth inhibition. These findings indicated that inhibition of fibroblast proliferation by Notch pathway activation is mediated, at least in part, through regulating Wnt1-independent, but Wnt11-dependent WISP-1 expression.     

Notch1 control of oligodendrocyte differentiation in the spinal cord

We have selectively inhibited Notch1 signaling in oligodendrocyte precursors (OPCs) using the Cre/loxP system in transgenic mice to investigate the role of Notch1 in oligodendrocyte (OL) development and differentiation. Early development of OPCs appeared normal in the spinal cord. However, at embryonic day 17.5, premature OL differentiation was observed and ectopic immature OLs were present in the gray matter. At birth, OL apoptosis was strongly increased in Notch1 mutant animals. Premature OL differentiation was also observed in the cerebrum, indicating that Notch1 is required for the correct spatial and temporal regulation of OL differentiation in various regions of the central nervous system. These findings establish a widespread function of Notch1 in the late steps of mammalian OPC development in vivo.     

Comparison of the adhesion and proliferation characteristics of HUVEC and two endothelial cell lines (CRL 2922 and CRL 2873) on various substrata

Endothelial cell coverage of blood-contacting devices is crucial to their eventual success in the clinic. Two established human cell lines derived from HUVEC (human umbilical vascular endothelial cells), CRL 2922 and CRL 2873, have been widely utilized to study and model endothelial cell biology. However, it is not clear if these two cell lines would be useful for modeling primary endothelial cell interaction with newly-formulated biomaterials in tissue engineering applications. Hence, this study was conducted to compare the adhesion and proliferation characteristics of HUVEC grown on seven different substrata, tissue culture polystyrene (TCPS), gelatin, chitosan, poly-L-lysine, hyaluronan, poly-L-lactic acid (PLLA), and polylactic-co-glycolic acid (PLGA). The short-term adhesive behavior (2 h) of HUVEC on the various substrata was not closely-replicated by either CRL 2873 or CRL 2922. This was likely because the 2 h timeframe is too short for identification of differences in the interaction among the three cell types grown on various substrata. There was much faster proliferation of CRL 2922 on all seven substrata when compared to HUVEC and CRL 2873. Moreover, the proliferation rates of CRL 2922 on the various substrata showed little variation. In contrast, HUVEC and CRL 2873 displayed similar trends in proliferation rates, with gelatin and TCPS yielding the highest rates, and PLLA and PLGA yielding the lowest rates. Hence, CRL 2873 is better suited for modeling primary endothelial cell interaction with newly-formulated biomaterials than CRL 2922. The advantage of using CRL 2873 over HUVEC for biomaterial screening is that it is immortalized and displays much less inter-batch variability than primary culture.     

Quantitative contribution of NHE2 and NHE3 to rabbit ileal brush-border Na+/H+ exchange

Intestinal neutral NaCl absorption, which is made up ofbrush-border (BB)Na⁺/H⁺exchange linked to BBCl/HCO₃exchange, is up- and downregulated as part of digestion and diarrhealdiseases. Glucocorticoids stimulate ileal NaCl absorption and BBNa⁺/H⁺exchange. Intestinal BB contains twoNa⁺/H⁺exchanger isoforms, NHE2 and NHE3, but their relative roles in rabbitileal BBNa⁺/H⁺exchange has not been determined. A technique to separate the contribution of NHE2 and NHE3 to ileal BBNa⁺/H⁺exchange activity was standardized by using an amiloride-related compound, HOE-694. Under basal conditions, both NHE2 and NHE3 contribute ~50% to ilealNa⁺/H⁺exchange. Glucocorticoids (methylprednisolone) increase BBNa⁺/H⁺exchange (2.5 times) but increase only ileal NHE3 activity (4.1 times),without an effect on NHE2 activity. Thus ileal BBNa⁺/H⁺exchange in animals treated with glucocorticoids is 69% via NHE3. Aquantitative Western analysis for NHE3 was developed, using as aninternal standard a fusion protein of the COOH-terminal 85 amino acidsof NHE3 and maltose binding protein. Glucocorticoid treatment increasedthe amount of BB NHE3. The quantitative Western analysis showed thatNHE3 makes up 0.018% of ileal BB protein in control rabbits and0.042% (2.3 times as much) in methylprednisolone-treated rabbits.Methylprednisolone treatment did not alter the amount of ileal BB NHE2protein. NHE3 turnover number was estimated to be 458 cycles/s underbasal conditions and 708 cycles/s in glucocorticoid-treated ileum. Thusmethylprednisolone stimulates ileal BBNa⁺/H⁺exchange activity only by an effect on NHE3 and not on NHE2; it does soprimarily by increasing the amount of BB NHE3, although it alsoincreases the NHE3 turnover number.

     

Effect of Different Adjuvants on Protection and Side-Effects Induced by Helicobacter suis Whole-Cell Lysate Vaccination

Helicobacter suis (H. suis) is a widespread porcine gastric pathogen, which is also of zoonotic importance. The first goal of this study was to investigate the efficacy of several vaccine adjuvants (CpG-DNA, Curdlan, Freund’s Complete and Incomplete, Cholera toxin), administered either subcutaneously or intranasally along with H. suis whole-cell lysate, to protect against subsequent H. suis challenge in a BALB/c infection model. Subcutaneous immunization with Freund’s complete (FC)/lysate and intranasal immunization with Cholera toxin (CT)/lysate were shown to be the best options for vaccination against H. suis, as determined by the amount of colonizing H. suis bacteria in the stomach, although adverse effects such as post-immunization gastritis/pseudo-pyloric metaplasia and increased mortality were observed, respectively. Therefore, we decided to test alternative strategies, including sublingual vaccine administration, to reduce the unwanted side-effects. A CCR4 antagonist that transiently inhibits the migration of regulatory T cells was also included as a new adjuvant in this second study. Results confirmed that immunization with CT (intranasally or sublingually) is among the most effective vaccination protocols, but increased mortality was still observed. In the groups immunized subcutaneously with FC/lysate and CCR4 antagonist/lysate, a significant protection was observed. Compared to the FC/lysate immunized group, gastric pseudo-pyloric metaplasia was less severe or even absent in the CCR4 antagonist/lysate immunized group. In general, an inverse correlation was observed between IFN-γ, IL-4, IL-17, KC, MIP-2 and LIX mRNA expression and H. suis colonization density, whereas lower IL-10 expression levels were observed in partially protected animals.     

The role of nurses in physician-assisted deaths in Belgium

Background

Belgium’s law on euthanasia allows only physicians to perform the act. We investigated the involvement of nurses in the decision-making and in the preparation and administration of life-ending drugs with a patient’s explicit request (euthanasia) or without an explicit request. We also examined factors associated with these deaths.

Methods

In 2007, we surveyed 1678 nurses who, in an earlier survey, had reported caring for one or more patients who received a potential life-ending decision within the year before the survey. Eligible nurses were surveyed about their most recent case.

Results

The response rate was 76%. Overall, 128 nurses reported having cared for a patient who received euthanasia and 120 for a patient who received life-ending drugs without his or her explicit request. Respectively, 64% (75/117) and 69% (81/118) of these nurses were involved in the physician’s decision-making process. More often this entailed an exchange of information on the patient’s condition or the patient’s or relatives’ wishes (45% [34/117] and 51% [41/118]) than sharing in the decision-making (24% [18/117] and 31% [25/118]). The life-ending drugs were administered by the nurse in 12% of the cases of euthanasia, as compared with 45% of the cases of assisted death without an explicit request. In both types of assisted death, the nurses acted on the physician’s orders but mostly in the physician’s absence. Factors significantly associated with a nurse administering the life-ending drugs included being a male nurse working in a hospital (odds ratio [OR] 40.07, 95% confidence interval [CI] 7.37–217.79) and the patient being over 80 years old (OR 5.57, 95% CI 1.98–15.70).

Interpretation

By administering the life-ending drugs in some of the cases of euthanasia, and in almost half of the cases without an explicit request from the patient, the nurses in our study operated beyond the legal margins of their profession.Medical end-of-life decisions with a possible or certain life-shortening effect occur often in end-of-life care.^1–5 The most controversial and ethically debated medical practice is that in which drugs are administered with the intention of ending the patient’s life, whether at the patient’s explicit request (euthanasia) or not. The debate focuses mainly on the role and responsibilities of the physician.⁶ However, physicians worldwide have reported that nurses are also involved in these medical practices, mostly in the decision-making and sometimes in the administration of the life-ending drugs.^1–3,7–9 Critical care,¹⁰ oncology¹¹ and palliative care nurses^12,13 have confirmed this by reporting their own involvement, particularly in cases of euthanasia.^14,15In Belgium, the law permits physicians to perform euthanasia under strict requirements of due care, one of which is that they must discuss the request with the nurses involved.¹⁶ There are no further explicit stipulations determining the role of nurses in euthanasia. Physician-assisted death is legally regulated in some other countries as well (e.g., the Netherlands, Luxemburg and the US states of Oregon and Washington State), without specifying the role of nurses. Reports from nurses in these jurisdictions are scarce, apart from some that are limited to particular settings, or lack details about their involvement.^13,14We conducted this study to investigate the involvement of nurses in Flanders, Belgium, in the decision-making and in the preparation and administration of life-ending drugs with, or without, a patient’s explicit request. We also examined patient- and nurse-related factors associated with the involvement of nurses in these deaths. In a related research article, Chambaere and colleagues describe the findings from a survey of physicians in Flanders about the practices of euthanasia and assisted suicide, and the use of life-ending drugs without an explicit request from the patient.¹⁷