Ecoenzymatic stoichiometry and microbial processing of organic matter in northern bogs and fens reveals a common P-limitation between peatland types

We compared carbon (C), nitrogen (N), and phosphorus (P) concentrations in atmospheric deposition, runoff, and soils with microbial respiration [dehydrogenase (DHA)] and ecoenzyme activity (EEA) in an ombrotrophic bog and a minerotrophic fen to investigate the environmental drivers of biogeochemical cycling in peatlands at the Marcell Experimental Forest in northern Minnesota, USA. Ecoenzymatic stoichiometry was used to construct models for C use efficiency (CUE) and decomposition (M), and these were used to model respiration (R_m). Our goals were to determine the relative C, N, and P limitations on microbial processes and organic matter decomposition, and to identify environmental constraints on ecoenzymatic processes. Mean annual water, C, and P yields were greater in the fen, while N yields were similar in both the bog and fen. Soil chemistry differed between the bog and fen, and both watersheds exhibited significant differences among soil horizons. DHA and EEA differed by watersheds and soil horizons, CUE, M, and R_m differed only by soil horizons. C, N, or P limitations indicated by EEA stoichiometry were confirmed with orthogonal regressions of ecoenzyme pairs and enzyme vector analyses, and indicated greater N and P limitation in the bog than in the fen, with an overall tendency toward P-limitation in both the bog and fen. Ecoenzymatic stoichiometry, microbial respiration, and organic matter decomposition were responsive to resource availability and the environmental drivers of microbial metabolism, including those related to global climate changes.     

The Novel Use of a Heterozygous Knockout Mouse for Embryofetal Development Assessment of a Glucokinase Activator

Glucokinase activators (GKAs), such as AZD1656, are designed as antihyperglycemic agents for diabetics and can cause dose‐limiting hypoglycemia in normal animals used in embryofetal development studies. Genetically modified heterozygous GK knockout (gk^del/wt) mice are less susceptible to severe GKA‐induced hypoglycemia than wild‐type mice due to their elevated baseline glucose levels. In this study, the gk^del/wt mouse was used as an alternative rodent strain for embryofetal development studies with AZD1656. Heterozygous global knockout gk^del/wt females were dosed with 20, 50, or 130 mg/kg/day of AZD1656 or vehicle for a minimum of 14 consecutive days before mating with wild‐type males and throughout organogenesis. Maternal effects were confined to slightly reduced food consumption, reduced body weight gain, and the pharmacologic effect of decreased plasma glucose. Fetuses were genotyped. Fetal weights at the high dose were slightly reduced but there was no effect on fetal survival. There were two specific major malformations, omphalocele and right‐sided aortic arch, with increased fetal incidence in mid‐ and high‐dose fetuses (e.g., omphalocele fetal incidence of 0.6, 0.7, 4.6, and 2% across the dose groups) plus increased incidences of minor abnormalities and variants indicative of either delayed or disturbed development. Fetal weight and abnormalities were unaffected by fetal genotype. The fetal effects are considered hypoglycemia related. There was no effect on embryofetal survival in the gk^del/wt mouse at AZD1656 exposures, which were 70× higher than those causing 75% fetal death in rabbits. This illustrates the value of genetically modified animals in unraveling target versus chemistry‐related effects.     

Clinical and Serological Features of Patients Referred through a Rheumatology Triage System because of Positive Antinuclear Antibodies

Background

The referral of patients with positive anti-nuclear antibody (ANA) tests has been criticized as an inappropriate use of medical resources. The utility of a positive ANA test in a central triage (CT) system was studied by determining the autoantibody profiles and clinical diagnoses of patients referred to rheumatologists through a CT system because of a positive ANA test.

Methods

Patients that met three criteria were included: (1) referred to Rheumatology CT over a three year interval; (2) reason for referral was a “positive ANA”; (3) were evaluated by a certified rheumatologist. The CT clinical database was used to obtain demographic and clinical information and a serological database was used to retrieve specific ANA and/or extractable nuclear antigen (ENA) test results. Clinical information was extracted from the consulting rheumatologist''s report.

Results

15,357 patients were referred through the CT system; 643 (4.1%) of these because of a positive ANA and of these 263 (40.9%) were evaluated by a certified rheumatologist. In 63/263 (24%) of ANA positive patients, the specialist provided a diagnosis of an ANA associated rheumatic disease (AARD) while 69 (26.2%) had no evidence of any disease; 102 (38.8%) had other rheumatologic diagnoses and 29 (11%) had conditions that did not meet AARD classification criteria. Of ANA positive archived sera, 15.1% were anti-DFS70 positive and 91.2% of these did not have an AARD.

Conclusions

This is the first study to evaluate the serological and clinical features of patients referred through a CT system because of a positive ANA. The spectrum of autoantibody specificities was wide with anti-Ro52/TRIM21 being the most common autoantibody detected. Approximately 15% of referrals had only antibodies to DFS70, the vast majority of which did not have clinical evidence for an AARD. These findings provide insight into the utility of autoantibody testing in a CT system.     

Increased expression of the pro-apoptotic Bcl2 family member PUMA and apoptosis by the muscle regulatory transcription factor MyoD in response to a variety of stimuli

We have previously reported that the level of MyoD expression correlates with the level of apoptosis that occurs in a subpopulation of skeletal myoblasts induced to differentiate by serum withdrawal. Herein we document that MyoD expression contributes to the level of apoptosis in myoblasts and fibroblasts in response to a variety of apoptotic stimuli. Specifically, re-expression of MyoD in skeletal myoblasts rendered defective for both differentiation and apoptosis by the expression of oncogenic Ras restores their ability to undergo both differentiation and apoptosis in response to serum withdrawal. Further, using a fibroblast cell line expressing an estrogen receptor:MyoD fusion protein, we have determined that addition of estrogen sensitizes these fibroblasts to apoptosis induced by serum withdrawal, or by treatment with etoposide or thapsigargin. RNAi mediated silencing of MyoD in either 23A2 or C2C12 myoblasts renders these cells resistant to apoptosis induced by serum withdrawal, or by treatment with etoposide or thapsigargin. Finally, MyoD mediated regulation of the apoptotic response to these various stimuli, in both myoblasts and fibroblasts, correlates with the level of induction of the pro-apoptotic Bcl2 family member PUMA.     

Manufacturing development and clinical production of NKG2D chimeric antigen receptor–expressing T cells for autologous adoptive cell therapy

Background aims

Adoptive cell therapy employing natural killer group 2D (NKG2D) chimeric antigen receptor (CAR)-modified T cells has demonstrated preclinical efficacy in several model systems, including hematological and solid tumors. We present comprehensive data on manufacturing development and clinical production of autologous NKG2D CAR T cells for treatment of acute myeloid leukemia and multiple myeloma (ClinicalTrials.gov Identifier: NCT02203825). An NKG2D CAR was generated by fusing native full-length human NKG2D to the human CD3ζ cytoplasmic signaling domain. NKG2D naturally associates with native costimulatory molecule DAP10, effectively generating a second-generation CAR against multiple ligands upregulated during malignant transformation including MIC-A, MIC-B and the UL-16 binding proteins.

The genes of plant signal transduction

Due to the enormous importance of plants as a source of both food and raw materials, an understanding of the control of their growth and development is imperative. Basic to this understanding is the identification of the genes that enable the plant to adapt to its environment while at the same time adhering to its basic developmental plan. The aims of this review are first to briefly summarize the various approaches that have been used to identify these plant signaling genes and second to give an overview of the genes that have been cloned so far and what these genes may tell us about the nature of signal transduction in plants.

The advent of modern molecular biology and molecular genetics has revolutionized our ability to unravel the complexities of plant signal transduction pathways. A whole battery of techniques are now available to identify the genes that control the plants development and ability to adapt to its environment.⁶⁵ Each technique has its own strengths and weaknesses and must be carefully selected by the researcher according to the question that he or she would like to ask.     

Relationship between the shape and the membrane potential of human red blood cells

Summary Microscopic observations of isotonic suspensions of human red blood cells demonstrate that cell shape is unaltered when the transmembrane electrical potential, orE _m , is set in the range –85 to +10 mV with valinomycin at varied external K⁺, or K _o .E _m was measured with the fluorescent potentiometric indicator, diS-C₃(5), as calibrated by a pH method. Repeating Glaser's experiments in which echinocytosis was attributed to hyperpolarization, we found that at low ionic strength the pH-dependent effects of amphotericin B appear to be unrelated toE _m . The effects of increased intracellular Ca²⁺, or Ca _o , on echinocytosis and onE _m are separable. With Ca ionophore A23187 half-maximal echinocytosis occurs at greater Ca _o than that which induces the half-maximal hyperpolarization associated with Ca-induced K⁺ conductance (Gardos effect). Thus, cells hyperpolarized by increased Ca _o remain discoidal when Ca is below the threshold for echinocytosis. With A23187 and higher Ca _o , extensive echinocytosis occurs in cells which are either hyperpolarized or at their resting potential. The Ca-activation curve for echinocytosis is left-shifted by low K _o , a new observation consistent with increased DIDS-sensitive uptake of⁴⁵Ca by hyperpolarized cells. These results support the following conclusions: (1) the shape and membrane potential of human red blood cells are independent under the conditions studied; (2) in cells treated with A23187, the Gardos effect facilitates echinocytosis by increasing Ca.