Expression of an Arabidopsis Sodium/Proton Antiporter Gene

Abstract Salinity is a major environmental stress that affects agricultural productivity worldwide. One approach to improving salt tolerance in crops is through high expression of the Arabidopsis gene AtNHX1, which encodes a vacuolar sodium/proton antiporter that sequesters excess sodium ion into the large intracellular vacuole. Sequestering cytosolic sodium into the vacuoles of plant cells leads to a low level of sodium in cytosol, which minimizes the sodium toxicity and injury to important enzymes in cytosol. In the meantime, the accumulation of sodium in vacuoles restores the correct osmolarity to the intracellular milieu, which favors water uptake by plant root cells and improves water retention in tissues under soils that are high in salt. To improve the yield and quality of peanut under high salt conditions, AtNHX1 was introduced into peanut plants through Agrobacterium-mediated transformation. The AtNHX1-expressing peanut plants displayed increased tolerance of salt at levels up to 150 mM NaCl. When compared to wild-type plants, AtNHX1-expressing peanut plants suffered less damage, produced more biomass, contained more chlorophyll, and maintained higher photosynthetic rates under salt conditions. These data indicate that AtNHX1 can be used to enhance salt tolerance in peanut

Sterile Neuroinflammation and Strategies for Therapeutic Intervention

Sterile neuroinflammation is essential for the proper brain development and tissue repair. However, uncontrolled neuroinflammation plays a major role in the pathogenesis of various disease processes. The endogenous intracellular molecules so called damage-associated molecular patterns or alarmins or damage signals that are released by activated or necrotic cells are thought to play a crucial role in initiating an immune response. Sterile inflammatory response that occurs in Alzheimer’s disease (AD), Parkinson’s disease (PD), stroke, hemorrhage, epilepsy, or traumatic brain injury (TBI) creates a vicious cycle of unrestrained inflammation, driving progressive neurodegeneration. Neuroinflammation is a key mechanism in the progression (e.g., AD and PD) or secondary injury development (e.g., stroke, hemorrhage, stress, and TBI) of multiple brain conditions. Hence, it provides an opportunity for the therapeutic intervention to prevent progressive tissue damage and loss of function. The key for developing anti-neuroinflammatory treatment is to minimize the detrimental and neurotoxic effects of inflammation while promoting the beneficial and neurotropic effects, thereby creating ideal conditions for regeneration and repair. This review outlines how inflammation is involved in the pathogenesis of major nonpathogenic neuroinflammatory conditions and discusses the complex response of glial cells to damage signals. In addition, emerging experimental anti-neuroinflammatory drug treatment strategies are discussed

Western blot band intensity is higher for serum samples from cancer patients.

<p>The average band intensity was greater in cancer group at 30, 37, 45, 65, and 100 kDa. This pattern was similar in cortex (A), hippocampus (B), and cerebellum (C). In addition, the pooled intensity from these regions was significantly increased in cancer relative to control at 65 (<i>P</i> = 0.044) and 45 kDa (<i>P</i> = 0.048) (D).</p

Detection of brain-directed autoantibodies in the serum of non-small cell lung cancer patients

<div><p>Antibodies against brain proteins were identified in the plasma of cancer patients and are defined to cause paraneoplastic neurological syndromes. The profiles of brain-directed antibodies in non-small cell lung cancer (NSCLC) are largely unknown. Here, for the first time, we compared autoantibodies against brain proteins in NSCLC (n = 18) against those present in age-matched non-cancer control subjects (n = 18) with a similar life-style, habit, and medical history. Self-recognizing immunoglobulin (IgG) are primarily directed against cells in the cortex (P = 0.008), hippocampus (P = 0.003–0.05), and cerebellum (P = 0.02). More specifically, IgG targets were prominent in the pyramidal, Purkinje, and granule cell layers. Furthermore, autoimmune IgG signals were localized to neurons (81%), astrocytes (48%), and endothelial (29%) cells. While cancer sera yielded overall higher intensity signals, autoantigens of 100, 65, 45, 37, and 30 kDa molecular weights were the most represented. Additionally, a group of 100 kDa proteins seem more prevalent in female adenocarcinoma patients (4/5, 80%). In conclusion, our results revealed autoantigen specificity in NSCLC, which implicitly depends on patient’s demographics and disease history. Patients at risk for lung cancer but with no active disease revealed that the immune profile in NSCLC is disease-dependent.</p></div

Schematic representation of the experimental design.

<p>Sera from lung cancer patients and non-cancer control individuals with similar lifestyles, habits, and age were filtered in 50 kDa molecular weight cut-off (MWCO) centrifugal filter devices to reduce small molecular weight proteins. These sera were then evaluated by immunohistochemistry and Western blotting performed on rat brain tissue and protein extract, respectively. Rats were sacrificed after intracardiac perfusion with PBS. Brains were then sectioned or dissected to harvest cortex, hippocampus, and cerebellum. Tissues were used for immunohistochemical detection to produce the results shown in Figs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181409#pone.0181409.g002" target="_blank">2</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181409#pone.0181409.g004" target="_blank">4</a>, whereas total protein isolated from the brain regions were analyzed by Western blotting as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181409#pone.0181409.g005" target="_blank">Fig 5</a>. The specific reactivity measured in Western blots was analyzed further in Figs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181409#pone.0181409.g006" target="_blank">6</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181409#pone.0181409.g007" target="_blank">7</a>.</p

Patients’ demographic properties.

<p>Patients’ demographic properties.</p

Autoantibodies are primarily directed against neuronal antigens.

<p>The figures show co-localization of cancer-derived autoreactive immunoglobulins (green) and neuronal (NeuN, red), astrocytic (GFAP, red), and endothelial (CD31, red) markers. Immunoreactivity for serum IgGs demonstrated staining of most cortical (A) and cerebellar (B) neurons. Small areas of ‘A’ were expanded in ‘E’ (cancer-IgG and anti-NeuN) and ‘F’. Some neurons were neither positive for cancer-IgG nor for anti-NeuN antibody (F, arrowheads). Purkinje cells were negative for NeuN (B, open arrows), nonetheless both Purkinje and granule cells immunoreacted with sera IgGs (B). A few IgG positive cells were defined as astrocytes (C) and endothelial cells (D). Small areas of ‘C’ and ‘D’ without DAPI staining were magnified to ‘G’ and ‘H’, respectively. The content of IgG in astrocytes was limited to the cytosol and to a few processes. The majority of cells in these panels were positive for cancer-IgGs (C, triangle arrows point; D, open triangle arrows point; green). A small percentage of cells remained unstained for IgGs (A, C, & D, simple arrows). I) Quantification of IgGs and cell-specific proteins in cortex. 81% of neurons were co-stained with cancer-derived IgG while 48% and 29% of astrocytes (by GFAP) and endothelial cells (CD31) were co-localized with cancer-IgG, respectively. NeuN, neuronal nuclei; GFAP, glial fibrillary acidic protein; CD31, cluster of differentiation 31.</p

A family of 100 kDa proteins is a primary target of serum IgGs from female lung adenocarcinoma patient.

<p>Gender- and disease-dependent (adeno vs squamous cell carcinomas) signals at 30, 37, 45, 65, and 100 kDa. Four of 5 female adenocarcinoma patients’ serum immunoglobulins detected an immunoblot band at 100 kDa in the cortical (A) and hippocampal (B) protein extracts. In cerebellar protein extracts, the maximum number of band detected was 4 (out of 7) at 100 kDa by the serum of male squamous cell carcinoma patient (C), which was the highest among all three regions. In the case of sera from male patients, proteins recognized by IgGs were variable in both adeno- and squamous carcinoma group. D) The pooled intensity from all three regions showed significant difference in adenocarcinoma male (<i>P</i> = 0.018, compared to squamous male). <i>P</i> values relative to squamous are provided in the graph. Adeno, adenocarcinoma; squam, squamous cell carcinoma.</p

Prevalence of individual bands on Western blot in cancer patients and non-cancer control individuals.

<p>Prevalence of individual bands on Western blot in cancer patients and non-cancer control individuals.</p

Sera from lung cancer patients are immunoreactive in rat brain.

<p>Sagittal brain sections were incubated with sera from cancer patients or non-cancer control subjects. A) No signal was detected in the absence of serum (negative control). B-C) Cancer-IgGs show robust immunoreactivity compared to control. Strong immunoreactivity was detected in the cortex, hippocampus, and cerebellum. D, F, H) Immunocytochemical staining with representative control (top row) and cancer (bottom row) sera. In cortex, staining intensity was significantly higher in cancer compared to control group (<i>P</i> = 0.008) (E). Autoantigen-directed immunoreactivity was observed in pyramidal neurons and their dendrites (arrows, D; also see Figs <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181409#pone.0181409.g003" target="_blank">3</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181409#pone.0181409.g004" target="_blank">4A</a>). F-G) Brain sections incubated in cancer sera had higher immunohistochemical intensity in the hippocampal areas, Baseline (<i>B</i>; <i>P</i> = 0.003), CA1 (<i>C</i>; <i>P</i> = 0.05), upper blade of the dentate gyrus (<i>U</i>; <i>P</i> = 0.006), hilus (<i>H</i>; <i>P</i> = 0.02), and lower blade of the dentate gyrus (<i>L; P</i> = 0.01). Areas selected for intensity quantification are shown in F (top). Autoreactive IgGs were also detected in blood vessels (arrows point, F; also see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0181409#pone.0181409.g004" target="_blank">Fig 4D</a>). H-I) Overall staining intensity in the cerebellum was greater with serum IgGs from cancer patients than control subjects (<i>P</i> = 0.02). The granule cell layer was more prominently stained with sera from cancer patients as pointed in a representative image (H, lower panel). The average difference in the immunoreactivity signals between cancer and control sera in the cortex, hippocampus, and cerebellum were 20.6, 17, and 15.1 arbitrary units, respectively. Con, control; Can, cancer; Data are shown as mean ± SEM for 7 patients.</p