Effects of prolonged drought on the anatomy of sun and shade needles in young Norway spruce trees

Predicted increases in the frequency and duration of drought are expected to negatively affect tree vitality, but we know little about how water shortage will influence needle anatomy and thereby the trees’ photosynthetic and hydraulic capacity. In this study, we evaluated anatomical changes in sun and shade needles of 20‐year‐old Norway spruce trees exposed to artificial drought stress. Canopy position was found to be important for needle structure, as sun needles had significantly higher values than shade needles for all anatomical traits (i.e., cross‐sectional needle area, number of tracheids in needle, needle hydraulic conductivity, and tracheid lumen area), except proportion of xylem area per cross‐sectional needle area. In sun needles, drought reduced all trait values by 10–40%, whereas in shade needles, only tracheid maximum diameter was reduced by drought. Due to the relatively weaker response of shade needles than sun needles in drought‐stressed trees, the difference between the two needle types was reduced by 25% in the drought‐stressed trees compared to the control trees. The observed changes in needle anatomy provide new understanding of how Norway spruce adapts to drought stress and may improve predictions of how forests will respond to global climate change.     

Moderation of calpain activity promotes neovascular integration and lumen formation during VEGF-induced pathological angiogenesis

Background

Successful neovascularization requires that sprouting endothelial cells (ECs) integrate to form new vascular networks. However, architecturally defective, poorly integrated vessels with blind ends are typical of pathological angiogenesis induced by vascular endothelial growth factor-A (VEGF), thereby limiting the utility of VEGF for therapeutic angiogenesis and aggravating ischemia-related pathologies. Here we investigated the possibility that over-exuberant calpain activity is responsible for aberrant VEGF neovessel architecture and integration. Calpains are a family of intracellular calcium-dependent, non-lysosomal cysteine proteases that regulate cellular functions through proteolysis of numerous substrates.

Methodology/Principal Findings

In a mouse skin model of VEGF-driven angiogenesis, retroviral transduction with dominant-negative (DN) calpain-I promoted neovessel integration and lumen formation, reduced blind ends, and improved vascular perfusion. Moderate doses of calpain inhibitor-I improved VEGF-driven angiogenesis similarly to DN calpain-I. Conversely, retroviral transduction with wild-type (WT) calpain-I abolished neovessel integration and lumen formation. In vitro, moderate suppression of calpain activity with DN calpain-I or calpain inhibitor-I increased the microtubule-stabilizing protein tau in endothelial cells (ECs), increased the average length of microtubules, increased actin cable length, and increased the interconnectivity of vascular cords. Conversely, WT calpain-I diminished tau, collapsed microtubules, disrupted actin cables, and inhibited integration of cord networks. Consistent with the critical importance of microtubules for vascular network integration, the microtubule-stabilizing agent taxol supported vascular cord integration whereas microtubule dissolution with nocodazole collapsed cord networks.

Conclusions/Significance

These findings implicate VEGF-induction of calpain activity and impairment of cytoskeletal dynamics in the failure of VEGF-induced neovessels to form and integrate properly. Accordingly, calpain represents an important target for rectifying key vascular defects associated with pathological angiogenesis and for improving therapeutic angiogenesis with VEGF.     

The physiological role of ascorbate as photosystem II electron donor: protection against photoinactivation in heat-stressed leaves

Previously, we showed that ascorbate (Asc), by donating electrons to photosystem II (PSII), supports a sustained electron transport activity in leaves in which the oxygen-evolving complexes were inactivated with a heat pulse (49°C, 40 s). Here, by using wild-type, Asc-overproducing, and -deficient Arabidopsis (Arabidopsis thaliana) mutants (miox4 and vtc2-3, respectively), we investigated the physiological role of Asc as PSII electron donor in heat-stressed leaves (40°C, 15 min), lacking active oxygen-evolving complexes. Chlorophyll-a fluorescence transients show that in leaves excited with trains of saturating single-turnover flashes spaced 200 ms apart, allowing continual electron donation from Asc to PSII, the reaction centers remained functional even after thousands of turnovers. Higher flash frequencies or continuous illumination (300 μmol photons m(-2) s(-1)) gradually inactivated them, a process that appeared to be initiated by a dramatic deceleration of the electron transfer from Tyr(Z) to P680(+), followed by the complete loss of charge separation activity. These processes occurred with half-times of 1.2 and 10 min, 2.8 and 23 min, and 4.1 and 51 min in vtc2-3, the wild type, and miox4, respectively, indicating that the rate of inactivation strongly depended on the Asc content of the leaves. The recovery of PSII activity, following the degradation of PSII proteins (D1, CP43, and PsbO), in moderate light (100 μmol photons m(-2) s(-1), comparable to growth light), was also retarded in the Asc-deficient mutant. These data show that high Asc content of leaves contributes significantly to the ability of plants to withstand heat-stress conditions.     

Retroviral MDR1 gene transfer into marrow-engrafting human peripheral blood progenitor cells results in preferential transgene expression in the immature myeloid compartment rather than in mature myeloid progeny in vivo

BACKGROUND: The objective of multidrug resistance-1 (MDR1) gene therapy is protection of the myeloid cell lineage. It is therefore important to examine the effect of retroviral transduction on myeloid maturation. Transfer of the human MDR1 gene can confer resistance to a variety of cytostatic drugs. For a safe application in humans it is paramount to follow-up the development of transduced cells. METHODS: We transduced human mobilized peripheral blood progenitor cells (PBPC) with a viral vector containing the human MDR1 cDNA and transplanted the transduced cells into non-obese diabetic severe combined immunodeficient (NOD/SCID) mice. The progeny of the transduced cells was analyzed in detail by flow cytometry. RESULTS: A detailed analysis by four-color flow cytometry showed that MDR1 transgene-expressing CD33+ myeloid cells were preferentially negative for the maturation-associated myeloid markers CD11b and CD10, while the untransduced CD33+ myeloid cells expressed significantly higher proportions of these Ag (P<0.01 each). There was no difference in the expression of B- or T-lymphoid Ag among the MDR1-transduced and untransduced lymphoid cells. DISCUSSION: These data indicate that retroviral MDR1 gene transfer results in preferential P-glycoprotein expression in myeloid progenitor cells, which is the target cell population for myelotoxicity of cytostatic drugs.     

Adipose Vascular Endothelial Growth Factor Regulates Metabolic Homeostasis through Angiogenesis

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Highlights? Two-way modulations of adipose VEGF were generated with aP2-Cre transgene ? Adipose VEGF KO reduces vasculature, increases hypoxia and inflammation in fat ? Adipose VEGF KO accelerates the development of metabolic disease in high-fat diet ? Induced adipose VEGF has opposite effect on fat and restores metabolic homeostasis     

Site-specific cassette exchange and germline transmission with mouse ES cells expressing phiC31 integrase

Currently two site-specific recombinases are available for engineering the mouse genome: Cre from P1 phage and Flp from yeast. Both enzymes catalyze recombination between two 34-base pair recognition sites, lox and FRT, respectively, resulting in excision, inversion, or translocation of DNA sequences depending upon the location and the orientation of the recognition sites. Furthermore, strategies have been designed to achieve site-specific insertion or cassette exchange. The problem with both recombinase systems is that when they insert a circular DNA into the genome (trans event), two cis-positioned recognition sites are created, which are immediate substrates for excision. To stabilize the trans event, functional mutant recognition sites had to be identified. None of the systems, however, allowed efficient selection-free identification of insertion or cassette exchange. Recently, an integrase from Streptomyces phage phiC31 has been shown to function in Schizosaccharomyces pombe and mammalian cells. This enzyme recombines between two heterotypic sites: attB and attP. The product sites of the recombination event (attL and attR) are not substrates for the integrase. Therefore, the phiC31 integrase is ideal to facilitate site-specific insertions into the mammalian genome.     

Causes of local recurrence after curative surgery for rectal cancer

The rate of local recurrence (LR) has been 20-40% after resective surgery for rectal cancer by the traditional - Miles or Dixon - operative technics. The authors performed curative resection in 358 patients with rectal cancer in a 10 year period (01.01.1990 - 31.12.2000) in the Surgical Department of Szeged University. Since 01.01.1996 the authors changed this type of surgery for the Heald technics (total mesorectal excision - TME - with sharp dissection, using the UltraCision device) for the surgical treatment of middle or lower third rectal cancer. To compare the results of the two procedures, the authors analysed their material in two periods: Period I: 01.01.1991 - 31.12.1992: 62 patients operated on with the traditional operative technics; LR 15% within 2 years after surgery. Period II: 01.01.1997 - 31.12.1998: 78 patients operated on with the Heald technics (TME with sharp dissection); LR 6.4% within 2 years after surgery. Based on their results, the authors found that the modern operative technics by Heald, used in the second period of the study, was a relevant factor decreasing LR from 15% to 6.4%, while the gender, age of the patients, ratio of the abdominoperineal extirpation versus anterior resection (APRE/AR) and the free margin of more than 3 cm proved to be irrelevant.     

Development of intron targeting (IT) markers for potato and cross-species amplification in Solanum nigrum (Solanaceae)

? Premise of the study: Intron Targeting (IT) primers were developed for potato using expressed sequence tags (EST) and NCBI database records to study genetic diversity. ? Methods and Results: Twenty-nine polymorphic intron targeting (IT) markers were generated and characterized from 30 samples of potato and 22 samples of Solanum nigrum to detect polymorphism. The number of alleles (A) per locus ranged from 2 to 7 in the analyzed populations, and the observed heterozygosity (H(O)) and expected heterozygosity (H(E)) from 0 to 0.833 and 0.750, respectively. All of the primers also amplified in the related species S. nigrum. ? Conclusions: The developed markers will provide valuable tools for genetic diversity analysis, genetic mapping, and marker-assisted breeding of potato and related Solanum species.