Finding balance: Tree-ring isotopes differentiate between acclimation and stress-induced imbalance in a long-term irrigation experiment.

Scots pine (Pinus sylvestris L.) is a common European tree species, and understanding its acclimation to the rapidly changing climate through physiological, biochemical or structural adjustments is vital for predicting future growth. We investigated a long-term irrigation experiment at a naturally dry forest in Switzerland, comparing Scots pine trees that have been continuously irrigated for 17 years (irrigated) with those for which irrigation was interrupted after 10 years (stop) and non-irrigated trees (control), using tree growth, xylogenesis, wood anatomy, and carbon, oxygen and hydrogen stable isotope measurements in the water, sugars and cellulose of plant tissues. The dendrochronological analyses highlighted three distinct acclimation phases to the treatments: irrigated trees experienced (i) a significant growth increase in the first 4 years of treatment, (ii) high growth rates but with a declining trend in the following 8 years and finally (iii) a regression to pre-irrigation growth rates, suggesting the development of a new growth limitation (i.e. acclimation). The introduction of the stop treatment resulted in further growth reductions to below-control levels during the third phase. Irrigated trees showed longer growth periods and lower tree-ring δ13 C values, reflecting lower stomatal restrictions than control trees. Their strong tree-ring δ18 O and δ2 H (O-H) relationship reflected the hydrological signature similarly to the control. On the contrary, the stop trees had lower growth rates, conservative wood anatomical traits, and a weak O-H relationship, indicating a physiological imbalance. Tree vitality (identified by crown transparency) significantly modulated growth, wood anatomical traits and tree-ring δ13 C, with low-vitality trees of all treatments performing similarly regardless of water availability. We thus provide quantitative indicators for assessing physiological imbalance and tree acclimation after environmental stresses. We also show that tree vitality is crucial in shaping such responses. These findings are fundamental for the early assessment of ecosystem imbalances and decline under climate change.

Hydrogen isotope fractionation in carbohydrates of leaves and xylem tissues follows distinct phylogenetic patterns: a common garden experiment with 73 tree and shrub species.

Recent methodological advancements in determining the nonexchangeable hydrogen isotopic composition (δ2 Hne ) of plant carbohydrates make it possible to disentangle the drivers of hydrogen isotope (2 H) fractionation processes in plants. Here, we investigated the influence of phylogeny on the δ2 Hne of twig xylem cellulose and xylem water, as well as leaf sugars and leaf water, across 73 Northern Hemisphere tree and shrub species growing in a common garden. 2 H fractionation in plant carbohydrates followed distinct phylogenetic patterns, with phylogeny reflected more in the δ2 Hne of leaf sugars than in that of twig xylem cellulose. Phylogeny had no detectable influence on the δ2 Hne of twig or leaf water, showing that biochemistry, not isotopic differences in plant water, caused the observed phylogenetic pattern in carbohydrates. Angiosperms were more 2 H-enriched than gymnosperms, but substantial δ2 Hne variations also occurred at the order, family, and species levels within both clades. Differences in the strength of the phylogenetic signals in δ2 Hne of leaf sugars and twig xylem cellulose suggest that the original phylogenetic signal of autotrophic processes was altered by subsequent species-specific metabolism. Our results will help improve 2 H fractionation models for plant carbohydrates and have important consequences for dendrochronological and ecophysiological studies.

Absence of canopy temperature variation despite stomatal adjustment in Pinus sylvestris under multidecadal soil moisture manipulation.

Global warming and droughts push forests closer to their thermal limits, altering tree carbon uptake and growth. To prevent critical overheating, trees can adjust their thermotolerance (Tcrit ), temperature and photosynthetic optima (Topt and Aopt ), and canopy temperature (Tcan ) to stay below damaging thresholds. However, we lack an understanding of how soil droughts affect photosynthetic thermal plasticity and Tcan regulation. In this study, we measured the effect of soil moisture on the seasonal and diurnal dynamics of net photosynthesis (A), stomatal conductance (gs ), and Tcan , as well as the thermal plasticity of photosynthesis (Tcrit , Topt , and Aopt ), over the course of 1 yr using a long-term irrigation experiment in a drought-prone Pinus sylvestris forest in Switzerland. Irrigation resulted in higher needle-level A, gs , Topt , and Aopt compared with naturally drought-exposed trees. No daily or seasonal differences in Tcan were observed between treatments. Trees operated below their thermal thresholds (Tcrit ), independently of soil moisture content. Despite strong Tcan and Tair coupling, we provide evidence that drought reduces trees' temperature optimum due to a substantial reduction of gs during warm and dry periods of the year. These findings provide important insights regarding the effects of soil drought on the thermal tolerance of P. sylvestris.

Synthesis of Sterically Encumbered Alkaline-Earth Metal Amides Applying the In Situ Grignard Reagent Formation.

Magnesium and calcium are too inert to deprotonate amines directly. For the synthesis of bulky amides alternative strategies are required and in the past, N-bound trialkylsilyl groups have been used to ease metalation reactions. The in situ Grignard reagent formation in stirred suspensions of magnesium or calcium with hydryl halide and imine in THF allows the synthesis of a plethora of amides with bulky silyl-free substituents. Ball milling protocols partially favor competitive side reactions such as aza-pinacol coupling reactions. Calcium is the advantageous choice for the in situ Grignard reagent formation and subsequent addition onto the imines yielding bulky calcium bis(amides) whereas the stronger reducing heavier alkaline-earth metals strontium and barium are less selective and hence, the aza-pinacol coupling reaction becomes competitive. Exemplary, the solid-state molecular structures of [(Et2 O)Mg(N(Ph)(CHPh2 )2 ] and [(Et2 O)2 Ca(N(Ph)(CHPh2 )2 ] have been determined.

In Situ Generation of Magnesium- and Calcium-Based Grignard Reagents for Amide Synthesis.

The alkaline-earth metals Mg and Ca are too inert for the direct metalation of primary and secondary amines. Consequently, activation prior to use is required. Alternatively, the Grignard reagents RMgX (R=alkyl, aryl, X=halide) can be applied in metalation of amines. However, such a straightforward procedure for the synthesis of alkylcalcium reagents is disadvantageous due to diverse side reactions, including Wurtz-type C-C coupling and ether degradation reactions. Therefore, suspensions of magnesium or calcium with amine can be treated in a smooth reaction with ethyl bromide in an ethereal solvent at room temperature. Intermediately formed RAeX (Ae=alkaline-earth metal, i. e., Mg, Ca) either metalates amines yielding the corresponding amides in an in situ Grignard metalation method (iGMM) or adds across C=N bonds of imines in an in situ Grignard addition method (iGAM). The amides R'2 N-AeX (Ae=Mg: Hauser bases) undergo Schlenk-type ligand exchange reactions yielding homoleptic Ae(NR'2 )2 and potentially sparingly soluble AeX2 .

A high-temperature water vapor equilibration method to determine non-exchangeable hydrogen isotope ratios of sugar, starch and cellulose.

The analysis of the non-exchangeable hydrogen isotope ratio (δ2 Hne ) in carbohydrates is mostly limited to the structural component cellulose, while simple high-throughput methods for δ2 Hne values of non-structural carbohydrates (NSC) such as sugar and starch do not yet exist. Here, we tested if the hot vapor equilibration method originally developed for cellulose is applicable for NSC, verified by comparison with the traditional nitration method. We set up a detailed analytical protocol and applied the method to plant extracts of leaves from species with different photosynthetic pathways (i.e., C3 , C4 and CAM). δ2 Hne of commercial sugars and starch from different classes and sources, ranging from -157.8 to +6.4‰, were reproducibly analysed with precision between 0.2‰ and 7.7‰. Mean δ2 Hne values of sugar are lowest in C3 (-92.0‰), intermediate in C4 (-32.5‰) and highest in CAM plants (6.0‰), with NSC being 2 H-depleted compared to cellulose and sugar being generally more 2 H-enriched than starch. Our results suggest that our method can be used in future studies to disentangle 2 H-fractionation processes, for improving mechanistic δ2 Hne models for leaf and tree-ring cellulose and for further development of δ2 Hne in plant carbohydrates as a potential proxy for climate, hydrology, plant metabolism and physiology.

Increasing temperature and vapour pressure deficit lead to hydraulic damages in the absence of soil drought.

Temperature (T) and vapour pressure deficit (VPD) are important drivers of plant hydraulic conductivity, growth, mortality, and ecosystem productivity, independently of soil water availability. Our goal was to disentangle the effects of T and VPD on plant hydraulic responses. Young trees of Fagus sylvatica L., Quercus pubescens Willd. and Quercus ilex L. were exposed to a cross-combination of a T and VPD manipulation under unlimited soil water availability. Stem hydraulic conductivity and leaf-level hydraulic traits (e.g., gas exchange and osmotic adjustment) were tracked over a full growing season. Significant loss of xylem conductive area (PLA) was found in F. sylvatica and Q. pubescens due to rising VPD and T, but not in Q. ilex. Increasing T aggravated the effects of high VPD in F. sylvatica only. PLA was driven by maximum hydraulic conductivity and minimum leaf conductance, suggesting that high transpiration and water loss after stomatal closure contributed to plant hydraulic stress. This study shows for the first time that rising VPD and T lead to losses of stem conductivity even when soil water is not limiting, highlighting their rising importance in plant mortality mechanisms in the future.

In Situ Grignard Metalation Method for the Synthesis of Hauser Bases.

The in situ Grignard Metalation Method (iGMM) is a straightforward one-pot procedure to quickly produce multigram amounts of Hauser bases R2 N-MgBr which are valuable and vastly used metalation reagents and novel electrolytes for magnesium batteries. During addition of bromoethane to a suspension of Mg metal and secondary amine at room temperature in an ethereal solvent, a smooth reaction yields R2 N-MgBr under evolution of ethane within a few hours. A Schlenk equilibrium is operative, interconverting the Hauser bases into their solvated homoleptic congeners Mg(NR2 )2 and MgBr2 depending on the solvent. Scope and preconditions are studied, and side reactions limiting the yield have been investigated. DOSY NMR experiments and X-ray crystal structures of characteristic examples clarify aggregation in solution and the solid state.

In situ Grignard Metalation Method, Part II: Scope of the One-Pot Synthesis of Organocalcium Compounds.

The in situ Grignard Metalation Method (iGMM) is a straightforward one-pot strategy to synthesize alkaline-earth metal amides in multi-gram scale with high yields via addition of bromoethane to an ethereal suspension of a primary or secondary amine and magnesium (Part I) or calcium (Part II). This method is highly advantageous because no activation of calcium is required prior to the reaction. Contrary to the magnesium-based iGMM, there are some limitations, the most conspicuous one is the large influence of steric factors. The preparation of Ca(hmds)2 succeeds smoothly within a few hours with excellent yields opening the opportunity to prepare large amounts of this reagent. Side reactions and transfer of the iGMM to substituted anilines and N-heterocycles as well as other H-acidic substrates such as cyclopentadienes are studied. Bulky amidines cannot be converted directly to calcium amidinates via the iGMM but stoichiometric calciation with Ca(hmds)2 enables their preparation.

One-Step Synthesis and Schlenk-Type Equilibrium of Cyclopentadienylmagnesium Bromides.

In the in situ Grignard metalation method (iGMM), the addition of bromoethane to a suspension of magnesium turnings and cyclopentadienes [C5 H6 (HCp), C5 H5 -Si(iPr)3 (HCpTIPS )] in diethyl ether smoothly yields heteroleptic [(Et2 O)Mg(CpR )(µ-Br)]2 (CpR =Cp (1), CpTIPS (2)). The Schlenk equilibrium of 2 in toluene leads to ligand exchange and formation of homoleptic [Mg(CpR )2 ] (3) and [(Et2 O)MgBr(µ-Br)]2 (4). Interfering solvation and aggregation as well as ligand redistribution equilibria hamper a quantitative elucidation of thermodynamic data for the Schlenk equilibrium of 2 in toluene. In ethereal solvents, mononuclear species [(Et2 O)2 Mg(CpTIPS )Br] (2'), [(Et2 O)n Mg(CpTIPS )2 ] (3'), and [(Et2 O)2 MgBr2 ] (4') coexist. Larger coordination numbers can be realized with cyclic ethers like tetrahydropyran allowing crystallization of [(thp)4 MgBr2 ] (5). The interpretation of the temperature-dependency of the Schlenk equilibrium constant in diethyl ether gives a reaction enthalpy ΔH and reaction entropy ΔS of -11.5 kJ mol-1 and 60 J mol-1 , respectively.

Scope and Limitations of the s-Block Metal-Mediated Pudovik Reaction.

The hydrophosphorylation of phenylacetylene with di(aryl)phosphane oxides Ar2 P(O)H (Pudovik reaction) yields E/Z-isomer mixtures of phenylethenyl-di(aryl)phosphane oxides (1). Alkali and alkaline-earth metal di(aryl)phosphinites have been studied as catalysts for this reaction with increasing activity for the heavier s-block metals. The Pudovik reaction can only be mediated for di(aryl)phosphane oxides whereas P-bound alkyl and alcoholate substituents impede the P-H addition across alkynes. The demanding mesityl group favors the single-hydrophosphorylated products 1-Ar whereas smaller aryl substituents lead to the double-hydrophosphorylated products 2-Ar. Polar solvents are beneficial for an effective addition. Increasing concentration of the reactants and the catalyst accelerates the Pudovik reaction. Whereas Mes2 P(O)H does not form the bis-phosphorylated product 2-Mes, activation of an ortho-methyl group and cyclization occurs yielding 2-benzyl-1-mesityl-5,7-dimethyl-2,3-dihydrophosphindole 1-oxide (3).

Alkaline-Earth Metal Bis[bis(trimethylsilyl)amide] Complexes with Weakly Coordinating 2,2,5,5-Tetramethyltetrahydrofuran Ligands.

The reaction of [(Me3Si)2N-Ae{µ-N(SiMe3)2}]2 with 2,2,5,5-tetramethyltetrahydrofuran in pentane yields the mononuclear complexes [(Me4thf)Ae{N(SiMe3)2}2] (Ae = Mg (1), Ca (2), Sr (3), and Ba (4)) with three-coordinate alkaline-earth metal centers. With increasing radius of the alkaline-earth metal atoms, the N-Ae-N bond angles decrease. These ether adducts significantly enhance the solubility of the bis(trimethylsilyl)amides of the alkaline-earth metals in hydrocarbon solvents. Contrary to the magnesium derivative 1, the heavier congeners dissociate into mononuclear [Ae{N(SiMe3)2}2] and free Me4THF without formation of sparingly soluble dinuclear [(Me3Si)2N-Ae{µ-N(SiMe3)2}]2.

Effectiveness of triclosan-coated PDS Plus versus uncoated PDS II sutures for prevention of surgical site infection after abdominal wall closure: the randomised controlled PROUD trial.

BACKGROUND: Postoperative surgical site infections are one of the most frequent complications after open abdominal surgery, and triclosan-coated sutures were developed to reduce their occurrence. The aim of the PROUD trial was to obtain reliable data for the effectiveness of triclosan-coated PDS Plus sutures for abdominal wall closure, compared with non-coated PDS II sutures, in the prevention of surgical site infections. METHODS: This multicentre, randomised controlled group-sequential superiority trial was done in 24 German hospitals. Adult patients (aged ≥18 years) who underwent elective midline abdominal laparotomy for any reason were eligible for inclusion. Exclusion criteria were impaired mental state, language problems, and participation in another intervention trial that interfered with the intervention or outcome of this trial. A central web-based randomisation tool was used to randomly assign eligible participants by permuted block randomisation with a 1:1 allocation ratio and block size 4 before mass closure to either triclosan-coated sutures (PDS Plus) or uncoated sutures (PDS II) for abdominal fascia closure. The primary endpoint was the occurrence of superficial or deep surgical site infection according to the Centers for Disease Control and Prevention criteria within 30 days after the operation. Patients, surgeons, and the outcome assessors were masked to group assignment. Interim and final analyses were by modified intention to treat. This trial is registered with the German Clinical Trials Register, number DRKS00000390. FINDINGS: Between April 7, 2010, and Oct 19, 2012, 1224 patients were randomly assigned to intervention groups (607 to PDS Plus, and 617 to PDS II), of whom 1185 (587 PDS Plus and 598 PDS II) were analysed by intention to treat. The study groups were well balanced in terms of patient and procedure characteristics. The occurrence of surgical site infections did not differ between the PDS Plus group (87 [14·8%] of 587) and the PDS II group (96 [16·1%] of 598; OR 0·91, 95% CI 0·66-1·25; p=0·64). Serious adverse events also did not differ between the groups-146 of 583 (25·0%) patients treated with PDS Plus had at least one serious adverse event, compared with 138 of 602 (22·9%) patients treated with PDS II; p=0·39). INTERPRETATION: Triclosan-coated PDS Plus did not reduce the occurrence of surgical site infection after elective midline laparotomy. Innovative, multifactorial strategies need to be developed and assessed in future trials to reduce surgical site infections. FUNDING: Johnson & Johnson Medical Limited.

Biochemical and biophysical drivers of the hydrogen isotopic composition of carbohydrates and acetogenic lipids.

The hydrogen isotopic composition (δ2H) of plant compounds is increasingly used as a hydroclimatic proxy; however, the interpretation of δ2H values is hampered by potential coeffecting biochemical and biophysical processes. Here, we studied δ2H values of water and carbohydrates in leaves and roots, and of leaf n-alkanes, in two distinct tobacco (Nicotiana sylvestris) experiments. Large differences in plant performance and biochemistry resulted from (a) soil fertilization with varying nitrogen (N) species ratios and (b) knockout-induced starch deficiency. We observed a strong 2H-enrichment in sugars and starch with a decreasing performance induced by increasing NO3-/NH4+ ratios and starch deficiency, as well as from leaves to roots. However, δ2H values of cellulose and n-alkanes were less affected. We show that relative concentrations of sugars and starch, interlinked with leaf gas exchange, shape δ2H values of carbohydrates. We thus provide insights into drivers of hydrogen isotopic composition of plant compounds and into the mechanistic modeling of plant cellulose δ2H values.

Enrichment of CD133-expressing cells in rectal cancers treated with preoperative radiochemotherapy is an independent marker for metastasis and survival.

BACKGROUND: The transmembrane glycoprotein CD133 (cluster of differentiation 133; also known as Prominin or PROM1) has been described as a potential stem cell marker in colorectal cancer and is associated with higher tumorigenic potential and resistance to radiochemotherapy (RCT). In this study, CD133 expression was evaluated in pre-RCT tumor biopsies and the corresponding post-RCT surgical specimens from patients with locally advanced rectal adenocarcinoma, and expression levels were correlated with histopathologic features and clinical follow-up. METHODS: One hundred twenty-six patients with International Union Against Cancer (UICC) stage II/III rectal cancer who received preoperative 5-fluorouracil (5-FU)-based RCT within the German Rectal Cancer Trials were investigated. Pre-RCT and post-RCT CD133 expression levels were determined using immunohistochemistry and were correlated with histopathologic parameters, tumor regression grade, cancer recurrence, and patient survival. RESULTS: Compared with pre-RCT biopsies, significantly higher CD133 expression was observed in tumor specimens (P = .01). However, no correlations were observed for either biopsies or tumor specimens between CD133 expression levels, histopathologic characteristics, or survival. In matched analyses of corresponding biopsy/tumor pairs, patients who had an increased fraction of CD133-expressing (CD133+) cells after preoperative RCT had significantly higher residual tumor stages (P = .02) and lower histopathologic tumor regression (P < .01). Moreover, these patients had significantly reduced disease-free survival and cancer-specific overall survival in univariate analysis (P < .001 and P = .004, respectively) and multivariate analysis (P = .003 and P = .024, respectively). CONCLUSIONS: The enrichment of CD133+ cancer cells during preoperative RCT was correlated with minor local tumor response, increased distant cancer recurrence, and decreased survival. The current results indicate that the up-regulation of intratumoral CD133 expression, in contrast to absolute pre-RCT and post-RCT CD133 levels, plays an important role in tumor progression and metastasis in patients with rectal cancer who are receiving neoadjuvant RCT.

Tree-ring isotopes from the Swiss Alps reveal non-climatic fingerprints of cyclic insect population outbreaks over the past 700 years.

Recent experiments have underlined the potential of δ2H in tree-ring cellulose as a physiological indicator of shifts in autotrophic versus heterotrophic processes (i.e., the use of fresh versus stored non-structural carbohydrates). However, the impact of these processes has not yet been quantified under natural conditions. Defoliator outbreaks disrupt tree functioning and carbon assimilation, stimulating remobilization, therefore providing a unique opportunity to improve our understanding of changes in δ2H. By exploring a 700-year tree-ring isotope chronology from Switzerland, we assessed the impact of 79 larch budmoth (LBM, Zeiraphera griseana [Hübner]) outbreaks on the growth of its host tree species, Larix decidua [Mill]. The LBM outbreaks significantly altered the tree-ring isotopic signature, creating a 2H-enrichment and an 18O- and 13C-depletion. Changes in tree physiological functioning in outbreak years are shown by the decoupling of δ2H and δ18O (O-H relationship), in contrast to the positive correlation in non-outbreak years. Across the centuries, the O-H relationship in outbreak years was not significantly affected by temperature, indicating that non-climatic physiological processes dominate over climate in determining δ2H. We conclude that the combination of these isotopic parameters can serve as a metric for assessing changes in physiological mechanisms over time.

Gastrointestinal stromal tumors.

INTRODUCTION: The gastrointestinal stromal tumor (GIST) is the most common mesenchymal tumor of the intestinal tract, known to be refractory to conventional chemotherapy or radiation. Its pathogenesis is defined by mutations within the KIT and PDGFRA gene, which constitutively activate KIT and PDGFRA oncoproteins, and serve as crucial diagnostic and therapeutic targets. DISCUSSION: Besides surgery, therapy with imatinib mesylate, which inhibits KIT kinase activity, represents the other cornerstone for the treatment of GIST. Still, the only curative option for GIST is given after complete surgical removal even in a metastatic setting, but recurrence is common, and the risk can be defined by surgical factors like incomplete resection, intraperitoneal rupture, or bleeding and tumor associated factors like tumor size, mitotic index, or localization. CONCLUSION: Consequently, adjuvant therapy with imatinib mesylate or other tyrosine kinase inhibitors is recommended for high-risk patients after complete resection. For unresectable and advanced GIST, a partial response or stable disease can be achieved in about 80% of patients with imatinib mesylate.

Sterically shielded primary anilides of the alkaline-earth metals of the type (thf)nAe(NH-Ar*)2 (Ae = Mg, Ca, Sr, and Ba; Ar* = bulky aryl).

Metalation of 2,4,6-triphenylphenylamine (H2N-C6H2-2,4,6-Ph3, 1a) and 4-methyl-2,6-bis(diphenylmethyl)aniline (2,6-bis(diphenylmethyl)-p-toluidine, H2N-C6H2-4-Me-2,6-(CHPh2)2, 2a) with dibutylmagnesium and Ae[N(SiMe3)2]2 with a stoichiometric 1 : 2 ratio in THF at room temperature yields the corresponding primary anilides [(thf)nAe{N(H)-C6H2-2,4,6-Ph3}2] (Ae/n = Mg/2 (1b), Ca/2 (1c), Sr/3 (1d), and Ba/3 (1e)) and [(thf)nAe{N(H)-C6H2-4-Me-2,6-(CHPh2)2}2] (Ae/n = Mg/2 (2b), Ca/3 (2c) and Sr/2 (2d)), respectively. The 1 : 1 reaction of Mg(n/sBu)2 and MgPh2 with 2a leads to the formation of heteroleptic [(thf)2Mg(R){N(H)-C6H2-4-Me-2,6-(CHPh2)2}] (R = n/sBu (2bBu), Ph (2bPh)). At 50 °C, the strontium complex 2d liberates one equivalent of 2avia intramolecular deprotonation of the triarylmethyl functionality yielding dinuclear [(thf)2Sr{N(H)-C6H2-4-Me-2-(CPh2)-6-(CHPh2)2}]2 (2d'). The barium compound is significantly more reactive and regardless of applied stoichiometry the isotypic barium congener [(thf)2Ba{N(H)-C6H2-4-Me-2-(CPh2)-6-(CHPh2)2}]2 (2e') forms. The molecular structures of 1c, 2d, 2d', and 2e' are stabilized by metal-phenyl π-interactions.