Important photosynthetic contribution of silique wall to seed yield-related traits in Arabidopsis thaliana.

In plants, green non-foliar organs are able to perform photosynthesis just as leaves do, and the seed-enclosing pod acts as an essential photosynthetic organ in legume and Brassica species. To date, the contribution of pod photosynthesis to seed yield and related components still remains largely unexplored, and in Arabidopsis thaliana, the photosynthetic activity of the silique (pod) is unknown. In this study, an Arabidopsis glk1/glk2 mutant defective in both leaf and silique photosynthesis was used to create tissue-specific functional complementation lines. These lines were used to analyze the contribution of silique wall photosynthesis to seed yield and related traits, and to permit the comparison of this contribution with that of leaf photosynthesis. Our results showed that, together with leaves, the photosynthetic assimilation of the silique wall greatly contributed to total seed yield per plant. As for individual components of yield traits, leaf photosynthesis alone contributed to the seed number per silique and silique length, while silique wall photosynthesis alone contributed to thousand-seed weight. In addition, enhancing the photosynthetic capacity of the silique wall by overexpressing the photosynthesis-related RCA gene in this tissue resulted in significantly increased seed weight and oil content in the wild-type (WT) background. These results reveal that silique wall photosynthesis plays an important role in seed-related traits, and that enhancing silique photosynthesis in WT plants can further improve seed yield-related traits and oil production. This finding may have significant implications for improving the seed yield and oil production of oilseed crops and other species with pod-like organs.

New efficient imidazolium aldoxime reactivators for nerve agent-inhibited acetylcholinesterase.

Herein, we described a new class of uncharged non-pyridinium reactivators for nerve agent-inhibited acetylcholinesterase (AChE). Based on a dual site binding strategy, we conjugated the imidazolium aldoxime to different peripheral site ligands (PSLs) of AChE through alkyl chains. Compared with the known quaternary pyridinium reactivators, two of the resulting conjugates (7g and 7h) were highlighted to be the first efficient non-pyridinium oxime conjugates exhibiting similar or superior ability to reactivate sarin-, VX- and tabun-inhibited AChE. Moreover, they were more broad-spectrum reactivators.

H5PV2Mo10O40 encapsulated in MIL-101(Cr): facile synthesis and characterization of rationally designed composite materials for efficient decontamination of sulfur mustard.

Currently extensive effort is compulsively expended to decontaminate efficiently banned chemical war agents. In this work, H5PV2Mo10O40 molecules have been encapsulated in mesoporous MIL-101(Cr), which features two types of mesoporous cages (internal diameters of 29 Å and 34 Å) and microporous windows (diameters of 12 Å and 16 Å), leading to the formation of a new composite H5PV2Mo10O40@MIL-101(Cr) through a simple impregnation method. The composite was characterized thoroughly by elemental analysis, FT-IR spectroscopy, powder X-ray diffraction, scanning electron microscopy, energy dispersive X-ray spectroscopy, TG/DTA, and textural analysis thereby confirming the encapsulation of the H5PV2Mo10O40 into MIL-101(Cr). The decontamination efficiency of sulfur mustard (4 µL HD in 40 µL of petroleum ether) by 20 mg of the composite is found to be 97.39% in 120 min under ambient conditions. GC-MS analysis on the decontaminated products using 2-chloroethyl ethyl sulfide (CEES), which has been widely used as a simulant of sulfur mustard, showed that MIL-101(Cr) just decontaminates CEES by adsorption, while CEES can be decontaminated under ambient conditions by a synergetic combination of adsorption of MIL-101(Cr) and subsequent chemical oxidation degradation to nontoxic 2-chloroethyl ethyl sulfoxide (CEESO) due to the presence of highly dispersed H5PV2Mo10O40 within the composites.

Fertility and early embryonic development toxicity of penequine hydrochloride in mice.

Penequine hydrochloride, a novel anticholinergic agent, was developed as an effective treatment for organophosphorus intoxication. The potential for penequine hydrochloride to induce fertility and early embryonic developmental toxicity was evaluated in AMMS-1 mice. Totally 320 healthy, sexual mature and nulliparous AMMS-1 mice were orally treated with the chemical in drinking water at dose levels of 0, 2.5, 12.5 and 62.5 mg/L from 60 days before cohabitation to successful copulation in 160 males and from two weeks before cohabitation to GD 6 in 160 females, respectively. All the parental mice were observed for body weights, water consumption and any abnormal change during treatment period. Caesarean sections were carried out on day 14 of pregnancy in half assumed-pregnant females, and all the intrauterine data were recorded. Pups naturally delivered by the other half females were weighed, and examined for viability, sex ratio and gross malformations. About 7 days after cohabitation period, all the paternal males were examined for epididymal and testicular weights, sperm number and sperm motility. The decreases in fertility/fecundity indices and maternal weight gain were found at high-dose level in both caesarean sections and natural delivery observations. The primary developmental toxicity of the chemical included decreases in relative organ (epididymis, liver and lung) weights at mid- and high-dose levels in pups on postnatal day (PND) 35. The cause of both the decreased fertility/fecundity indices in F0 males and the decreased relative organ weights in F1 pups are not well known but are presently under investigation. Under the experimental conditions, penequine hydrochloride did not produce any adverse effects (expect the decreases in certain relative organ weights) up to and including 12.5 mg/L (2.53 mg/kg/day in males and 2.19 mg/kg/day in females) corresponding to approximately 72 times above anticipated dosage in human.

Suppression of perfluoroisobutylene induced acute lung injury by pretreatment with pyrrolidine dithiocarbamate.

Perfluoroisobutylene (PFIB) is produced as a main by-product in large quantities by the fluoropolymer industry. As a highly toxic compound, even the case of brief inhalation of PFIB can result in acute lung injury (ALI), pulmonary edema and even death. To test for any preventive or therapeutic effects of pyrrolidine dithiocarbamate (PDTC), a NF-kappaB activation inhibitor, against PFIB inhalation-induced ALI, mice were exposed in a flow-past exposure system to PFIB and the prophylactic and therapeutic effects of PDTC were studied. The inhibitory effects of PDTC on ALI, the activation of NF-kappaB, as well as the expression of cytokines (IL-1beta and IL-8) after PFIB exposure were evaluated. The results demonstrated that pretreatment with PDTC (120 mg/kg, 30 min before PFIB exposure) could significantly lower the lung coefficient (wet lung-to-body weight ratio, dry lung-to-body weight ratio, water content in the lung, and lung wet-to-dry weight ratio) and protein content in bronchoalveolar lavage fluid (BALF), but no effects of PDTC were found when PDTC was treated after PFIB inhalation, suggesting a preventative effect rather than a therapeutic effect of PDTC. Furthermore, the above preventative effects of PDTC (when given at 30 min before PFIB exposure) on PFIB-induced lung injury were achieved in a dose-dependent manner. In support of these preventive effects of PDTC, our toxicological studies demonstrated that PFIB-inhalation induced a quick activation of NF-kappaB (0.5 h post PFIB exposure) and expression of IL-1beta and IL-8 (0.5 h and 1 h post PFIB exposure, respectively). Pretreatment with PDTC (120 mg/kg, 30 min before PFIB exposure) resulted in a significant inhibitive effect on the activation of NF-kappaB (0.5 h post PFIB exposure) and expression of IL-1beta and IL-8 (1 h post PFIB exposure). The mortality, the extent of lung injury of the mice indexed by lung coefficients, the content of total protein and albumin in BALF, as well as the lung histopathologic changes, were dramatically alleviated in PFIB exposure after pretreatment with PDTC, clearly suggesting that PDTC has a prophylactic role against PFIB inhalation-induced ALI, and that NF-kappaB activation might play a central role in initiating an acute inflammatory response and in causing injury to the lungs after PFIB inhalation.

Arginine-rich membrane-permeable peptides are seriously toxic.

The membrane-permeable peptides (MPP) such as undecapeptides TAT (YGRKKRRQRRR) and CTP (YGRRARRRRRR) have been receiving much attention for delivering various kinds of low membrane-permeability materials in vitro and in vivo. We have successfully used MPP in carrying various proteins through blood-brain barrier (BBB) in treatment of many kinds of nervous diseases. However, people always concentrate their mind on the efficacy and the mechanism of permeation of the conjugates across BBB, but overlook the toxicity of the membrane-permeable peptide itself. Once we injected intravenously not very large amounts of gamma-aminobutyric acid-MPP (GABA-MPP) to the mice, to our great surprise, the mice died within seconds with seizure, whereas the GABA control mice well survived. Thus, the importance of the toxicity of MPPs and their conjugates comes into the field of our vision. The low LD50 values of arginine-rich TAT (27.244 mg kg-1 ) and CTP (21.345 mg kg-1 ) per se in mice indicate that they all fall within the range of highly toxic chemicals. Among the arginine-rich peptides, R11 (RRRRRRRRRRR), a peptide composed purely of arginine residues, has the lowest LD50 value (16.5 mg kg-1 ) and manifests the highest toxicity, whereas TD (ACSSSPSKHCG), a peptide without arginine residue, shows a much lower toxicity and higher survival rate in mice. The mass percentage of arginine-rich MPP in the conjugate is critically important, the mass radio of arginine in the MPP appears a linear correlation with the toxicity. Thus we conclude, the arginine-rich MPPs are more suitable for using in the macro-molecular conjugates, but not in the small-molecular one.

BBB: Permeable Conjugate of Exogenic GABA.

Neurotransmitters are the key factors in ameliorating the symptoms of nervous system diseases. Stroke/cerebral ischemia has been proven to be caused by the excess release of excitatory amino acid glutamate in the brain, and the inhibitory neurotransmitter γ-aminobutyric acid (GABA) is considered to be the best choice to counteract the action of glutamate. Here, we show that GABA conjugated to a cytoplasmic transduction peptide (YGRRARRRRRR) by means of custom chemical synthesis could penetrate through the blood-brain barrier, increasing the GABA level in the plasma of rats and mice, which, as a result, display a state of calmness and somnolence.

Layer-by-layer assembly of Cu3(BTC)2 on chitosan non-woven fabrics: a promising haemostatic decontaminant composite material against sulfur mustard.

Cu3(BTC)2 (H3BTC = 1,3,5-benzenetricarboxylic acid) was anchored onto the surface of carboxymethylated chitosan non-woven fabrics by a controllable layer-by-layer technique in alternating solution baths of Cu(OAc)2·H2O and H3BTC solutions, and the resulting [Cu3(BTC)2]n@chitosan non-woven fabric composite materials (n = number of alternate deposition cycles) were thoroughly characterized. The results showed that the composite materials not only exhibited excellent decontamination ability against sulfur mustard (HD), with an enhanced degradation rate of HD with increasing n, but also possessed remarkable haemostasis performance. The degradation efficiency of sulfur mustard by [Cu3(BTC)2]4@chitosan was found to be much higher than that of pristine [Cu3(BTC)2]4. The inherent haemostatic capabilities of the chitosan non-woven fabrics were not affected by the growth of Cu3(BTC)2 on the surface of chitosan. Oral and histological toxicity examinations of the pristine Cu3(BTC)2 sample showed that damage and toxicity in the viscera of mice was very low, even if the intra-gastric administration dose of Cu3(BTC)2 reached a very high level (50 mg of Cu3(BTC)2 per kg of mouse). The results have shown that the as-prepared composite can be used safely as a promising haemostatic decontaminant with good recyclability against sulfur mustard.

Systematic Analysis of Hsf Family Genes in the Brassica napus Genome Reveals Novel Responses to Heat, Drought and High CO2 Stresses.

Drought and heat stress are major causes of lost plant crop yield. In the future, high levels of CO2, in combination of other abiotic stress factors, will become a novel source of stress. Little is known of the mechanisms involved in the acclimation responses of plants to this combination of abiotic stress factors, though it has been demonstrated that heat shock transcription factors (Hsfs) are involved in plant response to various abiotic stresses. In this study, we performed a genome-wide identification and a systematic analysis of genes in the Hsf gene family in Brassica napus. A total of 64 genes encoding Hsf proteins were identified and classified into 3 major classes: A, B and C. We found that, unlike in other eudicots, the A9 subclass is absent in rapeseed. Further gene structure analysis revealed a loss of the only intron in the DBD domain for BnaHsf63 and -64 within class C, which is evolutionarily conserved in all Hsf genes. Transcription profile results demonstrated that most BnaHsf family genes are upregulated by both drought and heat conditions, while some are responded to a high CO2 treatment. According to the combined RNA-seq and qRT-PCR analysis, the A1E/A4A/A7 subclasses were upregulated by both drought and heat treatments. Members in class C seemed to be predominantly induced only by drought. Among BnaHsf genes, the A2/A3/B2 subclasses were regulated by all three abiotic stresses. Members in A2/B2 subclasses were upregulated by drought and heat treatments, but were downregulated under high CO2 conditions. While the A3 subclass was upregulated by all the three abiotic stresses. Various stress-related cis-acting elements, enriched in promoter regions, were correlated with the transcriptional response of BnaHsfs to these abiotic stresses. Further study of these novel groups of multifunctional BnaHsf genes will improve our understanding of plant acclimation response to abiotic stresses, and may be useful for improving the abiotic stress resistance of crop varieties.

The pre- and post-natal toxicity of penequine hydrochloride in mice.

Penequine hydrochloride, a novel anticholinergic agent, was developed as an effective treatment for organophosphorus intoxication (e.g., soman poisoning). The current study was performed to assess the potential pre- and post-natal toxicity of penequine hydrochloride in mice. Approximately 120 timed-pregnant mice were assigned to four dose groups (n=30 per group). Dams were exposed orally to 0, 2.5, 12.5, 62.5 mg/L penequine hydrochloride in drinking water from gestation day 6 to lactation day 21. The F1 generation mice, which were not exposed directly to penequine hydrochloride as pups or as adults, were bred to produce F2 generation fetuses for the fertility test of the F1 population. Various pre- and post-natal measurements, including neurobehavioral tests, were performed with the F0 and F1 mice. Among the significant findings were decreases in water consumption, viability, organ weights and delay of physical landmarks in 62.5 mg/L groups. With the exception of treatment-unrelated abnormality in surface righting reflex in the F1 generation, penequine hydrochloride did not produce any adverse effects at doses up to and including 12.5 mg/L (equal to 2.5 mg/kg/day in mice) that were at least 75 times of human therapeutic dosage.

Novel nonquaternary reactivators showing reactivation efficiency for soman-inhibited human acetylcholinesterase.

Soman is a highly toxic nerve agent with strong inhibition of acetylcholinesterase (AChE), but of the few reactivators showing antidotal efficiency for soman-inhibited AChE presently are all permanently charged cationic oximes with poor penetration of the blood-brain barrier. To overcome this problem, uncharged reactivators have been designed and synthesized, but few of them were efficient for treating soman poisoning. Herein, we used a dual site biding strategy to develop more efficient uncharged reactivators. The ortho-hydroxylbenzaldoximes were chosen as reactivation ligands of AChE to prevent the secondary poisoning of AChE, and simple aromatic groups were used as peripheral site ligands of AChE, which were linked to the oximes in a similar way as that found in the reactivator HI-6. The in vitro experiment demonstrated that some of the resulting conjugates have robust activity against soman-inhibited AChE, and oxime 8b was highlighted as the most efficient one. Although not good as HI-6 in vitro, these new compounds hold promise for development of more efficient centrally acting reactivators for soman poisoning due to their novel nonquaternary structures, which are predicted to be able to cross the blood-brain barrier.

The prophylactic and therapeutic effects of cholinolytics on perfluoroisobutylene inhalation induced acute lung injury.

Perfluoroisobutylene (PFIB) is a kind of fluoro-olefin that is ten times more toxic than phosgene. The mechanisms of the acute lung injury (ALI) induced by PFIB inhalation remain unclear. To find possible pharmacological interventions, mice and rats were exposed to PFIB, and the prophylactic or therapeutic effects of 3-quinuclidinyl benzilate (QNB) and anisodamine were studied and confirmed. It was observed that the wet lung/body weight and the dry lung/body weight ratios at 24 h after PFIB exposure (130 mg/m(3) for 5 min) were significantly decreased when a single dose of QNB (5 mg/kg) was administered intraperitoneally either 30 min before exposure or 10 h after exposure. Anisodamine was without any prophylactic or therapeutic effects at single doses below 30 mg/kg. The effects of QNB against PFIB inhalation induced ALI were well evidenced by the significantly decreased mice mortality at 72 h, the total protein concentration in bronchoalveolar lavage fluid at 24 h after the PFIB exposure, as well as the ultrastructural observations. The analysis of the time courses of lung sulfhydryl concentration, myeloperoxidase (MPO) activity and hemorheology assay showed that the toxicity of PFIB may be due to consumption of lung protein sulfhydryl, influx of polymorphonuclear leukocytes (PMNs) into the lung, and increased peripheral blood viscosity at a low shear rate, all of which were partially blocked by QNB intervention except for PMN influx. The results suggest that cholinolytics might have prophylactic and therapeutic roles in PFIB inhalation induced ALI.

Monoclonal antibody, mAb 4C13, an effective detoxicant antibody against ricin poisoning.

Ricin is a glycoprotein produced in castor seeds and consists of two polypeptide chains named Ricin Toxin A Chain (RTA) and Ricin Toxin B Chain (RTB), linked via a disulfide bridge. Due to its high toxicity, ricin is regarded as a high terrorist risk for the public. However, antibodies can play a pivotal role in neutralizing the toxin. In this research, the anti-toxicant effect of mAb 4C13, a monoclonal antibody (mAb) established using detoxicated ricin as the immunized antigen, was evaluated. Compared with mAb 4F2 and mAb 5G6, the effective mechanism of mAb 4C13 was analyzed by experiments relating to its cytotoxicity, epitope on ricin, binding kinetics with the toxin, its blockage on the protein synthesis inhibition induced by ricin and the intracelluar tracing of its complex with ricin. Our result indicated that mAb 4C13 could recognize and bind to RTA, RTB and exert its high affinity to the holotoxin. Both cytotoxicity and animal toxicity of ricin were well blocked by pre-incubating the toxin with mAb 4C13. By intravenous injection, mAb 4C13 could rescue the mouse intraperitoneally (ip) injected with a lethal dose of ricin (20µg/kg) even at 6h after the intoxication and its efficacy was dependent on its dosage. This research indicated that mAb 4C13 could be an excellent candidate for therapeutic antibodies. Its potent antitoxic efficiency was related to its recognition on the specific epitope with very high affinity and its blockage of protein synthesis inhibition in cytoplasm followed by cellular internalization with ricin.

[Modeling of acute respiratory distress syndrome in canine after inhalation of perfluoroisobutylene and preliminary study on mechanisms of injury].

OBJECTIVE: To establish of acute respiratory distress syndrome (ARDS) model in canine after inhalation of perfluoroisobutylene (PFIB), and to observe the progressing of lung injury, and to study the mechanisms of injury. METHODS: A device of inhalation of PFIB for canine was made. The concentration of PFIB was 0.30 - 0.32 mg/L. Serum IL-6 and IL-8 were dynamically measured. Clinical manifestations, pathology of organs in canine were observed. RESULTS: (1) During inhalation, the concentration of PFIB remained stable; (2) After inhalation, blood arterial oxygen partial pressure fell gradually, and eventually met the criteria for diagnosing ARDS; (3) The level of IL-8 in serum rises significantly after inhalation (P < 0.05), whereas that of IL-6 was not obviously altered (P > 0.05); (4) Within 6 hours after inhalation, no abnormality in canine was observed, but afterwards symptoms gradually appeared, and typical breath of ARDS, such as high frequency and lower level could be seen in later phase; (5) Pathological examination showed severe congestion, edema and atelectasis in most part of both lungs, and signs of anoxia in other organs. CONCLUSIONS: (1) The device designed is capable of ensuring control of inhalation of PFIB; (2) Exposure to PFIB for 30 mins, canines all met the criteria for diagnosing ARDS 22 hours after inhalation, therefore the modeling is successful; (3) PFIB specifically damages the lung by causing excessive inflammation.

Injury of cell tight junctions and changes of actin level in acute lung injury caused by the perfluoroisobutylene exposure and the role of Myosin light chain kinase.

OBJECTIVES: To investigate the injury of cell tight junctions and change in actin level in the alveolus epithelial cells of the lung after perfluoroisobutylene (PFIB) exposure and the role of myosin light chain kinase (MLCK) in the injury. METHODS: Rats and mice were exposed to a sublethal dose of PFIB. The changes in tight junction zonula occludens-1 (ZO-1), actin and myosin light chain kinase (MLCK) were detected by immunofluorescence at 30 min, 1, 2, 4, 8, 16, 24, 48 and 72 h after PFIB exposure. The role of MLCK was analyzed by lung indices and the actin level. RESULTS: The normal ZO-1 immunofluorescence density and those after PFIB exposure were 71.63, 39.41, 37.59, 35.71, 33.22, 31.34, 31.61, 24.51, 40.03 and 44.71 respectively, The normal actin immunofluorescence density and those after PFIB exposure were 31.82, 36.46, 36.57, 41.60, 40.95, 35.41, 30.69, 19.96, 29.30 and 33.00 respectively, The normal MLCK immunofluorescence density and those after PFIB exposure were 61.21, 50.87, 48.37, 43.65, 41.96, 35.44, 31.77, 30.85, 33.10 and 38.20 respectively. When the MLCK inhibitor ML-7 was given in advance, pulmonary edema and actin degradation were suppressed. CONCLUSIONS: At an earlier stage, the increased permeability of the blood-air barrier after PFIB exposure is probably the result of injury of cell tight junctions that acts in concert with later changes in actin, resulting in an increase in permeability. MLCK could be a potential target for novel drug development for relief of acute lung injury.

Cell injuries of the blood-air barrier in acute lung injury caused by perfluoroisobutylene exposure.

OBJECTIVES: To investigate the complete process of cell injuries in the blood-air barrier after perfluoroisobutylene (PFIB) exposure. METHODS: Rats were exposed to PFIB (140 mg/m(3)) for 5 min. The pathological changes were evaluated by lung wet-to-dry weight ratio, total protein concentration of bronchoalveolar lavage fluid and HE stain. Ultrastructural changes were observed by transmission electron microscope. Apoptosis was detected by in situ apoptosis detection. Changes of actin in the lung tissue were evaluated by western blot assay. RESULTS: No significant pulmonary edema or increased permeability was observed within the first 4 h, post PFIB exposure. However, inflammatory cell infiltration and alveolar wall thickening were observed from 2 h. Destruction of the alveoli constitution integrity, edema and protein leakage were observed at 8 h. The injuries culminated at 24 h and then recovered gradually. The ultrastructural injuries of alveolar type I epithelial cells, alveolar type II epithelial cells and pulmonary microvascular endothelial cells were observed at 30 min post PFIB exposure. Some injuries were similar to apoptosis. Compared with control, more serious injuries were observed in PFIB-exposed rats after 30 min. At 8 h, some signs of cell necrosis were observed. The injuries culminated at 24 h and then ameliorated. The number of apoptotic cells abnormally increased at 30 min post PFIB exposure, the maximum appeared at 24 h, and then ameliorated gradually. Western blot analysis revealed that the level of actin in the lung showed no significant changes within the first 4 h post PFIB exposure. However, it decreased at 8 h, reached a nadir at 24 h, and then recovered gradually. CONCLUSIONS: The pathological processes were in progress persistently post PFIB exposure. The early injuries probably were the result of the direct attack of PFIB and the advanced injuries probably arose from the inflammatory reaction induced by PFIB.