KI-Catalyzed Oxidative Cyclization of Enamines and tBuONO to Access Functional Imidazole-4-Carboxylic Derivatives.

A simple protocol for the noble-metal-free oxidative cyclization of enamines and tBuONO has been developed. This KI-mediated formal [4 + 1] cycloamination reaction provides a practical strategy for the synthesis of imidazole-4-carboxylic derivatives using tBuONO both as an aminating reagent and oxidant. The reaction features wide substrate scope and good functional tolerance for enamine compounds, even the unactivated ones.

IBX-Promoted Oxidative Cyclization of N-Hydroxyalkyl Enamines: A Metal-Free Approach toward 2,3-Disubstituted Pyrroles and Pyridines.

An iodoxybenzoic acid-mediated selected oxidative cyclization of N-hydroxyalkyl enamines was developed. Through this strategy, a variety of 2,3-disubstituted pyrroles and pyridines were produced in good selectivity involving oxidation of alcohol, followed by condensation of aldehyde and α-C of enamines. Furthermore, this metal-free method has several advantages, including the use of environmentally friendly reagents, broad substrate scope, mild reaction conditions, and high efficiency.

Botulinum Toxin A Injection Using Esophageal Balloon Radiography Combined with CT Guidance for the Treatment of Cricopharyngeal Dysphagia.

Cricopharyngeal dysfunction, especially cricopharyngeal achalasia, is a common cause of dysphagia, while patients with brainstem stroke and medullary damage have a relatively high risk of cricopharyngeal achalasia. The aim of this article was to introduce an improved method of CT-guided method of injecting botulinum toxin A into the cricopharyngeus muscle using esophageal balloon radiography, and to assess the effect of the botulinum toxin A injection on swallowing performance. Seventeen patients with cricopharyngeal dysphagia were treated with botulinum toxin A injection using esophageal balloon radiography combined with CT guidance to the cricopharyngeal muscle. Primary outcome measures, including Functional Oral Intake Scale and Deglutition Handicap Index, were performed at baseline, 1 week, and 1 month after treatment. The Levene method was used to test the homogeneity of variance, and the Kruskal-Wallis test was used to compare the scores between the timepoints. Botulinum toxin A injection resulted in obvious improvement in 15 patients (88.2%) and no improvement in two patients (11.8%). Compared with the scores prior to treatment, the Functional Oral Intake Scale and Deglutition Handicap Index scores were significantly improved at 1 week (P < 0.001 and P = 0.008, respectively) and 1 month after the treatment (P = 0.001 and P < 0.001, respectively). Thus, CT-guided percutaneous injection of botulinum toxin A is probably a relatively safe, well-tolerated, and viable technique for the treatment of cricopharyngeal dysphagia caused by brainstem injury. Localization with a balloon radiography made the needle guidance easier to visualize.

Gene therapy by lentivirus-mediated RNA interference targeting extracellular-regulated kinase alleviates neuropathic pain in vivo.

BACKGROUNDS: Neuropathic pain is an abnormal pain, which is related to the activation of extracellular-regulated kinase (ERK) signaling. This study was to investigate the effects of ERK knockdown via lentivirus-mediated RNA interference on allodynia in rats with chronic compression of the dorsal root ganglia (DRG) and to uncover the potential mechanisms. METHODS: The model of chronic compression of the dorsal root ganglia (CCD) was established in rats by surgery. Gene silence was induced by injecting rats with lentivirus expressing ERK short hairpin RNA (shRNA). Behavioral test was performed by calculating paw withdrawal mechanical threshold (PWMT) and thermal paw withdrawal latency (TPWL). RESULTS: We firstly generated lentivirus expressing ERK shRNA to downregulate ERK gene expression both in vitro and in vivo by using Western blot analysis and quantitative reverse transcription polymerase chain reaction. In CCD, ERK mRNA, and protein levels in DRG neurons were dramatically increased, accompanied with decreased PWMT and TPWL. Lentivirus-mediated RNA interference decreased ERK gene expression in DRG neurons and normalized the PWMT and TPWL in CCD rats, but not in rats infected with lentivirus expressing negative control shRNA. Further, calcium responses of DRG neurons to the hypotonic solution and 4α-phorbol 12,13-didecanoate were enhanced in CCD rats, which were suppressed by lentivirus-mediated ERK gene silence. Finally, the levels of transient receptor potential vanilloid 4 gene expressions in DRG neurons and L4 to L5 spinal cord isolated from CCD rats were dramatically upregulated, which were reversed by lentivirus-mediated ERK gene knockdown. CONCLUSION: Lentivirus-mediated RNA interference (RNAi) silencing targeting ERK might reverse CCD-induced neuropathic pain in rats through transient receptor potential vanilloid 4.

Cloning and characterization of the pepper CaPAO gene for defense responses to salt-induced leaf senescence.

BACKGROUND: Pheophorbide a oxygenase (PAO) is an important enzyme in the chlorophyll catabolism pathway and is involved in leaf senescence. It opens the porphyrin macrocycle of pheophorbide a and finally forms the primary fluorescent chlorophyll catabolite. Previous studies have demonstrated the function of PAO during cell death. However, the characterizaton of PAO during leaf senescence induced by environmental factors is not well understood. METHODS: Homology-based cloning and RACE techniques were used to obtain the full-length cDNA of the CaPAO gene. CaPAO expression was determined by quantitative real-time PCR. Function of CaPAO gene were studied using virus-induced gene silencing and transgenic techniques with tobacco plants (Nicotiana tabacum). RESULTS: A novel PAO gene CaPAO was isolated from pepper (Capsicum annuum L.). The full-length CaPAO cDNA is comprised of 1838 bp, containing an open reading frame of 1614 bp, and encodes a 537 amino acid protein. This deduced protein belongs to the Rieske-type iron-sulfur superfamily, containing a conserved Rieske cluster. CaPAO expression, as determined by quantitative real-time PCR, was higher in leaves than roots, stems and flowers. It was upregulated by abscisic acid, methyl jasmonate and salicylic acid. Moreover, CaPAO was significantly induced by high salinity and osmotic stress treatments and also was regulated by Phytophthora capsici. The virus-induced gene silencing technique was used to silence the CaPAO gene in pepper plants. After 3 days of high salt treatment, the chlorophyll breakdown of CaPAO-silenced pepper plants was retarded. RD29A promoter-inducible expression vector was constructed and transferred into tobacco plant. After 7 days of salt treatment, the leaves of transgenic plants were severely turned into yellow, the lower leaves showed necrotic symptom and chlorophyll content was significantly lower than that in the control plants. CONCLUSIONS: The expression of CaPAO gene was induced in natural senescence and various stresses. The CaPAO gene may be related to defense responses to various stresses and play an important role in salt-induced leaf senescence.

Silencing of the CaCP gene delays salt- and osmotic-induced leaf senescence in Capsicum annuum L.

Cysteine proteinases have been known to participate in developmental processes and in response to stress in plants. Our present research reported that a novel CP gene, CaCP, was involved in leaf senescence in pepper (Capsicum annuum L.). The full-length CaCP cDNA is comprised of 1316 bp, contains 1044 nucleotides in open reading frame (ORF), and encodes a 347 amino acid protein. The deduced protein belongs to the papain-like cysteine proteases (CPs) superfamily, containing a highly conserved ERFNIN motif, a GCNGG motif and a conserved catalytic triad. This protein localized to the vacuole of plant cells. Real-time quantitative PCR analysis revealed that the expression level of CaCP gene was dramatically higher in leaves and flowers than that in roots, stems and fruits. Moreover, CaCP transcripts were induced upon during leaf senescence. CaCP expression was upregulated by plant hormones, especially salicylic acid. CaCP was also significantly induced by abiotic and biotic stress treatments, including high salinity, mannitol and Phytophthora capsici. Loss of function of CaCP using the virus-induced gene-silencing technique in pepper plants led to enhanced tolerance to salt- and osmotic-induced stress. Taken together, these results suggest that CaCP is a senescence-associated gene, which is involved in developmental senescence and regulates salt- and osmotic-induced leaf senescence in pepper.

Overexpression of the CaTIP1-1 pepper gene in tobacco enhances resistance to osmotic stresses.

Both the gene expression and activity of water channel protein can control transmembrane water movement. We have reported the overexpression of CaTIP1-1, which caused a decrease in chilling tolerance in transgenic plants by increasing the size of the stomatal pore. CaTIP1-1 expression was strongly induced by salt and mannitol stresses in pepper (Capsicum annuum). However, its biochemical and physiological functions are still unknown in transgenic tobacco. In this study, transient expression of CaTIP1-1-GFP in tobacco suspension cells revealed that the protein was localized in the tonoplast. CaTIP1-1 overexpressed in radicle exhibited vigorous growth under high salt and mannitol treatments more than wild-type plants. The overexpression of CaTIP1-1 pepper gene in tobacco enhanced the antioxidant enzyme activities and increased transcription levels of reactive oxygen species-related gene expression under osmotic stresses. Moreover, the viability of transgenic tobacco cells was higher than the wild-type after exposure to stress. The pepper plants with silenced CaTIP1-1 in P70 decreased tolerance to salt and osmotic stresses using the detached leaf method. We concluded that the CaTIP1-1 gene plays an important role in response to osmotic stresses in tobacco.

Effects of different fertilization rates on growth, yield, quality and partial factor productivity of tomato under non-pressure gravity irrigation.

To select the optimum fertilizer application under specific irrigation levels and to provide a reliable fertigation system for tomato plants, an experiment was conducted by using a microporous membrane for water-fertilizer integration under non-pressure gravity. A compound fertilizer (N:P2O5:K2O, 18:7:20) was adopted for topdressing at four levels, 1290 kg/ha, 1140 kg/ha, 990 kg/ha, and 840 kg/ha, and the locally recommended level of 1875 kg/ha was used as the control to explore the effects of different fertilizer application rates on growth, nutrient distribution, quality, yield, and partial factor of productivity (PFP) in tomato. The new regime of microporous membrane water-fertilizer integration under non-pressure gravity irrigation reduced the fertilizer application rate while promoting plant growth in the early and intermediate stages. Except for the 990 kg/ha fertilizer treatment, yields per plant and per plot for each fertilizer application rate were higher than or equal to those of the control. The new regime could effectively improve PFP and reduce soil nutrient enrichment. Fertilizer at 840 kg/ha showed the optimum results by increasing PFP by 75.72% as compared to control. In conclusion, the fertilizer rate at 840 kg/ha has not only maintained the productivity of soil but also tomato growth and quality of fruit which makes the non-pressure gravity irrigation a potential and cost-effective way for fertilizer application.

DOK3 is involved in microglial cell activation in neuropathic pain by interacting with GPR84.

Adaptor molecule downstream of kinase-3 (DOK3) is a vital regulator of innate immune responses in macrophages and B cells, and G-protein-coupled receptor 84 (GPR84) is significant in mediating the biosynthesis and maintenance of inflammatory mediators that are induced by neuropathic pain in microglia. In the present study, we determined the role of DOK3 in activating microglia-induced neuropathic pain and investigated the underlying mechanisms associated with GPR84. We found that knockdown of DOK3 in microglial cells dramatically reduced the levels of inflammatory factors, and we uncovered a physical association between DOK3 and GPR84 in the induction of inflammatory responses. We also observed that neuropathic pain and inflammatory responses induced by chronic constriction injury (CCI) of the sciatic nerve or intrathecal injection of a GPR84 agonist were compromised in DOK3-/- mice in vivo. Finally, enforced expression of DOK3 provoked inflammatory responses, and administration of pregabalin relieved neuropathic pain via inhibition of DOK3 expression. In conclusion, DOK3 induced neuropathic pain in mice by interacting with GPR84 in microglia. We hypothesize that targeting the adaptor protein DOK3 may open new avenues for pharmaceutical approaches to the alleviation of neuropathic pain in the spinal cord.

[Effects of sub-low temperature and drought stress on water transport and morphological anatomy of tomato plant]. / äºšä½Žæ¸©ä¸Žå¹²æ—±èƒè¿«å¯¹ç•ªèŒ„æ¤æ ªæ°´åˆ†ä¼ è¾“å’Œå½¢æ€è§£å‰–ç»“æž„çš„å½±å“.

We explored the effects of sub-low temperature and drought on water transport in tomato seedlings under normal temperature (25 ℃ day/18 ℃ night) and sub-low temperature (15 ℃ day/8 ℃ night) within the artificial climate chamber, and under normal irrigation (75%-85% field water holding capacity) and drought treatment (55%-65% field water holding capacity). We analyzed the effects of temperature and soil moisture on water transport, stomata and xylem vessel morpholo-gical and anatomical structure of tomato plants. The results showed that compared with condition of normal temperature + normal irrigation, drought treatment significantly reduced leaf water potential, transpiration rate, stomatal conductance, hydraulic conductance, sap flow rate, stomatal length, and diameter of leaf, stem and root conduit, and thus thickened the cell wall and enhanced the anti-embolism ability of conduit in leaf, stem and root. Leaf water potential, transpiration rate, stomatal conductance, hydraulic conductance, and conduit diameter in leaf, stem and root were significantly reduced by sub-low temperature treatment, but the stomata became larger, cell wall was thickened and the anti-embolism ability was enhanced in leaf, stem and root conduit. Under sub-low temperature condition, soil moisture did not affect leaf water potential, transpiration rate, stomatal conductance, hydraulic conductance, stomatal morphology, conduit structure of leaf and root. In conclusion, under drought treatment, the homeostasis in water relationship was obtained by the coordination of leaf, stem and root structure. Under sub-low temperature treatment, the regulation of water relationship was mainly dependent on the changes of conduit structure in leaf and root, which was less affected by soil moisture.

Constraint-induced movement therapy in treatment of acute and sub-acute stroke: a meta-analysis of 16 randomized controlled trials.

OBJECTIVE: The aim of this meta-analysis was to evaluate the clinical efficacy of constraint-induced movement therapy in acute and sub-acute stroke. DATA SOURCES: The key words were stroke, cerebrovascular accident, constraint-induced therapy, forced use, and randomized controlled trial. The databases, including China National Knowledge Infrastructure, WanFang, Weipu Information Resources System, Chinese Biomedical Literature Database, PubMed, Medline, Embase, the Cochrane Central Register of Controlled Trials, and the Cochrane Database of Systematic Reviews, were searched for studies on randomized controlled trials for treating acute or sub-acute stroke published before March 2016. DATA SELECTION: We retrieved relevant randomized controlled trials that compared constraint-induced movement therapy in treatment of acute or sub-acute stroke with traditional rehabilitation therapy (traditional occupational therapy). Patients were older than 18 years, had disease courses less than 6 months, and were evaluated with at least one upper extremity function scale. Study quality was evaluated, and data that met the criteria were extracted. Stata 11.0 software was used for the meta-analysis. OUTCOME MEASURES: Fugl-Meyer motor assessment of the arm, the action research-arm test, a motor activity log for amount of use and quality of movement, the Wolf motor function test, and a modified Barthel index. RESULTS: A total of 16 prospective randomized controlled trials (379 patients in the constraint-induced movement-therapy group and 359 in the control group) met inclusion criteria. Analysis showed significant mean differences in favor of constraint-induced movement therapy for the Fugl-Meyer motor assessment of the arm (weighted mean difference (WMD) = 10.822; 95% confidence intervals (95% CI): 7.419-14.226), the action research-arm test (WMD = 10.718; 95% CI: 5.704-15.733), the motor activity log for amount of use and quality of movement (WMD = 0.812; 95% CI: 0.331-1.293) and the modified Barthel index (WMD = 10.706; 95% CI: 4.417-16.966). CONCLUSION: Constraint-induced movement therapy may be more beneficial than traditional rehabilitation therapy for improving upper limb function after acute or sub-acute stroke.

Effect of TRPV4-p38 MAPK Pathway on Neuropathic Pain in Rats with Chronic Compression of the Dorsal Root Ganglion.

The aim of this study was to investigate the relationships among TRPV4, p38, and neuropathic pain in a rat model of chronic compression of the dorsal root ganglion. Mechanical allodynia appeared after CCD surgery, enhanced via the intrathecal injection of 4α-phorbol 12,13-didecanoate (4α-PDD, an agonist of TRPV4) and anisomycin (an agonist of p38), but was suppressed by Ruthenium Red (RR, an inhibitor of TRPV4) and SB203580 (an inhibitor of p38). The protein expressions of p38 and P-p38 were upregulated by 4α-PDD and anisomycin injection but reduced by RR and SB203580. Moreover, TRPV4 was upregulated by 4α-PDD and SB203580 and downregulated by RR and anisomycin. In DRG tissues, the numbers of TRPV4- or p38-positive small neurons were significantly changed in CCD rats, increased by the agonists, and decreased by the inhibitors. The amplitudes of ectopic discharges were increased by 4α-PDD and anisomycin but decreased by RR and SB203580. Collectively, these results support the link between TRPV4 and p38 and their intermediary role for neuropathic pain in rats with chronic compression of the dorsal root ganglion.

Cloning and expression analysis of CaPIP1-1 gene in pepper (Capsicum annuum L.).

Plant aquaporins are responsible for water transmembrane transport, which play an important role on abiotic and biotic stresses. A novel plasma membrane intrinsic protein of CaPIP1-1 was isolated from the pepper P70 according to transcriptome databases of Phytophthora capsici inoculation and chilling stress library. CaPIP1-1, which is 1155 bp in length with an open reading frame of 861 bp, encoded 286 amino acids. Three introns, exhibited CT/AC splice junctions, were observed in CaPIP1-1. The numbers and location of introns in CaPIP1-1 were the same as observed in tomato and potato. CaPIP1-1 was abundantly expressed in pepper fruit. Increased transcription levels of CaPIP1-1 were found in the different stresses, including chilling stress, salt stress, mannitol stress, salicylic acid, ABA treatment and Phytophthora capsici infection. The expression of CaPIP1-1 was downregulated by 50 µM HgCl2 and 100 µM fluridone. The pepper plants silenced CaPIP1-1 in cv. Qiemen showed growth inhibition and decreased tolerance to salt and mannitol stresses using detached leaf method.

Interaction of transient receptor potential vanilloid 4 with annexin A2 and tubulin beta 5.

The aim of the present study was to investigate if there exists an interaction of TRPV4 with annexin A2 and with tubulin beta 5 in transfected human embryonic kidney (HEK293) cells in vitro. Coimmunoprecipitation of the rat dorsal root ganglion was performed to validly conform the interaction of TRPV4 with the other two proteins. Gene fragments coding for the amino acids in protein were obtained. We conducted coimmunoprecipitation and immunofluorescence on the transfected cell samples. Coimmunoprecipitation experiments of transfected HEK293 cells revealed that TRPV4 and tubulin beta 5 associated together in a complex, whereas TRPV4 and annexin A2 did not. The immunofluorescence microscopy revealed a colocalization of TRPV4 with both the tubulin beta 5 and annexin A2. These results indicate an interaction between TRPV4 and tubulin beta 5 by associating together. However, the association between TRPV4 and annexin A2 may be mediated by some intermediate elements or just exists in some physiological conditions. Thus, TRPV4 channel function may be modulated by tubulin beta 5 and annexin A2 and their interactions may play a role in the mechanosensation in the pathogenesis of neuropathic pain.