Evaluation of right-to-left shunt on contrast-enhanced transcranial Doppler in patent foramen ovale-related cryptogenic stroke: Research based on imaging.

BACKGROUND: Cardiogenic embolism caused by patent foramen ovale (PFO) is a common etiology of cryptogenic stroke (CS), particularly in young and middle-aged patients. Studies about right-to-left shunt (RLS) detection using contrast-enhanced transcranial Doppler (c-TCD) are numerous. According to the time phase and number of microbubbles detected on c-TCD, RLS can be classified and graded. We hypothesized that the characteristics of an infarction lesion on diffusion-weighted imaging differs when combining the type and grade of RLS on c-TCD in patients with PFO-related CS. AIM: To explore the characteristics of infarction lesions on diffusion-weighted imaging when combining the RLS type and grade determined by c-TCD. METHODS: We retrospectively evaluated CS patients from August 2015 to December 2019 at a tertiary hospital. In total, 111 PFO-related CS patients were divided according to whether RLS was permanent (microbubbles detected both at resting state and after the Valsalva maneuver) or latent (microbubbles detected only after the Valsalva maneuver) on c-TCD. Each group was subdivided into small, mild and large RLS according to the grade of shunt on c-TCD. A normal control group was composed of 33 patients who suffered from simple dizziness. Intragroup and intergroup differences were analyzed in terms of clinical, laboratory and diffusion-weighted imaging lesion characteristics. The correlation between RLS grade evaluated by c-TCD and size of PFO determined by transesophageal echocardiography were also analyzed. RESULTS: In 111 patients with PFO-related CS, 68 had permanent RLS and 43 had latent RLS. Clinical characteristics and laboratory tests were not significantly different among the permanent RLS, latent RLS and normal control groups. The proportion of patients with multiple territory lesions in the permanent RLS group (50%) was larger than that in the latent RLS group (27.91%; P = 0.021). Posterior circulation was more likely to be affected in the latent RLS group than in the permanent RLS group (30.23% vs 8.82%, P = 0.004). Permanent-large and latent-large RLS were both more likely to be related to multiple (P trend = 0.017 and 0.009, respectively), small (P trend = 0.035 and 0.006, respectively) and cortical (P trend = 0.031 and 0.033, respectively) lesions. The grade of RLS evaluated by c-TCD was correlated to the size of PFO determined by transesophageal echocardiography (r = 0.758, P < 0.001). CONCLUSION: Distribution of the infarct suggested the possible type of RLS. Multiple, small and cortical infarcts suggest large RLS induced by a large PFO.

Neurons derived from human-induced pluripotent stem cells express mu and kappa opioid receptors.

Neuroprotection studies have shown that induced pluripotent stem (iPS) cells have the possibility to transform neuroprotection research. In the present study, iPS cells were generated from human renal epithelial cells and were then differentiated into neurons. Cells in the iPS-cell group were maintained in stem cell medium. In contrast, cells in the iPS-neuron group were first maintained in neural induction medium and expansion medium containing ROCK inhibitors, and then cultivated in neuronal differentiation medium and neuronal maturation medium to induce the neural stem cells to differentiate into neurons. The expression of relevant markers was compared at different stages of differentiation. Immunofluorescence staining revealed that cells in the iPS-neuron group expressed the neural stem cell markers SOX1 and nestin on day 11 of induction, and neuronal markers TUBB3 and NeuN on day 21 of induction. Polymerase chain reaction results demonstrated that, compared with the iPS-cell group, TUBB3 gene expression in the iPS-neuron group was increased 15.6-fold. Further research revealed that, compared with the iPS-cell group, the gene expression and immunoreactivity of mu opioid receptor in the iPS-neuron group were significantly increased (38.3-fold and 5.7-fold, respectively), but those of kappa opioid receptor had only a slight change (1.33-fold and 1.57-fold increases, respectively). Together, these data indicate that human iPS cells can be induced into mu opioid receptor- and kappa opioid receptor-expressing neurons, and that they may be useful to simulate human opioid receptor function in vitro and explore the underlying mechanisms of human conditions.

Specific matrix metalloproteinases and calcification factors are associated with the vulnerability of human carotid plaque.

The rupture of atherosclerotic plaque provokes the majority of acute cerebrovascular events. Studies have demonstrated that various matrix metalloproteinases (MMPs) may promote atherosclerotic plaque progression and rupture. However, results have been incongruous and the mechanisms of this remain obscured. Therefore, in the current study, carotid plaques were characterized by assessing the levels of MMPs and calcification factors, and evaluating their association with plaque vulnerability. Human carotid plaques were obtained from carotid endarterectomies, and classified into stable and vulnerable groups by ultrasonography and histological analyses. The mRNA and protein levels of MMPs, vascular endothelial growth factor (VEGF), bone sialoprotein 2 (BSP) and osteopontin were investigated by reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. Immunohistochemistry was used to localize MMP-2 and MMP-14 in stable and vulnerable plaques. The activation of various associated signaling pathways was also investigated using western blotting. The mRNA levels of MMP-2, -7, -9 and -14 were elevated in vulnerable plaques, among which expression of MMP-2 and -14 were the highest. Consistent with the mRNA levels, the protein levels of MMP-2 and -14 were also elevated. Immunohistochemistry also demonstrated positive staining of MMP-2 and MMP-14 in vulnerable plaques. Factors that indicate neovascularization and calcification, including VEGF and BSP, were concurrently elevated in vulnerable plaques. In addition, the protein levels of extracellular regulated kinase (ERK) and protein kinase C (PKC) were upregulated in vulnerable plaques. The current study provides novel insights into the MMP profiles of vulnerability plaques, and may assist in the development of novel methods for the diagnosis of plaque vulnerability and the prevention of plaque rupture.

Chitosan and spermine enhance drought resistance in white clover, associated with changes in endogenous phytohormones and polyamines, and antioxidant metabolism.

The interaction of chitosan and polyamines (PAs) could be involved mitigating drought stress in white clover (Trifolium repens L.). This research aimed to determine the effect of chitosan and PAs, and co-application of chitosan and PAs on improving drought tolerance associated with growth, phytohormones, polyamines and antioxidant metabolism. Plants were pretreated with or without 1gL-1 chitosan, 0.5mM spermine, or 1gL-1 chitosan+0.5mM spermine, then subjected to drought induced by polyethylene glycol (PEG) 6000 (-0.5MPa) in growth chambers for 14 days. Exogenous chitosan and spermine improved the level of PAs by regulating arginine decarboxylases, S-adenosyl methionine decarboxylase, copper-containing amine oxidase and polyamine oxidase activity, and expression of the genes encoding these enzymes under drought. Application of exogenous chitosan improved ABA content under normal and drought conditions. In addition, chitosan and spermine significantly enhanced the levels of cytokinin and GA, but reduced IAA levels during drought stress. Exogenous chitosan and spermine improved antioxidant defence, including enzyme activity, gene expression and the content of ascorbate and glutathione compounds, leading to a decline in superoxide anion radicals, H2O2 and malondialdehyde, effectively mitigating drought-induced oxidative damage. Other protective metabolites, such as total phenols and flavonoids, increased considerably under application of chitosan and spermine. These results suggest that chitosan-induced drought tolerance could be involved in PA metabolism, changes in endogenous phytohormones and antioxidant defence in white clover. Co-application of chitosan and spermine was more effective than either chitosan or spermine alone in mitigating drought stress.

Exogenous Application of GABA Improves PEG-Induced Drought Tolerance Positively Associated with GABA-Shunt, Polyamines, and Proline Metabolism in White Clover.

In order to investigate the physiological effects of exogenous γ-aminobutyric acid (GABA) on drought tolerance in white clover (Trifolium repens), GABA shunt, polyamines (PAs), and proline (Pro) metabolism were examined after plants pretreated with or without GABA (8 mM) and then exposed to water or 15% PEG-induced drought stress in growth chamber. In this study, exogenous application of GABA effectively alleviated drought-induced damage in leaves, as reflected by significantly higher relative water content, lower electrolyte leakage, lipid peroxidation, and leaf wilt. Exogenous GABA further promoted drought-induced increases in GABA transaminase and alpha ketone glutarate dehydrogenase activities, but inhibited glutamate decarboxylase activity under both control and drought conditions, resulting in an increase in endogenous glutamate (Glu) and GABA content. Besides, exogenous GABA could well accelerated PAs synthesis and suppressed PAs catabolism, which lead to the extremely enhanced different types of PAs content (free Put and Spd, insoluble bound Spd and Spm, soluble conjugated Spd and Spm, and total Put, Spd and Spm) under drought stress. In addition, exogenous GABA application further activated drought-induced Δ1-pyrroline-5-carboxylate synthetase and proline dehydrogenase activities, but suppressed drought-facilitated ornithine -δ-amino transferase activities, leading to a higher Pro accumulation and metabolism in GABA-pretreated plants in the middle and last period of drought. The results suggested that increased endogenous GABA by exogenous GABA treatment could improve drought tolerance of white clover associated with a positive regulation in the GABA-shunt, PAs and Pro metabolism.

CCL17 combined with CCL19 as a nasal adjuvant enhances the immunogenicity of an anti-caries DNA vaccine in rodents.

AIM: CCL19 and its receptor CCR7 are essential molecules for facilitating the trafficking of mature dendritic cells (DCs) and helping to establish a microenvironment in lymphoid tissues to initiate primary immune responses, whereas CCL17 is required in the CCR7-CCL19-dependent migration of DCs. In this study we examined whether co-administration of CCL17 and CCL19 could enhance the immunogenicity of an anti-caries DNA vaccine, pCIA-P, in rodents. METHODS: Plasmids encoding CCL17 (pCCL17/VAX) and CCL19 (pCCL19/VAX) were constructed. BALB/c mice were intranasally administered pCCL17/VAX, pCCL19/VAX, or pCCL17/VAX plus pCCL19/VAX, the migration of DCs to the spleen and draining lymph nodes (DLNs) was assessed with flow cytometry. The mice were co-administered pCIA-P; and the anti-PAc antibodies in the serum and saliva were detected with ELISA. Wistar rats were orally challenged with Streptococcus mutans and then administered pCIA-P in combination with pCCL17/VAX, pCCL19/VAX, or pCCL17/VAX plus pCCL19/VAX. The amount of S mutans sustained on rat molar surfaces was assessed using a colony forming assay. Caries activity was scored with the Keyes method. RESULTS: Co-administration of the CCL17 and CCL19 genes in mice caused a greater increase in the number of mature DCs in the spleen and DLNs compared with administration of CCL17 or CCL19 genes alone. CCL17 and CCL19 double-adjuvant plus pCIA-P induced significantly higher levels of anti-PAc salivary IgA and anti-PAc serum IgG antibody in mice, and strengthened the ability of pCIA-P in inhibiting the colonization of S mutans on rat tooth surfaces. The caries activity of the combined adjuvant group was significantly lower than that of the pCCL17/VAX or the pCCL19/VAX group. CONCLUSION: A nasal adjuvant consisting of a combination of CCL17 and CCL19 attracts more mature DCs to secondary lymphoid tissues, inducing enhanced antibody responses against the anti-caries DNA vaccine pCIA-P and reducing S mutans infection in rodents.

De novo Transcriptome Analysis and Molecular Marker Development of Two Hemarthria Species.

Hemarthria R. Br. is an important genus of perennial forage grasses that is widely used in subtropical and tropical regions. Hemarthria grasses have made remarkable contributions to the development of animal husbandry and agro-ecosystem maintenance; however, there is currently a lack of comprehensive genomic data available for these species. In this study, we used Illumina high-throughput deep sequencing to characterize of two agriculturally important Hemarthria materials, H. compressa "Yaan" and H. altissima "1110." Sequencing runs that used each of four normalized RNA samples from the leaves or roots of the two materials yielded more than 24 million high-quality reads. After de novo assembly, 137,142 and 77,150 unigenes were obtained for "Yaan" and "1110," respectively. In addition, a total of 86,731 "Yaan" and 48,645 "1110" unigenes were successfully annotated. After consolidating the unigenes for both materials, 42,646 high-quality SNPs were identified in 10,880 unigenes and 10,888 SSRs were identified in 8330 unigenes. To validate the identified markers, high quality PCR primers were designed for both SNPs and SSRs. We randomly tested 16 of the SNP primers and 54 of the SSR primers and found that the majority of these primers successfully amplified the desired PCR product. In addition, high cross-species transferability (61.11-87.04%) of SSR markers was achieved for four other Poaceae species. The amount of RNA sequencing data that was generated for these two Hemarthria species greatly increases the amount of genomic information available for Hemarthria and the SSR and SNP markers identified in this study will facilitate further advancements in genetic and molecular studies of the Hemarthria genus.

SSRs transferability and genetic diversity of three allogamous ryegrass species.

Simple sequence repeat (SSR) markers are widely applied in studies of plant molecular genetics due to their abundance in the genome, codominant nature, and high repeatability. However, microsatellites are not always available for the species to be studied and their isolation could be time- and cost-consuming. To investigate transferability in cross-species applications, 102 primer pairs previously developed in ryegrass and tall fescue were amplified across three allogamous ryegrass species including Lolium rigidum, Lolium perenne and Lolium multiflorum. Their highly transferability (100%) were evidenced. While, most of these markers were multiple loci, only 17 loci were selected for a robust, single-locus pattern, which may be due to the recentness of the genome duplication or duplicated genomic regions, as well as speciation. A total of 87 alleles were generated with an average of 5.1 per locus. The mean polymorphism information content (PIC) and observed heterozygosity (Ho) values at genus was 0.5532 and 0.5423, respectively. Besides, analysis of molecular variance (AMOVA) revealed that all three levels contributed significantly to the overall genetic variation, with the species level contributing the least (P<0.001). Also, the unweighted pair group method with arithmetic averaging dendrogram (UPGMA), Bayesian model-based STRUCTURE analysis and the principal coordinate analysis (PCoA) showed that accessions within species always tended to the same cluster firstly and then to related species. The results showed that these markers developed in related species are transferable efficiently across species, and likely to be useful in analyzing genetic diversity.

[Clinical Application of Immune-related Response Criteria in Evaluating Chinese Medical Treatme for Advanced Non-small Cell Lung Cancer].

OBJECTIVE: To evaluate the applicability of immune-related response criteria (irRC) in treating non-small cell lung cancer (NSCLC) by Chinese medicine (CM). METHODS: Totally 97 stage III a-IV NSCLC patients were predominantly treated with comprehensive CM. Curative effects were evaluated by three methods such as Response Evaluation Criteria in Solid Tumors (RECIST), Oncologic Curative Effect Evaluation Criteria of Chinese Medicine in Solid Tumor (draft, abbreviated as CM criteria), and irRC. The correspondency and consistency between irRC, RECIST and CM criteria were analyzed and compared. The objectivity of irRC in evaluating curative effect of Chinese medical treatment for NSCLC was assessed. RESULTS: The correspondency rate of irRC to RECIST was 59. 79% with Kappa value of 0. 379 (U test, P <0. 01). The two criteria had certain correspondence, but with an unsatisfactory consistency. The correspondency rate of irRC to CM criteria rate was 83. 51% with Kappa value of 0.751 (U test, P <0. 01). The two criteria had good correspondence and consistency. CONCLUSIONS: CM criteria had good consistency with CM criteria in evaluating curative effect for Chinese medical treatment of advanced NSCLC. Its results could objectively reflect features and advantages of CM for treating advanced NSCLC.

Spermine alleviates drought stress in white clover with different resistance by influencing carbohydrate metabolism and dehydrins synthesis.

The objective of this research was to analyse whether ameliorating drought stress through exogenously applied spermine (Spm) was related to carbohydrate metabolism, dehydrins accumulation and the transcription of genes encoding dehydrins in two white clovers (drought-susceptible cv. 'Ladino' and drought-resistant cv. 'Haifa') under controlled drying conditions for 10 days. The results show that the application of Spm effectively alleviates negative effects caused by drought stress in both cultivars. Exogenous Spm led to accumulation of more water-soluble carbohydrates (WSC), sucrose, fructose and sorbitol in both cultivars under drought stress, and also significantly elevated glucose content in leaves of drought-resistant cv. 'Haifa', but had no effect on drought-susceptible cv. 'Ladino'. Accordingly, the key enzyme activities of sucrose and sorbitol metabolism changed along with the application of Spm under drought stress. Spm induced a significant increase in sucrose phosphate synthase (SPS) or sorbitol dehydrogenase (SDH) activity, but decrease in sucrose synthetase (SS) activity when two cultivars were subjected to drought. In addition, the improved accumulation of dehydrins induced by exogenous Spm coincided with three genes expression which was responsible for dehydrins synthesis. But Spm-induced transcript level of dehydrin genes increased earlier in cv. 'Ladino' than that in cv. 'Haifa'. Thus, these results suggest that ameliorating drought stress through exogenously applied Spm may be associated with increased carbohydrate accumulation and dehydrins synthesis. There are differences between drought-susceptible and -resistant white clover cultivars related to Spm regulation of WSC metabolism and dehydrins expression.

Molecular insights into the genetic diversity of Hemarthria compressa germplasm collections native to southwest China.

Start codon targeted polymorphism (SCoT) analysis was employed to distinguish 37 whipgrass (Hemarthria compressa L.) clones and assess the genetic diversity and population structure among these genotypes. The informativeness of markers was also estimated using various parameters. Using 25 highly reproducible primer sets, 368 discernible fragments were generated. Of these, 282 (77.21%) were polymorphic. The number of alleles per locus ranged from five to 21, and the genetic variation indices varied. The polymorphism information content (PIC) was 0.358, the Shannon diversity index (H) was 0.534, the marker index (MI) was 4.040, the resolving power (RP) was 6.108, and the genotype index (GI) was 0.782. Genetic similarity coefficients (GS) between the accessions ranged from 0.563 to 0.872, with a mean of 0.685. Their patterns observed in a dendrogram constructed using the unweighted pair group method with arithmetic mean analysis (UPGMA) based on GS largely confirmed the results of principal coordinate analysis (PCoA). PCoA was further confirmed by Bayesian model-based STRUCTURE analysis, which revealed no direct association between genetic relationship and geographical origins as validated by Mantel's test (r = 0.2268, p = 0.9999). In addition, high-level genetic variation within geographical groups was significantly greater than that between groups, as determined by Shannon diversity analysis, analysis of molecular variance (AMOVA) and Bayesian analysis. Overall, SCoT analysis is a simple, effective and reliable technique for characterizing and maintaining germplasm collections of whipgrass and related species.

Exogenous spermidine improves seed germination of white clover under water stress via involvement in starch metabolism, antioxidant defenses and relevant gene expression.

This study was designed to determine the effect of exogenous spermidine (Spd) (30 µM) on white clover seed germination under water stress induced by polyethylene glycol 6000. Use of seed priming with Spd improved seed germination percentage, germination vigor, germination index, root viability and length, and shortened mean germination time under different water stress conditions. Seedling fresh weight and dry weight also increased significantly in Spd-treated seeds compared with control (seeds primed with distilled water). Improved starch metabolism was considered a possible reason for this seed invigoration, since seeds primed with Spd had significantly increased α-amylase/ß-amylase activities, reducing sugar, fructose and glucose content and transcript level of ß-amylase gene but not transcript level of α-amylase gene. In addition, the physiological effects of exogenous Spd on improving seeds' tolerance to water deficit during germination were reflected by lower lipid peroxidation levels, better cell membrane stability and significant higher seed vigour index in seedlings. Enhanced antioxidant enzyme activities (superoxide dismutase, peroxidase, catalase and ascorbate peroxidase), ascorbate-glutathione cycle (ASC-GSH cycle) and transcript level of genes encoding antioxidant enzymes induced by exogenous Spd may be one of the critical reasons behind acquired drought tolerance through scavenging of reactive oxygen species (ROS) in water-stressed white clover seeds. The results indicate that Spd plays an important function as a stress-protective compound or physiological activator.

Genetic variability and population structure of the potential bioenergy crop Miscanthus sinensis (Poaceae) in Southwest China based on SRAP markers.

The genus Miscanthus has great potential as a biofuel feedstock because of its high biomass, good burning quality, environmental tolerance, and good adaptability to marginal land. In this study, the genetic diversity and the relationship of 24 different natural Miscanthus sinensis populations collected from Southwestern China were analyzed by using 33 pairs of Sequence Related Amplified Polymorphism (SRAP) primers. A total of 688 bands were detected with 646 polymorphic bands, an average of 19.58 polymorphic bands per primer pair. The average percentage of polymorphic loci (P), gene diversity (H), and Shannon's diversity index (I) among the 24 populations are 70.59%, 0.2589, and 0.3836, respectively. The mean value of total gene diversity (HT) was 0.3373±0.0221, while the allelic diversity within populations (HS) was 0.2589±0.0136 and the allelic diversity among populations (DST) was 0.0784. The mean genetic differentiation coefficient (Gst=0.2326) estimated from the detected 688 loci indicated that there was 76.74% genetic differentiation within the populations, which is consistent with the results from Analysis of Molecular Variance (AMOVA) analysis. Based upon population structure and phylogenetic analysis, five groups were formed and a special population with mixed ancestry was inferred indicating that human-mediated dispersal may have had a significant effect on population structure of M. sinensis. Evaluating the genetic structure and genetic diversity at morphological and molecular levels of the wild M. sinensis in Southwest China is critical to further utilize the wild M. sinensis germplasm in the breeding program. The results in this study will facilitate the biofuel feedstock breeding program and germplasm conservation.

Effect of naphthalene acetic acid on adventitious root development and associated physiological changes in stem cutting of Hemarthria compressa.

In order to find a way to induce rooting on cuttings of Hemarthria compressa cv. Ya'an under controlled conditions, a project was carried out to study the effect of naphthalene acetic acid (NAA) on rooting in stem cuttings and related physiological changes during the rooting process of Hemarthria compressa. The cuttings were treated with five concentrations of NAA (0, 100, 200 300, 400 mg/l) at three soaking durations (10, 20, 30 minutes), and cuttings without treatment were considered as control. Samples were planted immediately into pots after treatment. IAA-oxidase (IAAO) activity, peroxidase (POD) activity and polyphenol oxidase (PPO) activity were determined after planting. Results showed that NAA had positive effect on rooting at the concentration of 200 mg/l compared to other concentrations at 30 days after planting (DAP). Among the three soaking durations, 20 minutes (min) of 200 mg/l NAA resulted in higher percentages of rooting, larger numbers of adventitious roots and heavier root dry weight per cutting. The lowest IAAO activity was obtained when soaked at 200 mg/l NAA for 20 min soaking duration. This was consistent with the best rooting ability, indicating that the lower IAAO activity, the higher POD activity and PPO activity could be used as an indicator of better rooting ability for whip grass cuttings and might serve as a good marker for rooting ability in cuttings.

Intraoperative microvascular Doppler monitoring in intracranial aneurysm surgery.

BACKGROUND: Surgical treatment of intracranial aneurysms is often compromised by incomplete exclusion of the aneurysm or stenosis of parent vessels. Intraoperative microvascular Doppler (IMD) is an attractive, noninvasive, and inexpensive tool. The present study aimed to evaluate the usefulness and reliability of IMD for guiding clip placement in aneurysm surgery. METHODS: A total of 92 patients with 101 intracranial aneurysms were included in the study. IMD with a 1.5-mm diameter, 20-MHz microprobe was used before and after clip application to confirm aneurysm obliteration and patency of parent vessels and branching arteries. IMD findings were verified postoperatively with digital subtraction angiography (DSA) or dual energy computed tomography angiography (DE-CTA). Ninety consecutive patients, harboring 108 aneurysms, who underwent surgery without IMD was considered as the control group. RESULTS: The microprobe detected all vessels of the Circle of Willis and their major branches. Clips were repositioned in 24 (23.8%) aneurysms on the basis of the IMD findings consistent with incomplete exclusion and/or stenosis. IMD identified persistent weak blood flow through the aneurismal sac of 11 of the 101 (10.9%) aneurysms requiring clip adjustment. Stenosis or occlusion of the parent or branching arteries as indicated by IMD necessitated immediate clip adjustment in 19 aneurysms (18.8%). The mean duration of the IMD procedure was 4.8 minutes. The frequency of clip adjustment (mean: 1.8 times per case) was associated with the size and location of the aneurysm. There were no complications related to the use of IMD, and postoperative angiograms confirmed complete aneurysm exclusion and parent vessel patency. About 8.3% (9/108) aneurysms were unexpectedly incompletely occluded, and 10.2% (11/108) aneurysms and parent vessel stenosis without IMD were detected by postoperative DSA or DE-CTA. IMD could reduce the rate of residual aneurysm and unanticipated vessel stenosis which demonstrated statistically significant advantages compared with aneurysm surgery without IMD. CONCLUSION: IMD is a safe, easily performed, reliable, and valuable tool that is suitable for routine use in intracranial surgery, especially in complicated, large, and giant aneurysms with wide neck or without neck.

Co-delivery of ccl19 gene enhances anti-caries DNA vaccine pCIA-P immunogenicity in mice by increasing dendritic cell migration to secondary lymphoid tissues.

AIM: To investigate how co-delivery of the gene encoding C-C chemokine ligand-19 (CCL-19) affected the systemic immune responses to an anti-caries DNA vaccine pCIA-P in mice. METHODS: Plasmid encoding CCL19-GFP fusion protein (pCCL19/GFP) was constructed by inserting murine ccl19 gene into GFP-expressing vector pAcGFP1-N1. Chemotactic effect of the fusion protein on murine dendritic cells (DCs) was assessed in vitro and in vivo using transwell and flow cytometric analysis, respectively. BALB/c mice were administered anti-caries DNA vaccine pCIA-P plus pCCL19/GFP (each 100 µg, im) or pCIA-P alone. Serum level of anti-PAc IgG was assessed with ELISA. Splenocytes from the mice were stimulated with PAc protein for 48 h, and IFN-γ and IL-4 production was measured with ELISA. The presence of pCCL19/GFP in spleen and draining lymph nodes was assessed using PCR. The expression of pCCL19/GFP protein in these tissues was analyzed under microscope and with flow cytometry. RESULTS: The expression level of CCL19-GFP fusion protein was considerably increased 48 h after transfection of COS-7 cells with pCCL19/GFP plasmids. The fusion protein showed potent chemotactic activity on DCs in vitro. The level of serum PAc-specific IgG was significantly increased from 4 to 14 weeks in the mice vaccinated with pCIA-P plus pCCL19/GFP. Compared to mice vaccinated with pCIA-P alone, the splenocytes from mice vaccinated with pCIA-P plus pCCL19/GFP produced significantly higher level of IFN-γ, but IL-4 production had no significant change. Following intromuscular co-delivery, pCCL19/GFP plasmid and fusion protein were detected in the spleen and draining lymph nodes. Administration of CCL19 gene in mice markedly increased the number of mature DCs in secondary lymphoid tissues. CONCLUSION: CCL19 serves as an effective adjuvant for anti-caries DNA vaccine by inducing chemotactic migration of DCs to secondary lymphoid tissues.

[The feasibility of training anesthesiologists with hemodynamic models of miniature pigs].

OBJECTIVE: To investigate the feasibility of training anesthesiologists with hemodynamic models of miniature pigs. METHODS: Eight miniature experimental pigs were chosen; ECG, SpO2, PetCO2, ABP, CVP, SV, SVV and PAP were monitored after anesthesia. The research was divided into two parts. Part one: the blood of pig was taken from its artery, air was injected into its body thought jugular vein, 15%KCl and 0.75% bupivacaine were given by peripheral vein respectively. The resuscitation would not be implemented unless the monitoring data changes were significantly and were recorded. Trainees looked on only during that time. Part two: Trainees were trained for placement of Swan-Ganz catheter after the sheath had been success- fully intubated into pig's right jugular vein. Trainees were trained for placement of femoral artery guided by ultrasound.Scores related to trainees' performance were written down. RESULTS: (1) Monitoring data (ABP, SV, SVV, CVP, PetCO2, PAP, SpO2 , ECG) changes were significantly ( P < 0.05 or P < 0.01), which indicated that the models had been successfully established. The evaluation scale displays: 81.3% of trainees thought this research improved their under- standing of hemodynamic changes; 78.2% thought that it helped them know how to deal with these circumstances; 71.9% thought this training was meaningful. (2) It was improved for students' skill to place Swan-Ganz catheter (P < 0.05), whereas the skill for placement of artery catheter by ultrasound was not significantly improved (P > 0.05). CONCLUSION: It was feasible to use a hemodynamic model of miniature pigs as an assisted teaching method to improve resident anesthesiologists' ability to understand and cope with clinical hemodynamic changes.