[Identification and functional analysis of jasmonic acid signaling repressor McJAZ8 gene in Mentha canadensis].

Mentha canadensis, as a plant with medicinal and culinary uses, holds significant economic value. Jasmonic acid signaling repressor JAZ protein has a crucial role in regulating plant response to adversity stresses. The M. canadensis McJAZ8 gene is cloned and analyzed for protein characterization, protein interactions, and expression patterns, so as to provide genetic resources for molecular breeding of M. canadensis for stress tolerance. This experiment will analyze the protein structural characteristics, subcellular localization, protein interactions, and gene expression of McJAZ8 using bioinformatics, yeast two-hybrid(Y2H), transient expression in tobacco leaves, qRT-PCR, and other technologies. The results show that:(1)The full length of the McJAZ8 gene is 543 bp, encoding 180 amino acids. The McJAZ8 protein contains conserved TIFY and Jas domains and exhibits high homology with Arabidopsis thaliana AtJAZ1 and AtJAZ2.(2)The McJAZ8 protein is localized in the nucleus and cytoplasm.(3)The Y2H results show that McJAZ8 interacts with itself or McJAZ1/3/4/5 proteins to form homologous or heterologous dimers.(4)McJAZ8 is expressed in different tissue, with the highest expression level in young leaves. In terms of leaf sequence, McJAZ8 shows the highest expression level in the fourth leaf and the lowest expression level in the second leaf.(5) In leaves and roots, the expression of McJAZ8 is upregulated to varying degrees under methyl jasmonate(MeJA), drought, and NaCl treatments. The expression of McJAZ8 shows an initial upregulation followed by a downregulation pattern under CdCl_2 treatment. In leaves, the expression of McJAZ8 tends to gradually decrease under CuCl_2 treatment, while in roots, it initially decreases and then increases before decreasing again. In both leaves and roots, the expression of McJAZ8 is downregulated to varying degrees under AlCl_(3 )treatment. This study has enriched the research on jasmonic acid signaling repressor JAZ genes in M. canadensis and provided genetic resources for the molecular breeding of M. canadensis.

[Cloning and expression pattern analysis of abscisic acid receptor gene McPYL4 in Mentha canadensis].

Mentha canadensis is a traditional Chinese herb with great medicinal and economic value. Abscisic acid(ABA) receptor PYLs have important roles in plant growth and development and response to adversity. The M. canadensis McPYL4 gene was cloned, and its protein characteristics, gene expression, and protein interactions were analyzed, so as to provide genetic resources for genetic improvement and molecular design breeding for M. canadensis resistance. Therefore, the protein characteristics, subcellular localization, gene expression pattern, and protein interactions of McPYL4 were analyzed by bioinformatics analysis, transient expression of tobacco leaves, RT-qPCR, and yeast two-hybrid(Y2H) techniques. The results showed that the McPYL4 gene was 621 bp in length, encoding 206 amino acids, and its protein had the conserved structural domain of SRPBCC and was highly homologous with Salvia miltiorrhiza SmPYL4. McPYL4 protein was localized to the cell membrane and nucleus. The McPYL4 gene was expressed in all tissue of M. canadensis, with the highest expression in roots, followed by leaves, and it showed a pattern of up-regulation followed by down-regulation in leaves 1-8. In both leaves and roots, the McPYL4 gene responded to the exogenous hormones ABA, MeJA, and the treatments of drought, AlCl_3, NaCl, CdCl_2, and CuCl_2. Moreover, McPYL4 was up-regulated for expression in both leaves and roots under the MeJA treatment, as well as in leaves treated with AlCl_3 stress for 1 h, whereas McPYL4 showed a tendency to be down-regulated in both leaves and roots under other treatments. Protein interactions showed that McPYL4 interacted with AtABI proteins in an ABA-independent manner. This study demonstrated that McPYL4 responded to ABA, JA, and several abiotic stress treatments, and McPYL4 was involved in ABA signaling in M. canadensis and thus in the regulation of leaf development and various abiotic stresses in M. canadensis.

Cross-sectional study on prevalence and risk factors for falls among the elderly in communities of Guangdong province, China.

OBJECTIVE: This study aims to investigate the prevalence and risk factors of falls among the elderly in Guangdong, China. METHODS: A cross-sectional study was conducted in six communities of Guangdong province. People over 60 years old were selected with multistage random-cluster sampling. Data on falls within the previous 12 months and fall-related risk factors were collected through a face-to-face interview. RESULTS: The prevalence of falls among older adults was 11.9% (95% CI: 11.0% to 12.8%) among 5374 interviewees. The common injuries caused by falls were bruises/scrapes (40.0%) and fractures (15.5%), and most people fall while doing housework (35.0%). Univariate analysis showed that 14 factors were associated with falls among older adults, including gender, age, residence, occupation, education level, balance ability, situation of cognition, disease, depression, living arrangement, marital status, the behaviour of exercise, drinking and drug use (p<0.05). Multivariate analysis showed that the associated factors of falls among older adults included woman (OR=1.68, 95% CI: 1.40 to 2.02), age from 70 to 79 years (OR=1.31, 95% CI: 1.09 to 1.58), age over 80 (OR=1.63, 95% CI: 1.25 to 2.13), impaired balance ability (OR=1.45, 95% CI: 1.20 to 1.75), exercise several times per month (OR=1.69, 95% CI: 1.13 to 2.53), polypharmacy (OR=1.54, 95% CI: 1.19 to 2.00), cognition impairment (OR=1.35, 95% CI: 1.08 to 1.69), mild depression (OR=1.89, 95% CI: 1.47 to 2.45) and moderate depression (OR=3.07, 95% CI: 1.99 to 4.73). CONCLUSIONS: The hazards caused by falls to the elderly in China cannot be ignored. A multidimensional customised fall prevention programme should be considered to reduce the risk of falls among the elderly based on the results above.

Association of ACE I/D gene polymorphism with vesicoureteral reflux susceptibility in children: a meta-analysis.

BACKGROUND AND OBJECTIVE: Many studies have been conducted to investigate the association between angiotensin-converting enzyme (ACE) insertion (I)/deletion (D) gene polymorphism and vesicoureteral reflux (VUR) susceptibility. However, the results from those studies are still conflicting. We performed a meta-analysis of studies relating the ACE I/D gene polymorphism to the risk of VUR. METHOD: We searched the databases of PubMed, Embase, Cochrane Library and CBM-disc (China Biological Medicine Database) as of 1 March 2011, and recruited the eligible investigations for this meta-analysis. RESULTS: Ten investigations were identified for the analysis of association between ACE I/D gene polymorphism and VUR risk: six in Caucasians, three in East-Asians and one in a Turkish population. All the investigations were performed in children. There was no marked association between ACE I/D gene polymorphism and VUR susceptibility/renal scar for overall populations, Caucasians and East-Asians. In the Turkish population, D allele and DD genotype were associated with the VUR susceptibility/renal scar. Furthermore, ACE I/D gene polymorphism was not associated with VUR progression. CONCLUSIONS: D allele and DD genotype are risk factors for the VUR susceptibility/renal scar in Turkish children. However, more case-control association investigations on larger, stratified populations are required in the future.

Relationship between angiotensin-converting enzyme insertion/deletion gene polymorphism and systemic lupus erythematosus/lupus nephritis: a systematic review and metaanalysis.

OBJECTIVE: Results from studies of the association between angiotensin-converting enzyme (ACE) insertion/deletion (I/D) gene polymorphism and systemic lupus erythematosus (SLE)/lupus nephritis (LN) are controversial. We performed this metaanalysis to evaluate the relationship between ACE I/D gene polymorphism and SLE/LN and to explore whether the ACE D allele or DD genotype could become a predictive marker for risk of SLE/LN. METHODS: Association studies were identified from the databases of PubMed, Embase, Cochrane Library and CBM-disc (China Biological Medicine Database) as of May 1, 2011, and eligible investigations were synthesized using a metaanalysis method. Results were expressed with OR for dichotomous data, and 95% CI were calculated. RESULTS: Sixteen investigations were identified for the analysis of association between ACE I/D gene polymorphism and SLE, consisting of 1959 patients with SLE and 2078 controls. In the overall populations, there was a marked association between D allele or DD genotype and SLE susceptibility (D: OR 1.29, 95% CI 1.04-1.58, p = 0.02; DD: OR 1.60, 95% CI 1.17-2.19, p = 0.003), and DD homozygous was associated with LN risk (OR 2.78, 95% CI 1.26-6.11, p = 0.01). In the subgroup analysis, DD genotype associated with SLE risk was observed in Asians; no other association was found in Asians, whites, Africans, and Brazilians. CONCLUSION: D allele and DD homozygous are significant genetic molecular markers to predict SLE susceptibility, and DD genotype is a valuable marker to predict the LN risk. More investigations are required to clarify the association of the D allele or DD homozygous with SLE/LN susceptibility.

Curcumin protects intestinal mucosal barrier function of rat enteritis via activation of MKP-1 and attenuation of p38 and NF-κB activation.

BACKGROUND: Intestinal mucosa barrier (IMB) dysfunction results in many notorious diseases for which there are currently few effective treatments. We studied curcumin's protective effect on IMB and examined its mechanism by using methotrexate (MTX) induced rat enteritis model and lipopolysaccharide (LPS) treated cell death model. METHODOLOGY/PRINCIPAL FINDINGS: Curcumin was intragastrically administrated from the first day, models were made for 7 days. Cells were treated with curcumin for 30 min before exposure to LPS. Rat intestinal mucosa was collected for evaluation of pathological changes. We detected the activities of D-lactate and diamine oxidase (DAO) according to previous research and measured the levels of myeloperoxidase (MPO) and superoxide dismutase (SOD) by colorimetric method. Intercellular adhesion molecule-1 (ICAM-1), tumor necrosis factor α (TNF-α) and interleukin 1ß (IL-1ß) were determined by RT-PCR and IL-10 production was determined by ELISA. We found Curcumin decreased the levels of D-lactate, DAO, MPO, ICAM-1, IL-1ß and TNF-α, but increased the levels of IL-10 and SOD in rat models. We further confirmed mitogen-activated protein kinase phosphatase-1 (MKP-1) was activated but phospho-p38 was inhibited by curcumin by western blot assay. Finally, NF-κB translocation was monitored by immunofluorescent staining. We showed that curcumin repressed I-κB and interfered with the translocation of NF-κB into nucleus. CONCLUSIONS/SIGNIFICANCE: The effect of curcumin is mediated by the MKP-1-dependent inactivation of p38 and inhibition of NF-κB-mediated transcription. Curcumin, with anti-inflammatory and anti-oxidant activities may be used as an effective reagent for protecting intestinal mucosa barrier and other related intestinal diseases.

[Effect of simvastatin on monocyte CX3CR1 expression in patients with acute coronary syndrome].

OBJECTIVE: To observe the effect of simvastatin on expression of CX3CR1 in the monocytes in patients with acute coronary syndrome and investigate the non-lipid mechanisms of statins against atherosclerosis. METHODS: The expression of CX3CR1 in the monocytes was measured by quantitative real-time RT-PCR in 63 patients with acute coronary syndrome confirmed by coronary arteriography after treatment with simvastatin at 10(-7) approximately 10(-5) mol/L for 4, 8 and 12 h, respectively. RESULTS: CX3CR1 expression in the monocytes treated with different concentrations of simvastatin was significantly lower than that in the control cells (P<0.05), and the expression in the cells treated with the agent for different time lengths was also significantly lower than that in the control cells (P<0.01). CONCLUSION: Simvastatin can reduce CX3CR1 expression in the monocytes of the patients with acute coronary syndrome in a concentration- and time-dependent manner, so as to reduce the inflammation and stabilize the vascular plaques.

[Subcellular distribution and translocation of hepatitis B virus core protein in HepG2.2.15 cells].

OBJECTIVES: The hepatitis B virus core protein has been found in nuclei, cytoplasm, or both of hepatocytes transfected with HBV DNA. It is still unclear whether intact core particles could pass through nuclear pores and what could be the mechanism regulating the subcellular localization of the core protein. This study on the distribution of core protein in hepatocytes and its translocation has a potential advantage to learn more about the HBV life cycle. METHODS: Dimethyl sulphoxide (DMSO, 2%), which effects hepatic differentiation, and/or 1 micro mol/L heteroaryldihydropyrimidine Bay41-4109, which interferes with the assembly of core particles, were added into HepG2.2.15 cell culture system for 4 days. The hepatitis B virus core antigen (HBcAg) and hepatitis B virus surface antigen (HBsAg) were stained with fluorescent immunocytochemistry and then observed under a confocal microscope. HBcAg in cytoplasm and nuclei were respectively extracted and analyzed using Western blot. HBV covalently closed circular DNA (cccDNA) was detected by using selective PCR method. RESULTS: The HBcAg was mostly expressed in the cytoplasm and weak signals of cccDNA were detected in the control HepG2.2.15 cells. After DMSO treatment, the expression of HBcAg in cytoplasm was increased about 2.5-fold; the expression of HBcAg and cccDNA in nuclei also increased. With the use of Bay41-4109, the signal of HBcAg in cytoplasm decreased 2/3, but it increased in the nuclei, and cccDNA decreased in the nuclei. When the HepG2.2.15 cells were treated both with DMSO and Bay41-4109, cord-liked distribution of HBsAg was observed in the cytoplasm. HBcAg in cytoplasm was decreased 1/2 but the HBcAg in the nuclei increased about 5-fold, whereas the cccDNA was almost negative. CONCLUSION: In HepG2.2.15 cells, the core protein is mainly assembled as a formation of core particles in the cytoplasm and they are blocked by the nuclear membrane. Bay41-4109 interferes with the assembly of core particles and the dissociated core proteins are able to enter the nuclei. DMSO promotes the nuclear entry of core protein/core particles and facilitates the formation of cccDNA.

[Preparation of a monoclonal antibodies against Plasmodium falciparum glutamate dehydrogenase and establishment of colloidal gold-immunochromatographic assay].

OBJECTIVE: To prepare a monoclonal antibodies (mAbs) against glutamate dehydrogenase (GDH) of Plasmodium falciparum (FCC1/HN strain) and establish colloidal gold-immunochromatographic assay (GICA) for diagnosis of Plasmodium falciparum malaria. METHODS: Recombinant GDH was used to immunize Balb/C mice and the mAbs against GDH were prepared using hybridoma technique followed by identification of IgG isotype and its affinity. Protein-G affinity chromatography was employed to purify the antibodies, which were labeled with colloidal gold for establishment of GICA for Plasmodium falciparum detection. RESULTS: Six mAbs were obtained and identified as IgG1(kappa) of IgG isotypes with affinity constants (Kaff) ranging from 1 x 10(-8) to 2.8 x 10(-10). GICA had a sensitivity of 86.66%; and specificity of 96.43%; for Plasmodium falciparum detection compared with routine microscopic examination. CONCLUSION: The established GICA is rapid and accurate for Plasmodium falciparum detection with such potential utility as for instant diagnosis of Plasmodium falciparum malaria.

[Preparation of monoclonal antibodies against multiple antigens].

OBJECTIVE: To prepare monoclonal antibodies (mAbs) against multiple antigens by single cell fusion. METHODS: BALB/c mice were immunized with the multiple antigens, namely alpha fetoprotein (AFP), carcinoembryonic antigen (CEA), HBsAgiHBcAg and HBeAg, and hybridomas were employed using PEG as the fusing agent. The hybridoma cells were respectively screened with AFP, CEA, HBsAg, HBcAg and HBeAg by enzyme-linked immunosorbent assay and limited dilution. The mAbs were purified by protein G affinity chromatography, its subtype was identified, the affinity constants (K(a)) were determined and the specificity was analyzed by Western blotting. RESULTS: Twenty hybridoma cell lines were obtained by single cell fusion, including 5 cell lines against AFP, 6 against CEA, 3 against HBsAg, 4 against HBcAg, and 2 against HBeAg. The subtypes of some hybridoma cell lines positive for the mAbs were identified as the immunoglobulin G1 (IgG1), with K(a) ranging from 1x10(9) M(-1) to x10(11) M(-1). Western blot analysis showed that all the mAbs strongly and specifically bound to their respective antigens. CONCLUSION: The mAbs against multiple antigens have been obtained by single cell-fusion, which increases the production of mAbs and reduces the time of preparation.