Acupuncture for patients with chronic functional constipation: A randomized controlled trial.

BACKGROUND: Acupuncture is used to treat chronic functional constipation (CFC) in China, despite limited evidence. We aim to assess the effectiveness and safety of acupuncture in managing CFC. METHODS: A multicenter randomized controlled trial was performed involving 684 patients with CFC; the patients were randomly allocated to receive He acupuncture (n = 172), Shu-mu acupuncture (n = 171), He-shu-mu acupuncture (n = 171), or oral administration of mosapride (n = 170). Sixteen sessions of acupuncture were given in the treatment duration of 4 weeks. The primary outcome was the change in spontaneous bowel movements (SBMs) at week 4 (at the end of treatment) compared to baseline. The secondary outcomes included stool consistency (Bristol scale), the degree of straining during defecation, and adverse events. KEY RESULTS: The SBMs increased in all the four groups at week 4, and the magnitude of increase was equivalent in the four groups (He acupuncture, 2.7 [95% CI, 2.3-3.1]; Shu-mu acupuncture, 2.7 [95% CI, 2.3-3.0]; He-shu-mu acupuncture, 2.2 [95% CI, 1.9-2.5]; and mosapride, 2.4 [95% CI, 2.0-2.9]; P = .226). However, the change in SBMs at week 8 was significantly smaller in mosapride group (1.4 [95% CI, 1.0-1.8]) than the three acupuncture groups (2.4 [95% CI, 2.1-2.7], 2.3 [95% CI, 1.9-2.7], 2.1 [95% CI, 1.7-2.5] in He, Shu-mu, and He-shu-mu group, respectively, P = .005). CONCLUSIONS & INTERFERENCES: The three acupuncture treatments were as effective as mosapride in improving stool frequency and stool consistency in CFC, but the magnitude of the treatment effect is unknown due to the lack of sham acupuncture control.

Cloning and characterization of the UBC gene from lotus (Nelumbo nucifera).

Protein ubiquitination is extensively involved in the regulation of a considerable number of physiological processes in plant cells. E2 (ubiquitin-conjugating enzyme, UBC), one of the essential enzymes of eukaryotic ubiquitination, catalyzes protein ubiquitination together with E1 and E3. In this study, we cloned four full-length cDNA NnUBCs of Nelumbo nucifera. With the same coding sequence length of 459 bp and coding 153 amino acids, these four genes are highly homologous with the AtUBC1 and AtUBC2 of Arabidopsis thaliana. Quantitative fluorescence polymerase chain reaction showed that these four genes exhibited different expression patterns in different tissues of N. nucifera. Overall, the expression of NnUBC3 was the highest in all plant tissues. Tests of different stress treatments showed that NnUBC3 plays an important role in response to heat, salt, and drought stresses in N. nucifera. Moreover, transgenic Arabidopsis plants (Atubc1-1Atubc2-1 mutant) expressing NnUBC3 presented a wild-type phenotype, indicating that NnUBC3 performs the same function as AtUBC1 and AtUBC2.

Molecular and characterization of NnPPO cDNA from lotus (Nelumbo nucifera) in rhizome browning.

The complete cDNA (NnPPO) of polyphenol oxidase in Nelumbo nucifera was successfully isolated, using Rapid amplification cDNA end (RACE) assays. The full-length cDNA of NnPPO was 2069 bp in size, containing a 1791 bp open reading frame coding 597 amino acids. The putative NnPPO possessed the conserved active sites and domains for PPO function. Phylogenetic analysis revealed that NnPPO shared high homology with PPO of high plants, and the homology modeling proved that NnPPO had the typical structure of PPO family. In order to characterize the role of NnPPO, Real-time PCR assay demonstrated that NnPPO mRNA was expressed in different tissues of N. nucifera including young leave, rhizome, flower, root and leafstalk, with the highest expression in rhizome. Patterns of NnPPO expression in rhizome illustrated its mRNA level was significantly elevated, which was consistent with the change of NnPPO activity during rhizome browning. Therefore, transcriptional activation of NnPPO was probably the main reason causing rhizome browning.

Development and characterization of genic-SSR markers from different Asia lotus (Nelumbo nucifera) types by RNA-seq.

Nelumbo nucifera is an important economic vegetable and traditional medicine, but available genetic resources remain limited. Next generation sequencing has proven to be a rapid and effective means of identifying genic simple sequence repeat (genic-SSR) markers. This study developed genic-SSRs for N. nucifera using Illumina sequencing technology to assess diversity across cultivated and wild lotus. A total of 105,834 uni-contigs were produced with an average read length of 722 bp. Exactly 11,178 genic-SSR loci were identified in 9523 uni-contigs. Di-nucleotide (64.5%) was the most abundant SSR, followed by tri-nucleotide (23%), tetra-nucleotide (8.9%), penta-nucleotide (2.5%), and hexa-nucleotide (1%) repeat types. The most common di- and tri-nucleotide repeat motifs were AG/CT (51%) and AAG/CTT (8%), respectively. Based on these SSRs sequences, 6568 primer pairs were designed, of which 72 primers were randomly selected for synthesis and validation, and 38 in-silico polymorphic primers were obtained using in-house perl scripts. A total of 110 primers were screened in the lotus samples and the results showed that 101 primers yielded amplification products, of which 80 were polymorphs. The number of alleles ranged from 2 to 17 and the PIC (polymorphism information content) ranged from 0.19 to 0.87 with a mean value of 0.55. An Unweighted Pair Group Method with Arithmetic Mean (UPGMA) dendrogram based on Jaccard's similarity coefficients showed that the correlation between geographical source and genotype was low. This study describes the distribution of genic-SSRs in the expressed portion of the lotus genome. These genic-SSRs have an important role to play in molecular mapping, diversity analysis, and marker-assisted selection strategies in Nelumbo.

Identification and characterization of chalcone synthase cDNAs (NnCHS) from Nelumbo nucifera.

Chalcone synthase (CHS) catalyzes the first committed step in flavonoids biosynthetic pathway. In this study, six full-length cDNAs (NnCHS) encoding CHS from Nelumbo nucifera were successfully isolated, using rapid amplification cDNA end (RACE) assay. The obtained cDNAs were 1426 bp in size, containing a 1167 bp open reading frame coding 389 amino acids. Exons-intron architecture of NnCHS gene was illustrated, consisting two exons inserted by a 426 bp intron. The putative NnCHS possessed all the conserved active sites for CHS function as well as the family signature. Phylogenetic analysis revealed that NnCHS shared high homology with CHS from high plants, and the homology-based structural modeling showed that NnCHS had the typical structure of CHS. Moreover, Real-time PCR assays demonstrated that NnCHS mRNAs were expressed in various tissues of N. nucifera, with the highest expression in red flower and lowest level in the leaves. Moreover, patterns of NnCHS expression illustrated short-time wounding or low temperature significantly induced the up-regulation of NnCHS mRNA.

CBF-dependent signaling pathway: a key responder to low temperature stress in plants.

Plants under low temperature (LT) stress exhibit a C-repeat binding factor (CBF)-dependent responsive pathway. The transcription factors in the CBF family, existing in multiple plant species, are the key regulators of the cold-responsive (COR) genes. CBF1 and CBF3 are regulated in a different way from CBF2, and CBF4 is the only known CBF gene definitely involved in abscisic acid (ABA)-dependent signaling pathways. RAP2.1 and RAP2.6 are the downstream regulators under CBFs. The upstream regulators of the CBF named inducer of CBF expression (ICE) acts as a positive regulator of CBFs. Meanwhile, these CBF signaling pathway components could associate with many other transcription activators and repressors in regulating gene expression when plants are under LT stress. HOS1 negatively regulates ICE1, which down regulates MYB15, an upstream repressor of CBFs. ZAT12 participates in the repression of CBFs, while ZAT10 and FRY2 negatively regulate the CBF-target genes. ADF5 was recently also found to repress CBFs. LOS2 works against ZAT10, and LOS4 positively regulates CBFs. SFR6 is involved in the modification of CBFs to activate the COR genes, and SIZ1-dependent sumoylation plays a positive role in the regulation of ICE1. The utilization of CBF-dependent signaling components has a broad perspective in the field of plant breeding for enhancing crop LT tolerance.

Analysis of the interindividual conservation of T cell receptor alpha- and beta-chain variable regions gene in the peripheral blood of patients with systemic lupus erythematosus.

The aim of this study was to find conserved motifs in specific T cell receptor (TCR) alpha- and beta-chains, and to analyse the association between complementarity determining region 3 (CDR3) spectratype and systemic lupus erythematosus (SLE) activity. TCR alpha-and beta-chain CDR3 spectratypes were analysed in 20 SLE patients. The CDR3 spectratypes of three patients were monitored over time, and the CDR3 regions of clonally expanded T cells were sequenced. CDR3 spectratype analysis showed prominent usage of TCR AV8, AV14, AV23, AV30, AV31, BV2, BV8, BV11, BV14, BV16, BV19 and BV24 families in SLE patients. The CDR3 spectratype showed dynamic change correlating with SLE activity. The sequence of the CDR3 region in clonally expanded T cells suggested a conserved GGX amino acid motif in both alpha- and beta-chains. The Ja34 and Jb2s1 region genes were found in high frequency. Both TCR Valpha and Vbeta gene usage is highly restricted in SLE, suggesting that the TCRs recognize a limited number of antigenic epitopes. The conserved motifs and limited use of joining region genes may indicate the recognition of similar antigenic epitopes in multiple individuals.

[An in vitro and in vivo study of antitumor effects of rHTNF-alpha on human ovarian cancer].

We examined the cytotoxic activities of recombinant human tumor necrosis factor (rHTNF-alpha) and five chemotherapeutic agents, CTX, 5-Fu, VCR, DDP, KSM, against two human ovarian cancer cell lines, OVCAR3 and CAOV3, using the MTT assay. The results showed that cytotoxicities of rHTNF-alpha at 5 x 10-5 x 10(4) u/ml against OVCAR3 cell line for 24 h exposure were from 14.2 +/- 6.8% to 67.2 +/- 3.0%, and those against CAOV3 cell line were from 8.2 +/- 4.3% to 60.9 +/- 1.3%. The cytotoxic effects of all five chemotherapeutic agents against the two cell lines were much lower than that of rHTNF-alpha. Further, we studied the combined anticancer potential of rHTNF-alpha with chemotherapeutic agents against the two cell lines. Various degrees of synergism in cytotoxicities of DDP or KSM in combination with rHTNF-alpha were observed. The cytotoxic effect of rHTNF-alpha on CAOV3 cell were also morphologically observed under phase contrast and electron microscope. Based on experiment in vitro, the in vivo anticancer activity of rHTNF-alpha alone or in combination with KSM was examined against human ovarian cancer OVCAR3 subcutaneously transplanted in nude mice. After 8 weeks of treatment, a statistically significant difference of mean tumor volume was found between the control group and groups that received rHTNF-alpha or rHTNF-alpha plus KSM (P < 0.01).

Recombinant proteins VP1 and VP3 of hepatitis A virus prime for neutralizing response.

Six overlapping genomic regions of capsid proteins VP1 and VP3 of hepatitis A virus (HAV) inserted into the expression vectors pBD or pUR respectively expressed beta-galactosidase-HAV fusion proteins. The recombinant proteins were poorly soluble so they were difficult to detect by human anti-HAV sera in radioimmunoassay, but the fusion proteins dissolved in sodium dodecyl sulfate reacted with human and rabbit anti-HAV-positive sera in immunoblots. Antisera against VP1 and VP3 recombinant proteins reacted with the respective structural proteins of HAV in immunoblots. Two recombinant proteins, one including the first 120 amino acids of the N-terminus of VP1 and the other containing all of VP1 except for the first 60 N-terminal amino acids, induced a transient neutralizing antibody response in rabbits. Antisera directed against other regions of VP1 and VP3 neither neutralized viral infectivity nor recognized native virus in a competitive radioimmunoassay. However, when immunized animals were challenged with a sub-immunogenic dose of HAV, all animals responded with stable virus-neutralizing antibodies.