Cross-feeding between cyanobacterium Synechococcus and Escherichia coli in an artificial autotrophic-heterotrophic coculture system revealed by integrated omics analysis.

BACKGROUND: Light-driven consortia, which consist of sucrose-secreting cyanobacteria and heterotrophic species, have attracted considerable attention due to their capability for the sustainable production of valuable chemicals directly from CO2. In a previous study, we achieved a one-step conversion of sucrose secreted from cyanobacteria to fine chemicals by constructing an artificial coculture system consisting of sucrose-secreting Synechococcus elongateus cscB+ and 3-hydroxypropionic acid (3-HP) producing Escherichia coli ABKm. Analyses of the coculture system showed that the cyanobacterial cells grew better than their corresponding axenic cultures. To explore the underlying mechanism and to identify the metabolic nodes with the potential to further improve the coculture system, we conducted integrated transcriptomic, proteomic and metabolomic analyses. RESULTS: We first explored how the relieved oxidative stress affected cyanobacterial cell growth in a coculture system by supplementing additional ascorbic acid to CoBG-11 medium. We found that the cell growth of cyanobacteria was clearly improved with an additional 1 mM ascorbic acid under axenic culture; however, its growth was still slower than that in the coculture system, suggesting that the improved growth of Synechococcus cscB+ may be caused by multiple factors, including reduced oxidative stress. To further explore the cellular responses of cyanobacteria in the system, quantitative transcriptomics, proteomics and metabolomics were applied to Synechococcus cscB+. Analyses of differentially regulated genes/proteins and the abundance change of metabolites in the photosystems revealed that the photosynthesis of the cocultured Synechococcus cscB+ was enhanced. The decreased expression of the CO2 transporter suggested that the heterotrophic partner in the system might supplement additional CO2 to support the cell growth of Synechococcus cscB+. In addition, the differentially regulated genes and proteins involved in the nitrogen and phosphate assimilation pathways suggested that the supply of phosphate and nitrogen in the Co-BG11 medium might be insufficient. CONCLUSION: An artificial coculture system capable of converting CO2 to fine chemicals was established and then analysed by integrated omics analysis, which demonstrated that in the coculture system, the relieved oxidative stress and increased CO2 availability improved the cell growth of cyanobacteria. In addition, the results also showed that the supply of phosphate and nitrogen in the Co-BG11 medium might be insufficient, which paves a new path towards the optimization of the coculture system in the future. Taken together, these results from the multiple omics analyses provide strong evidence that beneficial interactions can be achieved from cross-feeding and competition between phototrophs and prokaryotic heterotrophs and new guidelines for engineering more intelligent artificial consortia in the future.

hsa-MicroRNA-28-5p Inhibits Diffuse Large B-Cell Lymphoma Cell Proliferation by Downregulating 14-3-3ζ Expression.

MicroRNAs (miRNAs) participate in the comprehensive biological process of several cancer types. In our former study, we found that hsa-microRNA- (miR-)28-5p was downregulated, but tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activating protein zeta (14-3-3ζ or YWHAZ) was upregulated in diffuse large B-cell lymphoma (DLBCL) tissues. We predicted that YWHAZ was a target gene for hsa-miR- 28-5p using bioinformatics analysis. Our goal was to reveal the role of hsa-miR-28-5p in DLBCL. YWHAZ was tested by immunohistochemistry (IHC) in formalin-fixed paraffin-embedded (FFPE) tissues of 137 DLBCL tissues, and the expression of hsa-miR-28-5p and YWHAZ was examined by quantitative real-time polymerase chain reaction (qRT-PCR) in 15 fresh and frozen DLBCL tissues. To study the functional roles of the downregulated hsa-miR-28-5p in DLBCL, a Cell Counting Kit-8 assay was conducted to estimate cell proliferation. Transient transfection of miRNA mimics was performed to overexpress hsa-miR-28-5p, and flow cytometry was performed to examine cell apoptosis and cell cycle progression. A dual-luciferase reporter assay was employed to explore the relationship between hsa-miR-28-5p and YWHAZ. Western blotting and qRT-PCR were used to investigate the function of hsa-miR-28-5p in YWHAZ expression. hsa-miR-28-5p was found to be significantly downregulated in DLBCL tissues and cell lines. Functional studies showed that hsa-miR-28-5p overexpression inhibited cell viability and proliferation, and YWHAZ was predicted to be a target of hsa-miR-28-5p. Dual-luciferase reporter assay, Western blotting, and qRT-PCR verified that hsa-miR-28-5p negatively regulated YWHAZ expression by directly targeting its 3' untranslated regions in DLBCL cells. hsa-miR-28-5p may suppress the growth of DLBCL cells by inhibiting YWHAZ expression. These findings could provide novel targets for DLBCL diagnosis and therapy.

[Effect of acupuncture on serum inflammatory cytokines and intestinal flora in rats with stress-induced gastric ulcer].

OBJECTIVE: To investigate the effect of acupuncture on serum inflammatory cytokines and intestinal flora in rats with stress-induced gastric ulcer (SGU), and to explore the mechanism of acupuncture in the treatment of SGU. METHODS: Sprague-Dawley rats were randomly divided into blank, model, acupuncture, and medication groups, with 7 rats in each group. Restraint water-immersion stress was used to establish the model of SGU. The rats in the acupuncture group were given acupuncture at "Zhongwan"(CV12) and bilateral "Zusanli"(ST36) for 20 min, once a day for 5 days, and those in the medication group were given 2 mL solution of Omeprazole enteric-coated tablets (0.2 mg/kg) by gavage, once a day for 5 days. The Guth method was used to calculate the gastric mucosa damage index, HE staining was used to observe the histopathological changes of the gastric mucosa, ELISA was used to measure the serum levels of interleukin-4 (IL-4) and interleukin-6 (IL-6), and 16S rDNA sequencing method was used to observe the change in intestinal flora. RESULTS: Compared with the blank group, the model group had a significant increase in gastric mucosa damage index (P<0.01), markedly pathological changes of the gastric mucosa shown by HE staining, a significant reduction in the content of serum IL-4 (P<0.01), and a significant increase in the content of serum IL-6 (P<0.01), as well as a significant reduction in Bacteroidetes and Firmicutes at the phylum level. Compared with the model group, the acupuncture group and the medication group had a significant reduction in gastric mucosa damage index (P<0.01, P<0.05). HE staining showed reduced pathological changes of the gastric mucosa, as well as a significant increase in the content of serum IL-4 (P<0.01, P<0.05) and a significant reduction in the content of serum IL-6 (P<0.01, P<0.05). As for the intestinal flora, there was a significant increase in Bacteroidetes. Compared with the medication group, the gastric mucosa damage index was decreased （P<0.05），the content of serum IL-4 was significantly increased （P<0.05）, while the content of serum IL-6 significantly decreased (P<0.05) in the acupuncture group. CONCLUSION: Acupuncture at CV12 and ST36 can down-regulate the content of serum IL-6, up-regulate the content of serum IL-4, maintain the relative homeostasis of inflammatory cytokines, and regulate the community structure of intestinal flora, and thus help to repair the damage of gastric mucosa.

[Effect of acupuncture on intestinal flora in rats with stress gastric ulcer].

OBJECTIVE: To observe the effect of acupuncture at "Baihui" (GV 20), "Zhongwan" (CV 12) and "Zusanli" (ST 36) on intestinal flora in rats with stress gastric ulcer (SGU) , and to explore the mechanism of acupuncture promoting SGU recovery. METHODS: Thirty-one SPF SD rats were randomly divided into a control group (7 rats), a model control group (8 rats), an acupuncture group (8 rats) and a medication group (8 rats). The rats in the model group, acupuncture group and medication group were selected to applied the improved restraint water-immersion stress method to establish the SGU model. After modeling, the rats in the control group and model group were fixed and restrained for 20 min every day for a total of 5 days; the rats in the acupuncture group were intervented with acupuncture at "Baihui" (GV 20), "Zhongwan" (CV 12) and "Zusanli" (ST 36), once a day, 20 min each time, and twisting needle for 30 s every 5 min for a total of 5 days; the rats in the medication group were gavaged by solution of omeprazole enteric-coated tablet (200 mg/mL), 2 mL for each rat, once a day. Guth method was used to calculate the gastric mucosal damage index (GMDI), HE staining was used to observe the pathological changes of gastric mucosa, and 16SrDNA identification was used to detect the structural abundance of intestinal flora. RESULTS: Compared with the control group, the GMDI of rats in the model group was increased (P<0.01), the gastric mucosal pathological changes were significant, and the intestinal flora richness index Chao1, Observed species and diversity index Shannon were all decreased (P<0.05), the diversity index Simpson was increased (P<0.05). Compared with the model group, the GMDI of rats in the acupuncture group and medication group was reduced (P<0.01, P<0.05), the gastric mucosal damage degree was reduced, and the intestinal flora richness index Chao1, Observed species and diversity index Shannon were all increased (P<0.05) and the diversity index Simpson decreased (P<0.05). Compared with the medication group, the GMDI of rats in the acupuncture group was reduced (P<0.01), the recovery of gastric mucosal injury was better than that of the medication group. CONCLUSION: Acupuncture can effectively improve gastric mucosal injury of SGU, and the mechanism may be related to increasing the diversity of intestinal flora and promoting the correction of the disordered intestinal flora.

Optimization of extraction of loquat flowers polyphenolics and its antioxidant and anti-polyphenol oxidase properties.

In this study, the conditions of extraction of loquat flowers polyphenolics were optimized through response surface methodology (RSM). Proper extraction conditions were: solid to liquid ratio 1 g per 50 mL and ethanol concentration 50% at 61°C for 9 min. Furthermore, the antioxidant and anti-polyphenol oxidase (PPO) activity of purified total polyphenolics (PTP) were investigated. PTP displayed strong antioxidant activity with IC50 values of 126.3 ± 8.9, 162.4 ± 6.3 and 94.97 mg ascorbic acid equivalent/g dry weight (mg AAE/d.w.) for ABTS, DPPH, and FRAP assays. In addition, PTP has a substantial inhibitory activity on PPO (IC50 = 115 ± 9.2 µg/mL). From the kinetics analysis, it was proved to be a reversible and mixed-type inhibitor of PPO with KI and KIS values of 76.77 µg/mL and 227.86 µg/mL, respectively. Further, the molecular mechanism underlying the inhibition of PPO by PTP was investigated by molecular docking techniques. The results showed that PTP units could form interaction with the catalytic pocket of PPO through the interaction with amino acid residues in the enzyme active center. The antioxidant activities of PTP together with its effect on PPO activity provide a strong starting point for their practical usage in the food industry.

[Moxibustion improved gastric ulcer by reducing contents of corticotrophin-releasing hormone and adrenocorticotropic hormone in serum and hypothalamus-pituitary tissues in rats with stress-induced gastric ulcer].

OBJECTIVE: To observe the effect of moxibustion of "Zhongwan" (CV12) and "Zusanli" (ST36) on histopathological changes of the gastric mucosa and contents of corticotropin-releasing hormone(CRH) and adrenocorticotrophic hormone(ACTH) in the serum, hypothalamus and pituitary tissues in rats with stress-induced gastric ulcer(SGU), so as to reveal its mechanisms underlying improvement of SGU. METHODS: A total of 28 male SD rats were randomly divided into blank control, model, moxibustion, and medication groups, with 7 rats in each group. The SGU model was established by water immersion restraint stress for 3 h. Moxibustion was applied to "Zhongwan"(CV12) and bilateral "Zusanli" (ST36) for 20 min, once a day for 5 days, and rats of the medication group were treated by gavage of Omeprazole enteric-coated tablets (0.2 mg/kg) once a day for 5 days. The gastric mucosal damage index (ulcer index, UI) was measured to assess the injury severity according to Guth's me-thods. Histopathological changes of the gastric mucosa were determined by H．E. staining. The contents of CRH in serum and hypothalamus and ACTH in serum and pituitary gland tissue were assayed by using ELISA. RESULTS: Outcomes of H．E. staining showed gastric mucosal epithelia defect, disordered arrangement of glands, obvious mucosal hyperemia and edema, exudation of a large number of red blood cells, swelling of mucosal cells with necrosis of nuclei in the model group. These situations were relatively milder in the moxibustion and medication groups. After modeling, the UI, and the contents of CRH in the serum and hypotha-lamus, and ACTH in the serum and pituitary tissue were significantly increased in comparison with the blank control group (P<0.01). Following the intervention, the UI, and contents of CRH in the serum and hypothalamus, and ACTH in the serum and pituitary were all down-regulated in both medication and moxibustion groups relevant to the model group (P<0.05, P<0.01). The therapeutic effect of moxibustion was notably superior to that of medication in down-regulating serum CRH and ACTH (P<0.05). No significant differences were found between the medication and moxibustin interventions in lowering the UI, hypothalamic CRH and pituitary ACTH levels (P>0.05)．. CONCLUSION: Moxibustion can relieve gastric mucosal injury induced by stress in water immersion restraint stress rats, which may be associated with its effects in down-regulating the levels of CRH and ACTH in se-rum, hypothalamus and pituitary tissues (inhibition of activities of hypothalamic-pituitary-adrenocortical axis)．.

Effectiveness comparisons of acupuncture for chronic prostatitis/chronic pelvic pain syndrome: A Bayesian network meta-analysis protocol.

BACKGROUND: Chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) is a common urinary system disease in the male population. Recent studies have shown that acupuncture can alleviate the pain caused by CP/CPPS to a certain extent and improve the quality of life of patients. This study used a network meta-analysis (NMA) to compare the effectiveness and safety of different forms of acupuncture on CP/CPPS. METHODS AND ANALYSIS: We will search for PubMed, Cochrane Library, AMED, EMbase, WorldSciNet; Nature, Science online and China Journal Full-text Database, China Biomedical Literature CD-ROM Database, and related randomized controlled trials (RCTs) included in the China Resources Database. The time is limited from the construction of the library to December 2018. The quality of the included RCTs will be evaluated with the risk of bias tool and evidence will be evaluated by grading of recommendations assessment, development, and evaluation. STATA 13.0 and WinBUGS 1.4.3 through the GeMTC package will be used to perform a NMA to synthesize direct and indirect evidence. RESULTS: The results of this NMA will be submitted to a peer-reviewed journal for publication. TRIAL REGISTRATION NUMBER: PROSPERO CRD42018111408.

[Effect of Electroacupuncture on Cerebro-cortex Caspase-3 Expression and Blood Lipid Levels in Hyperlipemia Rats with Cerebral Ischemia].

OBJECTIVE: To observe the effect of electroacupuncture (EA) stimulation of "Fenglong" (ST 40), "Sanyinjiao" (SP 6) plus manual acupuncture (MA) stimulation of "Shuigou" (GV 26) and "Baihui" (GV 20) on Caspase-3 protein expression in the cerebral cortex of rats with hyperlipemia and cerebral ischemia(HL-CI),so as to reveal its mechanisms underlying improvement of HL-CI. METHODS: Forty-five rats were randomly divided into normal control,sham operation,model,EA group I(EA+MA was given for 14 days, i.e., 7 days before CI, and 7 days more after HL-CI)and EA group Ⅱ (EA+MA was given for only 7 days after HL-CI),with 9 rats being in each group. The HL-CI model was established by feeding the animals with high fat forage for 6 weeks and then making an occlusion of the unilateral middle cerebral artery by regional application of quantitative paper adsorbing 50% FeCl3 solution (10 µL). Rats of the sham operation group were treated with the same procedures only without application of FeCl3 solution. For rats of the EA group I,EA (1-3 mA, 2 Hz/100 Hz) was applied to bilateral acupoints SP 6 and ST 40 (for 20 min),and MA stimulation applied to GV 26 and GV 20. EA was conducted once daily for 7 days after 6 weeks' high fat fo-rage feeding, and EA+MA intervention was conducted once daily for 7 days after CI modeling. For rats in the EA group Ⅱ, EA+MA was applied to the same 4 acupoints once a day for 7 days only after CI modeling. The neurological impairment was assessed by Zea Longa's scoring. The blood sample was taken from the abdominal aorta for measuring the contents of serum cholesterol (CHO),triglyceride (TG) and high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C). Pathological changes of the cerebral cortex were observed after H.E. staining, and the expression of cerebro-cortex Caspase-3 was analyzed by immunohistochemistry. RESULTS: Following modeling,the neurological score,CHO, TG and LDL-C contents, and the number of Caspase-3 positive cells as well as Caspase-3 immunoactivity level were significantly increased in the model group(P<0.05), while serum HDL-C level was obviously decreased(P<0.05). After the treatment,the increased neurological score, CHO, TG and LDL-C contents, and the number of Caspase-3 positive cells and Caspase-3 immunoactivity level were considerably decreased in the EA group I and Ⅱ(P<0.05)while the decreased HDL-C level was notably increased relevant to the model group(P<0.05). The effects of the EA group I were notably superior to those of EA group Ⅱ in down-regulating the neurological score,CHO, TG and LDL-C levels and the expression of Caspase-3 protein(P<0.05). No significant differences were found between the normal control and sham operation groups in the neurological scores 20 min and 7 days after modeling and Caspase-3 expression levels (P>0.05). H.E. staining showed a reduction of the apoptotic cells and inflammatory cells in both EA group I and Ⅱ. CONCLUSIONS: Both EA and EA+MA interventions can improve neurological function in HL-CI rats,which may be related to their effects in adjusting the levels of serum lipids and down-regulating the expression of cell apoptosis-related Caspase-3 protein in the ischemic cortex. Moreover, the cerebral ischemia injury may be lightened by EA-lowering hyperlipemia first.

[Systematic Evaluation and Meta-analysis on Acupuncture for Peptic Ulcer].

OBJECTIVE: To systematically evaluate the clinical efficacy of acupuncture for peptic ulcer. METHODS: Randomized controlled trials of acupuncture for peptic ulcer were searched from China National Knowledge Infrastructure (CNKI), WanFang Database, Chinese Scientific and Technological Journals (VIP), China Biomedicine (CBM), PubMed and the Cochrane Library from the establishment time of databases to September, 2016. Data extraction and quality evaluation were implemented for the literature which met the inclusive criteria. The RevMan 5.3 software was used to make Meta-analysis. RESULTS: Sixteen papers including 1 570 patients of peptic ulcer were included. The results of Meta-analysis showed that there was no statistical significance between acupuncture and western medicine in the effective rate, the healing rate of ulcer area and the HP negative rate (all P>0.05); the recurrence rate of acupuncture was significantly lower than that of western medicine[RR=0.35, 95%CI (0.14, 0.84), P<0.05]. Acupuncture plus western medicine was significantly different from simple western medicine in the effective rate, the healing rate of ulcer area and the recurrence rate[RR=1.20,95% CI (1.04, 1.38), P=0.01; RR=1.29, 95% CI (1.06, 1.58), P=0.01;RR=0.27, 95% CI (0.16, 0.45), P<0.00001]. The analysis of evidence grade (GRADE) pre-sented that the healing rate of ulcer area and the HP negative rate of acupuncture were "low grade", and others were "extremely low grade". CONCLUSIONS: Acupuncture combined with western medicine has some advantages for peptic ulcer compared with the conventional western medicine, which needs further confirmation due to the lower evidence grade. Larger samples, randomized controlled trials with high quality are highly recommended.

[Effects of Direct Moxibustion of "Ganshu"(BL 18) on the Contents of T cells in Peripheral Blood in Rats with Precancerous Lesion of Primary Hepatocellular Carcinoma].

OBJECTIVE: To observe the effect of moxibustion stimulation of "Ganshu"(BL 18) region on contents of T cells in the peripheral blood in rats with Diethylnitrosamine (DEN)-induced hepatocellular carcinoma (HCC), so as to explore its effective in improving immunoregulatory function. METHODS: Seventy male Wistar rats were randomly divided into control group (n=10), model group(n=15), direct moxibustion-15 s group(n=15), direct moxibustion-30 s group (n=15) and ginger-separated moxibustion group(n=15). The primary HCC precancerous lesion model was established by intraperitoneal injection of DEN (50 mg/kg), once every 3 days for 10 weeks. Moxibustion was applied to bilateral "Ganshu"(BL 18) for about 15 min (3 moxa-cones), or 30 min (six moxa-cones), with or without ginger-slice separation, once every other day for 10 consecutive weeks. The contents of T cells of CD 3+, CD 4+, CD 8+ and CD 4+/CD 8+ ratio in the peripheral blood were detected with Flow Cytometey(FCM), pathological changes of liver were observed by light microscope after hematoxylin-eosin(HE) stain. RESULTS: Compared to the control group, the contents of blood CD 3+ and CD 4+ T cells and ratio of CD 4+/CD 8+ were significantly down-regulated, while that of CD 8+T cells was obviously increased in the model group(P<0.05,P<0.01). After moxibustion intervention, the decreased CD 3+ and CD 4+T cells and CD 4+/CD 8+ levels and the increased CD 8+T cell contents were reversed in all the 3 moxibustion groups (P<0.05, P<0.01), except CD 3+ in the ginger-separated moxibustion group (P>0.05). There were no significant differences among the three moxibustion groups in the CD 3+, CD 4+, CD 8+ and CD 4+/CD 8+ levels (P>0.05). In addition, the pathological changes of liver tissue as central vein deviation or absence, disordered arrangement of hepatic cords, narrowing of the hepatic sinusoid, hyperplasia of collagen fibers, formation of tuberosis, unevenness of liver cells with nuclear anachromasis and higher heteromorphism, and macronucleus oncocytes in primary HCC rats were not observed or milder after moxibustion intervention. CONCLUSIONS: Direct moxibustion and ginger-separated moxibustion can improve pathological changes of hepatic cells in rats with HCC, which may be associated with its actions in raising blood CD 3+ and CD 4+ T cell contents and reducing CD 8+ T levels to improve immune function.