Genetically predicted gut microbiome and risk of oral cancer.

PURPOSE: Mounting evidence suggests a possible link between gut microbiome and oral cancer, pointing to some potential modifiable targets for disease prevention. In the present study, Mendelian randomization (MR) was used to explore whether there was a causal link between gut microbiome and oral cancer. METHODS: The single nucleotide polymorphisms (SNPs) significantly associated with gut microbiome were served as instrumental variables. MR analyses were performed using genetic approaches such as inverse variance weighting (IVW), MR Egger and weighted median, with IVW as the primary approach, supplemented by MR Egger and weighted median. Mendelian randomization pleiotropy residual sum and outlier (MR-PRESSO) and MR-Egger regression were used to detect the presence of horizontal pleiotropy and identify outlier SNPs. RESULTS: Causal effect estimates indicated that genetically predicted abundance of Prevotellaceae was associated with higher risk of oral cancer (odds ratio (OR) 1.80, 95% confidence interval (CI) 1.16-2.81, p = 0.009). There was no evidence of notable heterogeneity and horizontal pleiotropy. CONCLUSION: Genetically derived estimates suggest that Prevotellaceae may be associated with the risk of oral cancer. Such robust evidence should be given priority in future studies and explore the underlying mechanisms.

Volume Incentive Spirometry Reduces Pulmonary Complications in Patients After Open Abdominal Surgery: A Randomized Clinical Trial.

Objective: To compare the effect of diaphragmatic breathing and volume incentive spirometry (VIS) on hemodynamics, pulmonary function, and blood gas in patients following open abdominal surgery under general anesthesia. Methods: A total of 58 patients who received open abdominal surgery were randomly assigned to the control group (n=29) undergoing diaphragmatic breathing exercises and the VIS group (n=29) undergoing VIS exercises. All the participants performed the six-minute walk test (6MWT) preoperatively to evaluate their functional capacity. Hemodynamic indexes, pulmonary function tests, and blood gas indexes were recorded before surgery and on the 1st, 3rd, and 5th postoperative day. Results: The functional capacity was not significantly different between the two groups during the preoperative period (P >0.05). At 3 days and 5 days postoperatively, patients in the VIS group had a significantly higher SpO2 than that in the control group (P <0.05). Pulmonary function test values were reduced in both two groups postoperatively when compared to the preoperative values but improved for three and five days afterward (P <0.05). Of note, the significantly elevated levels of peak expiratory flow (PEF), forced expiratory volume in the first second (FEV1), forced vital capacity (FVC), and FEV1/FVC ratio were observed on the 1st, 3rd, and 5th postoperative days in the VIS group compared with those in the control group (P <0.05). Besides, bass excess (BE), and pH values were significantly higher in the VIS group on the 1st postoperative day than those in the control group (P <0.05). Conclusion: Diaphragmatic breathing and VIS could improve postoperative pulmonary function, but VIS exercise might be a better option for improving hemodynamics, pulmonary function, and blood gas for patients after open abdominal surgery, hence lowering the incidence of postoperative pulmonary complications.

Therapeutic effects of total saikosaponins from Radix bupleuri against Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disease characterized by memory loss and cognitive dysfunction in the elderly, with amyloid-beta (Aß) deposition and hyperphosphorylation of tau protein as the main pathological feature. Nuclear factor 2 (Nrf2) is a transcription factor that primarily exists in the cytosol of hippocampal neurons, and it is considered as an important regulator of autophagy, oxidative stress, and inflammation. Total saikosaponins (TS) is the main bioactive component of Radix bupleuri (Chaihu). In this study, it was found that TS could ameliorate cognitive dysfunction in APP/PS1 transgenic mice and reduce Aß generation and senile plaque deposition via activating Nrf2 and downregulating the expression of ß-secretase 1 (BACE1). In addition, TS can enhance autophagy by promoting the expression of Beclin-1 and LC3-II, increasing the degradation of p62 and NDP52 and the clearance of phosphorylated tau (p-tau), and reducing the expression of p-tau. It can also downregulate the expression of nuclear factor-κB (NF-κB) to inhibit the activation of glial cells and reduce the release of inflammatory factors. In vitro experiments using PC12 cells induced by Aß, TS could significantly inhibit the aggregation of Aß and reduce cytotoxicity. It was found that Nrf2 knock-out weakened the inhibitory effect of TS on BACE1 and NF-κB transcription in PC12 cells. Moreover, the inhibitory effect of TS on BACE1 transcription was achieved by promoting the binding of Nrf2 and the promoter of BACE1 ARE1. Results showed that TS downregulated the expression of BACE1 and NF-κB through Nrf2, thereby reducing the generation of Aß and inhibiting neuroinflammation. Furthermore, TS can ameliorate synaptic loss and alleviate oxidative stress. In gut microbiota analysis, dysbiosis was demonstrated in APP/PS1 transgenic mice, indicating a potential link between gut microbiota and AD. Furthermore, TS treatment reverses the gut microbiota disorder in APP/PS1 mice, suggesting a therapeutic strategy by remodeling the gut microbe. Collectively, these data shows that TS may serve as a potential approach for AD treatment. Further investigation is needed to clarify the detailed mechanisms underlying TS regulating gut microbiota and oxidative stress.

Design, synthesis, and biofunctional evaluation of novel pentacyclic triterpenes bearing O-[4-(1-piperazinyl)-4-oxo-butyryl moiety as antiproliferative agents.

A series of pentacyclic triterpenoids derivatives bearing O-[4-(1-piperazinyl)-4-oxo-butyryl moiety has been synthesized and investigated for their potential antiproliferative activities. Pentacyclic triterpenoids derivative compounds were synthesized by a four or six step synthetic procedure. The activity studies were evaluated using Cell Counting Kit-8 method, and Western blotting analysis on A549 cells, MCF-7 cells and Hela cells. Compounds methyl 3-O-[4-(1-piperazinyl)-4-oxo-butyryl]olean-12-ene-28-oate (OA-4) and compound 2-O-[4-(1-piperazinyl)-4-oxo-butyryl]-3,23-dihydroxyurs-12-ene-28-oate (AA-5) were found to be promising antiproliferative agents. These compounds show potentiality for further optimization as antitumor drugs.

[The identification and verification of species-specific microRNAs and their precursors in Lampetra japonica].

MicroRNAs (miRNAs) negatively regulate genes which are involved in various biological processes of metabolism at both transcriptional and post-transcriptional levels. In recent years, the existence and function of miRNAs have been extensively studied in plants and animals with the application of deep sequencing and microarray technology. In this study, small RNAs from leucocytes of Lampetra japonica (L. japonica) were sequenced using the second generation high-throughput sequencing technology. A total of 5 207 787 small RNA sequences were identified, and 4 739 346 of them assembled into 10 989 variants. Based on sequence similarity analysis, the sequences of these variants matched known miRNAs of 306 conserved families, among which 6 conserved miRNA family members expressed at an extremely high level which reflected the conservatism of miRNAs among species. In addition, 70 unannotated sequences were predicted to be new miRNAs, and 34 of them were further verified expressing in antigen-treated L. japonica leucocytes by miRNA microarray assay. Moreover, the minimal folding free energy indexes for 16 of the 34 miRNA precursors exceed 0.85, indicating the existence of species-specific miRNAs in L. japonica which may play important roles in regulating, growth, development and disease response of L. japonica leukocytes.

Inhibition of human enterovirus 71 replication by pentacyclic triterpenes and their novel synthetic derivatives.

A large number of bioactive pentacyclic triterpenoids have been shown to have multiple biological activities. This study was conducted to evaluate the inhibitory activities of 6 newly synthesized and novel pentacyclic triterpenoids against enterovirus 71 (EV71). The parent compound, ursolic acid (UA), showed the greatest inhibitory activity against EV71, while oleanolic acid (OA), asiatic acid (AA), and synthetic derivatives of 18-ß-glycyrrhetinic acid (GA) and OA also exhibited inhibitory effects, although to lesser extents. The results suggest these compounds show potential for further optimization as antiviral candidates for treatment of EV71 infections.

[Recent advance in pharmacological therapies for proliferative vitreoretinopathy].

Proliferative vitreoretinopathy (PVR) is a leading blinding disease, which is often associated with ocular trauma, rhegmatogenous retinal detachment and diabetic retinopathy. PVR involves the vitreous and retina and its occurrence is characterized by vitreoretinal cells migration, transformation and excessive proliferation which lead to the formation of pre- or sub-retinal membrane or membrane formation in the vitreous. The subsequent contraction of the membrane can lead to retinal detachment and loss of vision. At present, vitrectomy is the standard treatment modality for the treatment of PVR. However, this procedure is expensive and post-operative vision is often unsatisfactory. With the advances of biological studies, the pathogenesis of PRV becomes clear, and the corresponding pharmacological intervention studies targeting the relevant pathways developed rapidly. This review is aiming to highlight the new developments in pharmacological prevention and treatment for PVR.

[Methyl 3, 4-dihydroxyphenylacetate prevents enterovirus 71 proliferation in rhabdomyosarcoma cells].

This article presented the inhibitory activity of methyl 3, 4-dihydroxyphenylacetate on the enterovirus 71 (EV71) infection. The EV71 VP1 capsid protein expression levels were analyzed with Western blotting. Results revealed that the compound is able to inhibit EV71 replication in rhabdomyosarcoma (RD) cells. After being incubated with the compound at a concentration of 0.01 microg x microL(-1) for 48 h, the level of EV71 vp1 mRNA in RD cells decreased by (76.83 +/- 2.47)%. The cytotoxic activity of the compound was evaluated against RD cells by a MTT assay. The results showed that the compound had low toxicity with a CC50 of 0.072 6 microg x microL(-1). These findings suggest that methyl 3, 4-dihydroxyphenylacetate is a novel compound for antiviral therapies against EV71, which merited further investigation.

[Effect of the compound of poly lactic-co-glycolic acid and bone marrow stromal cells modified by osteoprotegerin gene on the periodontal regeneration in Beagle dog periodontal defects].

OBJECTIVE: To evaluate the effect of the osteoprotegerin (OPG) gene-modified autologous bone marrow stromal cells (BMSCs) on regeneration of periodontal defects, and to provide new experimental evidence to explore the gene therapy for periodontal disease. METHODS: pSecTag2/B-opg was transduced into BMSCs by lipofectamine 2000. The expression of OPG protein in the BMSCs was detected by immunocytochemistry and Western blot. Inverted phase contrast microscope and scanning electron microscopy (SEM) were used to observe the morphology and proliferation of the BMSCs(OPG) on on the surface of the poly lactic-co-glycolic (PLGA). Horizontal alveolar bone defect (4 mmx4 mmx 3 mm) were surgically created in the buccal aspect of the mandibular premolar, and were randomly assigned to receive BMSCs(OPG)-PLGA (cells/material/OPG), BMSCs-PLGA (cells/material), PLGA (material), or root planning only (blank control). The animals were euthanized at 6 weeks post surgery for histological analysis. The height of new alveolar bone and cementum and the formation of new connective tissue were analyzed and compared. All data were statistically analyzed using the q test. RESULTS: The BMSCs transfected by human OPG gene can highly express OPG protein. SEM observations demonstrated that BMSCs(OPG) were able to proliferate and massively colonize on the scaffolds structure. After 6 weeks, the height of new alveolar bone and cementum and the formation of new connective tissue were significantly greater in the experimental group than in the control groups (P < 0.05). CONCLUSION: BMSCs(OPG)-PLGA can significantly promote the regeneration of dog's periodontal bone defects. Gene therapy utilizing OPG may offer the potential for periodontal tissue engineering applications.

[Role of caveolin-1 down-regulation by iRNA in human hepatocyte proliferation].

OBJECTIVE: To investigate the role of caveolin-1 down-regulation in human hepatocyte proliferation in vitro. METHODS: The expression vector psiRNA-CAV1 was constructed and transfected into Chang liver cells (CHL). The caveolin-1 down-regulated cell clones were selected by the antibiotic zeocin. The proliferation of the cell strain CAV7 was examined by MTT, in which untransfected CHL and HepG2 cells were set as controls. Expression of caveolin-1, Akt, Erk1/2, p-Akt and p-Erk1/2 in the transfected and control cells was detected by Western blot. RESULTS: After caveolin-1 expression was down-regulated by RNAi, CHL increased faster at first (24 h and 72 h, P<0.05; 96 h, P<0.01), but slower later. P-Akt and p-Erk1/2 expressions were down-regulated, indicating that the growth and proliferation related Akt and Erk1/2 pathways were inhibited after caveolin-1 down-regulation. CONCLUSION: Caveolin-1 may play an important role in hepatocyte proliferation.

Using the concept of Chou's pseudo amino acid composition to predict protein subcellular localization: an approach by incorporating evolutionary information and von Neumann entropies.

The rapidly increasing number of sequence entering into the genome databank has called for the need for developing automated methods to analyze them. Information on the subcellular localization of new found protein sequences is important for helping to reveal their functions in time and conducting the study of system biology at the cellular level. Based on the concept of Chou's pseudo-amino acid composition, a series of useful information and techniques, such as residue conservation scores, von Neumann entropies, multi-scale energy, and weighted auto-correlation function were utilized to generate the pseudo-amino acid components for representing the protein samples. Based on such an infrastructure, a hybridization predictor was developed for identifying uncharacterized proteins among the following 12 subcellular localizations: chloroplast, cytoplasm, cytoskeleton, endoplasmic reticulum, extracell, Golgi apparatus, lysosome, mitochondria, nucleus, peroxisome, plasma membrane, and vacuole. Compared with the results reported by the previous investigators, higher success rates were obtained, suggesting that the current approach is quite promising, and may become a useful high-throughput tool in the relevant areas.

[Establishment and identification of transiently expression system of bone marrow stromal cells modified by osteoprotegerin gene].

OBJECTIVE: In oder to treat periodontitis by using tissue engineering and gene engineering technology, the article established an transient expression system of bone marrow stromal cells (BMSC) modified by osteoprotegerin (OPG) gene and detected its expression using eukaryotic secreted expression pSecTag2/B-OPG plasmid. METHODS: By solation and culture of BMSC in vitro, the identified recombined plasmid was transiently transfected into BMSC by Lipofectamine 2000 and OPG expression in BMSC was determined by RT-PCR and Western blot in 6 weeks. RESULTS: The fragments of the recombinant plasmid digested with Hind III, EcoR I and BamH I and examined by 10 g/L agarose electrophoresis, were consistent with predicted size. The sequence of OPG was identical to the sequence provided by GeneBank [gi:33878056]. OPG transcribing in BMSC was confirmed by RT-PCR and OPG sustainable expressing in BMSC was detected by Western blot in 39 days. CONCLUSION: The transiently expression system of BMSC modified by OPG gene was successfully established.

TPR domain of Ser/Thr phosphatase of Aspergillus oryzae shows no auto-inhibitory effect on the dephosphorylation activity.

A Ser/Thr phosphatase gene cloned from Aspergillus oryzae, aoppt, revealed that the tetratricopeptide repeat (TPR) and catalytic domains of the full-length AoPPT are located at the N- and C-terminal regions, respectively, similar to those of human Ser/Thr phosphatase 5 (PP5) and yeast Ppt1. Four different regions of AoPPT, namely, a full-length polypeptide, the catalytic domain, the catalytic domain plus C-terminal 15 amino-acid residues and the TPR domain were expressed in Escherichia coli and their roles in dephosphorylation activity were examined, using p-nitrophenyl phosphate as the substrate. The full-length AoPPT showed the highest dephosphorylation activity while the catalytic domain had the lowest activity. The activity of the catalytic domain was not inhibited by the presence of the TPR domain and arachidonic acid did not increase the activity of the full-length enzyme. These findings suggest that the integrity of the entire enzyme would be necessary for its full activity to be expressed.

[A follow up study of total IgM, IgG, nucleoprotein and spike protein antibodies against severe acute respiratory syndrome (SARS) coronavirus in patients with SARS].

OBJECTIVE: To survey the dynamic changing and persistence of the special antibodies, including total IgM, IgG, nucleocapsid protein and spike protein antibodies, against severe acute respiratory syndrome coronavirus (SARS-CoV) in patients with SARS. METHODS: 146 cases, all clinically diagnosed as SARS with positive SARS-CoV IgG, were followed up. 362 serum samples were collected from the onset of the disease to 660 days afterward. Total IgM and IgG against SARS-CoV were tested with commercial ELISA kits. For recombinant nucleoprotein and spike protein, we developed an ELISA to test these two antibodies. RESULTS: Within 20 days of the onset, the positive rate of anti-SARS-CoV IgM was 46.5% (20/43); it reached a peak after 21 - 40 days (80.6%, 25/31). Then, the positive rate of IgM went down gradually to 8.2% (6/73) until 550 days after the onset. The patient's IgG positive rate was lower (34.9%, 15/43) than that of IgM within 20 days of the onset. Then it went up rapidly to 100%. It remained positive (98.6%, 70/71) until 600 - 660 days after the onset. When N-IgG and S-IgG were tested 40 days after the onset of the disease at three different times, the positive rate of N-IgG (92.5%, 37/40) was higher than that of S-IgG (67.5%, 27/40), but the two structure protein antibodies were always lower than the total IgG. CONCLUSIONS: In SARS patients with definite clinical and etiological diagnosis, the highest positive rate of the antibodies against SARS-CoV was found at 21 - 40 days after the onset. IgM disappeared almost 500 days (91.8%) after the onset. Total IgG positive rate could reach 100% and 98.6% and the positivity might persist nearly two years. So it is speculated that the total IgG antibody may be positive 3 to 5 years after infection, but it seems that N-IgG and S-IgG keep positive shorter in time than total IgG antibody.

[The construction of pSecTag2/B vector modified by osteoprotegerin].

PURPOSE: In order to treat periodontitis by using tissue engineering and gene engineering technology, we constructed mammalian secreted expression pSecTag2/B-OPG vector, the cDNA sequence encoding osteoprotegerin(OPG) obtaining from 293 cell. METHODS: The primers were designed based on the human OPG cDNA sequence. Total mRNA was isolated from 293 cells and RT-PCR was performed .The fragment was recombined into pSecTag2/B vector and sequenced by automatic sequence analyzer. RESULTS: The sequences of OPG cDNA from 293 cell by RT-PCR were completely identical to the sequences provided by GenBank [gi:33878056]. After polymerase chain reaction and the recombinant plasmid digesting with Hind III,EcoR I and BamH I,1% agarose electrophoresis showed several fragments, which were consistent with predicted size. CONCLUSION: From cultured 293 cells the cDNA has been cloned and pSecTag2/B-OPG vector has been constructed successfully. Supported by Research Fund (No.03043304) from Anhui Provincial Natural Science Foundation.

RANTES gene single nucleotide polymorphisms and expression in patients with chronic hepatitis B virus infection.

BACKGROUND: Regulated on activation, normal T-cell expressed and secreted (RANTES) plays a critical role in T-lymphocyte activation and proliferation. The process is involved in both acute and chronic phases of inflammation. The present study was to ascertain the possible correlations between chronic hepatitis B virus (HBV) infection and the RANTES gene polymorphisms and their expression. METHODS: The study included 130 HBV negative healthy donors and 152 patients with chronic hepatitis B (CHB) virus infection. The polymerase chain reaction (PCR) and restriction fragment length polymorphisms (RFLPs) were used to detect RANTES gene single nucleotide polymorphisms (SNPs). RANTES levels in the platelet depleted plasma were detected by enzyme linked immunosorbent assay (ELISA). RESULTS: RANTES alleles -403G, -28C and In1.1T were the predominant alleles in the subjects studied. No significant correlation was found between CHB infection and the RANTES alleles, while a significant correlation was found between CHB infection and increased RANTES expression in platelet depleted plasma (P < 0.05). CONCLUSIONS: SNPs in RANTES gene do not affect chronic HBV infection or the outcome of interferon-alpha treatment in patients positive for HBV "e" antigen (HBeAg+). However, patients with CHB infection express the higher levels of plasma RANTES, which is thus associated with CHB infection.

[Immunological characteristics of primary biliary cirrhosis patients].

OBJECTIVE: To study the clinical significance of the immunological characteristics in patients with primary biliary cirrhosis (PBC). METHODS: 3000 patients with abnormal liver functions were examined for anti-nuclear antibodies (ANA), anti-mitochondrial antibodies (AMA), anti-smooth muscle antibodies (SMA) and anti-liver kidney microsomal antibody (LKM) using immunofluorescent assays (IFA). LKM-1, liver cytosolic-1 (LC-1), soluble liver antigen (SLA)/liver- pancreas antigen (LP) and subtype of AMA (M2, M4, M9) as well as ANA profile were detected by an immune blotting assay and an enzyme-linked immune absorbent assay (ELISA), respectively. Cytokines were tested by flow cytometry. RESULTS: Of the 3000 patients with liver diseases, 52 (1.7%) were diagnosed with PBC. All the PBC cases were positive for AMA and M2. 94% of them showed high titer of AMA (> or = 1:320), and in 79% of them M2 was >200 RU/L, and 78% of them were ANA positive. Three main fluorescent patterns of ANA seen were nuclear membrane, nuclear dots and centromere patterns. Sjogren's Syndrome A/B (SS-A/B), homogenous, nucleolar or nuclear granular patterns were seen in only a few patients. IgM, ALP and GGT in PBC patients were significantly higher than those in hepatitis B related liver cirrhosis patients. The levels of IL-6, IL-10, TNF-alpha and IFN-gamma in PBC patients were higher than in the normal controls. Among the 52 PBC patients, 5 had autoimmune liver disease overlap syndromes. Two of them were SLA/LP positive, indicated as AIH type III and PBC overlapping, and 1 was LKM-1 positive showing AIH type II overlapping PBC, and 2 had ANA positive and were identified as AIH and PBC by liver biopsy. CONCLUSION: The percentage of PBC in Chinese liver disease patients is about 1% to 2%. Most of the PBC patients have high levels of AMA and AMA-M2, IgM, ALP, GGT and several cytokines, indicating that abnormality of humeral and cellular immunity may be associated with the pathogenesis of PBC.

Effects of octreotide on expression of L-type voltage-operated calcium channels and on intracellular Ca2+ in activated hepatic stellate cells.

BACKGROUND: The contractility of hepatic stellate cells (HSCs) may play an important role in the pathogenesis of cirrhosis with portal hypertension. The aim of this study was to research the effects of octreotide, an analogue of somatostatin, on intracellular Ca2+ and on the expression of L-type voltage-operated calcium channels (L-VOCCs) in activated HSCs, and to try to survey the use of octreotide in treatment and prevention of cirrhosis with portal hypertension complications. METHODS: HSC-T6, an activated HSCs line, was plated on small glass coverslips in 35-mm culture dishes at a density of 1 x 10(5)/ml, and incubated in DMEM media for 24 hours. After the cells were loaded with Fluo-3/AM, intracellular Ca2+ was measured by Laser Scanning Confocal Microscopy (LSCM). The dynamic changes in activated HSCs of intracellular Ca2+, stimulated by octreotide, endothelin-1, and KCl, respectively, were also determined by LSCM. Each experiment was repeated six times. L-VOCC expression in HSCs was estimated by immunocytochemistry. RESULTS: After octreotide stimulation, a significant decrease in the intracellular Ca2+ of activated HSCs was observed. However, octreotide did not inhibit the increases in intracellular Ca2+ after stimulation by KCl and endothelin-1. Moreover, octreotide did not significantly affect L-VOCC expression. These results suggest that neither L-VOCC nor endothelin-1 receptors in activated HSCs are inhibited by octreotide. CONCLUSIONS: Octreotide may decrease portal hypertension and intrahepatic vascular tension by inhibiting activated HSCs contractility through decreases in intracellular Ca2+. The somatostatin receptors in activated HSCs may be inhibited by octreotide.