In the last decade, the Streptococcus pneumoniae population has changed, mainly due to the abuse of antibiotics. The aim of this study was to determine the genetic structure of 144 S. pneumonia serotype 14 isolates collected from children with acute respiratory infections during 1997–2012 in China.
All isolated pneumococci were tested for their sensitivity to 11 kinds of antibiotics with the E-test method or disc diffusion. The macrolides resistance genes ermB and mefA, as well as the sulfamethoxazole-trimethoprim resistance gene dihydrofolate reductase (DHFR) were detected by polymerase chain reaction (PCR). The sequence types (STs) were analyzed with multilocus sequence typing (MLST).
From 1997 to 2012, the percentage of serotype 14 S. pneumonia isolates in the whole isolates increased. All of the 144 serotype 14 S. pneumonia isolates were susceptible to amoxicillin-clavulanic acid, vancomycin and levofloxacin. No penicillin resistant isolate was found, and the intermediate rate was as low as 0.7 %. Erythromycin resistance was confirmed among 143 isolates. The ermB gene was determined in all erythromycin resistant isolates, and the mefA gene was positive additionally in 13 of them. The non-susceptibility rate to the tested cephalosporins increased from 1997–2012. All trimethoprim-resistant isolates contained the Ile100-Leu mutation. Overall, 30 STs were identified, among which ST876 was the most prevalent, followed by ST875. During the study period, the percentage of CC876 increased from 0 % in 1997–2000 to 96.4 % in 2010–2012, whereas CC875 decreased from 84.2 to 0 %. CC876 showed higher non-susceptibility rates to β-lactam antibiotics than CC875.
The percentage of serotype 14 S. pneumonia isolates increased over time in China. The increase of resistance to β-lactam antibiotics in this serotype isolates was associated with the spread of CC876.
Streptococcus pneumoniae; Serotypes; Antibiotic resistance; Children; Epidemiology
Objective: To evaluate the medium to long-term curative effects of surgical long segmental fixation and fusion in degenerative scoliosis (DS). Patients and methods: From January 2001 to December 2011, 56 DS patients underwent long segmental fixation and fusion. Clinical data, including visual analogue scale (VAS) scores, Oswestry disability index (ODI), lumbar lordosis angles, coronary Cobb angles and postoperative complications were followed up for 2 to 12 years postoperatively. Results: VAS and ODI scores were significantly improved 1 year postoperatively compared to the preoperative values (P = 0.000). Coronary Cobb angles were significantly improved three months postoperatively (P = 0.001) but ≥ 1 year after surgery there was no further significant improvement compared to the preoperative values (P = 0.585). The lumbar lordosis angle was not significantly changed postoperatively (P > 0.05). Conclusions: Favorable medium to long-term curative effects can be achieved by long segmental fixation and fusion. Ideally, the fixation and fusion segments should be longer than the segments affected by scoliosis. The restoration of the lumbar lordosis angle is the key to rebuilding sagittal balance, which is closely correlated with a patient’s clinical symptoms and quality of life.
Degeneration; internal fixation; postoperative complications; scoliosis
Aims: This study is to determine if expression level of microRNA-143 (miR-143) and cyclooxygenase-2 (COX-2) are related to the occurrence and development of degenerative scoliosis. Methods: A total of 30 patients with degenerative scoliosis, 30 patients with adolescent idiopathic scoliosis were enrolled in this study. For control, 30 patients with spinal burst fractures were also enrolled in this study. Real-time PCR and western blotting was performed to measure the expression levels of COX-2 in intervertebral disc tissues, peripheral blood and cerebrospinal. Expression levels of miR-143 in intervertebral disc tissues, peripheral blood and cerebrospinal were detected by real-time PCR. Results: The expression levels of COX-2 were increased in intervertebral disc tissues, peripheral blood and cerebrospinal of patients with degenerative scoliosis when compared with those of patients with adolescent idiopathic scoliosis and spinal burst fractures (P < 0.05). However, the expression levels of miR-143 were decreased in intervertebral disc tissues, peripheral blood and cerebrospinal of patients with degenerative scoliosis when compared with those of patients with adolescent idiopathic scoliosis and spinal burst fractures (P < 0.05). Conclusions: COX-2 is highly expressed whereas miR-143 is lowly expressed in patients with degenerative scoliosis. Decreased expression of miR-143 may be related to the aggravation of degenerative scoliosis by regulation of COX-2.
Degenerative scoliosis; cyclooxygenase-2; microRNA-143; intervertebral disc tissues
Commensal bacteria and innate immunity play a major role in the development of colorectal cancer (CRC). We propose that selected commensals polarise colon macrophages to produce endogenous mutagens that initiate chromosomal instability (CIN), lead to expression of progenitor and tumour stem cell markers, and drive CRC through a bystander effect.
Primary murine colon epithelial cells were repetitively exposed to Enterococcus faecalis-infected macrophages, or purified trans-4-hydroxy-2-nonenal (4-HNE)—an endogenous mutagen and spindle poison produced by macrophages. CIN, gene expression, growth as allografts in immunodeficient mice were examined for clones and expression of markers confirmed using interleukin (IL) 10 knockout mice colonised by E. faecalis.
Primary colon epithelial cells exposed to polarised macrophages or 4-hydroxy-2-nonenal developed CIN and were transformed after 10 weekly treatments. In immunodeficient mice, 8 of 25 transformed clones grew as poorly differentiated carcinomas with 3 tumours invading skin and/or muscle. All tumours stained for cytokeratins confirming their epithelial cell origin. Gene expression profiling of clones showed alterations in 3 to 7 cancer driver genes per clone. Clones also strongly expressed stem/progenitor cell markers Ly6A and Ly6E. Although not differentially expressed in clones, murine allografts positively stained for the tumour stem cell marker doublecortin-like kinase 1. Doublecortin-like kinase 1 and Ly6A/E were expressed by epithelial cells in colon biopsies for areas of inflamed and dysplastic tissue from E. faecalis-colonised IL-10 knockout mice.
These results validate a novel mechanism for CRC that involves endogenous CIN and cellular transformation arising through a microbiome-driven bystander effect.
Colon Carcinogenesis; Colonic Bacteria; Genetic Instability; Macrophages; Stem Cells
Haemophilus influenzae is one of the main pathogens that cause community-acquired respiratory infections in children. Our previous study showed that H. influenzae is the second most common pathogen causing pneumonia and accounts for 30–50% of bacterial meningitis among Chinese children. H. influenzae carriage in children and its resistance to commonly used antimicrobials varies widely both geographically and over time.
Surveys of the nasopharyngeal carriage of H. influenzae in children younger than 5 years of age with acute respiratory tract infection (ARI) were conducted in Beijing Children’s Hospital, China in 2000, 2002, 2010, and 2012. The overall annual carriage rates of H. influenzae among children younger than 5 years of age with ARI were 35.5%, 20.6%, 14.4%, and 18.7%, and the percentages of H. influenzae isolates producing β-lactamase were 4%, 13%, 27.1%, and 31%, respectively. The percentages of susceptibility to ampicillin progressively decreased from 96% (2000) to 87% (2002) to 63% (2010) to 61% (2012). All of the ampicillin-resistant isolates were found to be beta-lactamase producers. The susceptibility to tetracycline increased from 54% (2000) to 60% (2002) to 91.5% (2010) to 94.5% (2012). No statistically significant differences were observed in the susceptibility to cefaclor, cefuroxime, sulfamethoxazole, and chloramphenicol. Amoxicillin/clavulanic acid and ceftriaxone were the most effective antimicrobials for the isolates of H. influenzae across the 10-year period.
This report on the H. influenzae carriage rates in children and the susceptibility of these bacteria to commonly used antibiotics showed that H. influenzae carriage decreased from 2000 to 2012. Additionally, the percentage of β-lactamase-producing isolates increased while their susceptibility to ampicillin progressively decreased during this time. These results indicate that the appropriate empirical antimicrobial therapy should be changed for pediatric patients in China.
Haemophilus influenzae; Antimicrobial susceptibility; Acute upper respiratory tract infection; Pediatrics
Background: Many studies have examined the association between interleukin-6 (IL-6) gene polymorphisms and bone mineral density (BMD). However, the results remain conflicting. To assess the relationship more precisely, a meta-analysis was performed. Methods: The PubMed, Embase, Chinese BioMedical Literature (CBM), Wanfang, and China National Knowledge Infrastructure (CNKI) database were searched for relevant articles published up to March 2013. Weighted mean difference (WMD) and 95% confidence interval (95% CI) were calculated using a fixed-effects or random-effects model. Results: A total of 16 articles with 11,957 subjects were investigated in this meta-analysis. Overall, −634C/G polymorphism was significantly associated with BMD at the femoral neck (WMD, −0.016 g/cm2; 95% CI, −0.028 to −0.003 g/cm2), lumbar spine (WMD, −0.049 g/cm2; 95% CI, −0.069 to −0.030 g/cm2), and whole body (WMD, −0.023 g/cm2; 95% CI, −0.037 to −0.009 g/cm2) for GG versus CC+CG. In subgroup analyses stratified by ethnicity, individuals carrying −634GG genotype had a significantly lower mean BMD at any skeletal site examined, compared with individuals with −634CC or −634CG genotype in Asian populations. For −174G/C polymorphism, the BMD differences between CC+CG and GG genotype were 0.004 g/cm2 at the distal radius (95% CI, 0.004 to 0.005 g/cm2), 0.011 g/cm2 at the trochanter (95% CI, 0.002 to 0.020 g/cm2), and 0.017 g/cm2 at the Ward's triangle (95% CI, 0.003 to 0.032 g/cm2). No significant publication bias was observed in either the −634C/G or −174G/C polymorphism. Conclusions: This suggests that there are modest effects of the −634C/G and −174G/C polymorphisms on BMD. Large-scale and well-designed studies are required to further investigate gene–gene and gene–environment interactions on IL-6 polymorphisms and BMD in various populations.
This study aims to investigate the clinical features of invasive community-acquired Staphylococcus aureus (CA-SA) infection in Chinese children and analyze its molecular features.
Clinical data and invasive CA-SA isolates were prospectively collected. Pediatric risk of mortality (PRISM) score was used for disease severity measurement. Molecular typing was then performed, followed by expression analysis for virulence genes.
Among 163 invasive CA-SA infection cases, 71 (43.6%) were methicillin-resistant SA (MRSA) infections and 92 (56.4%) were methicillin-susceptible SA (MSSA). A total of 105 (64.4%) children were younger than 1 year old, and 79.7% (129/163) were under 3 years age. Thirteen kinds of diseases were observed, in which bacteremia and pneumonia accounted for 65.6% (107/163) and 52.8% (86/163), respectively. A total of 112 (68.1%) patients had two or more infective sites simultaneously, and four cases (2.5%) died. CA-MSSA more frequently caused multi-sites infections, bacteremia, and musculoskeletal infection than MRSA. A total of 25 sequence types (STs) were detected. MRSA mainly comprised ST59 (49/71, 69%), whereas the most frequent clonotypes were ST88 (15/92, 16.3%), ST25 (13/92, 14.1%), ST7 (13/92, 14.1%), ST2155 (12/92, 13%), and ST188 (9/92, 9.8%) for MSSA. Seven STs were common to both MSSA and MRSA groups. No differences in clinical presentation or PRISM score were found between the two groups or among different ST. The expression levels of the four known virulence genes varied among the six main ST clones.
Invasive CA-SA infections were characterized by high incidence and multi-site infections in young children in China. The clinical manifestations of CA-MSSA were more frequently associated with multi-site infections, bacteremia and musculoskeletal infection than those of CA-MRSA. Isolated genotypes may be relevant to the expressions of virulence genes, but not to clinical manifestations.
Electronic supplementary material
The online version of this article (doi:10.1186/s12879-014-0582-4) contains supplementary material, which is available to authorized users.
Staphylococcus aureus; Community-acquired; Invasive infection; Child; Molecular epidemiology
Bacterial meningitis is more common in the neonatal period than any other time in life; however, it is still a challenge for the evidence based diagnosis. Strategy for identification of neonatal bacterial meningitis pathogens is presented by evaluating three different available methods to establish evidence-based diagnosis for neonatal bacterial meningitis.
The cerebrospinal fluid samples from 56 neonates diagnosed as bacterial meningitis in 2009 in Beijing Children’s Hospital were analyzed in the study. Two PCR based molecular assays, real-time fluorescence quantitative PCR (RT-PCR) and multiplex PCR based-reverse line blot hybridization (mPCR/RLB), were used to assess 7 common neonatal meningitis bacterial pathongens, including Escherichia coli, Staphylococcus aureus, Listerisa monocytogenes, Neisseria meningitidis, Haemophilus influenzae, Streptococcus pneumoniae, and Streptococcus agalactiae. The findings in examinations of two assays were compared with the results obtained bacterial culture tests.
Bacterial meningitis was identified in five cases (9%) by CSF cultures, 25 (45%) by RT-PCR and 16 (29%) by mPCR/RLB. One strain of S. epidermidis and one of E. faecalis were identified using mPCR/RLB but not by RT-PCR. In contrast, cultures identified one strain of S. pneumoniae which was missed by both PCR assays. Overall, the bacterial pathogens in 28 cases were identified with these three methods. Both RT-PCR and mPCR/RLB assays were more sensitive than bacterial culture, (p < 0.05).
Our study confirmed that both RT-PCR and mPCR/RLB assays have better sensitivity than bacterial culture. They are capable of detecting the pathogens in CSF samples with negative culture results.
Neonate; Bacterial meningitis; Bacterial pathogens identification; Multiplex real-time fluorescence quantitative PCR (RT-PCR); Multiplex PCR based-reverse line blot hybridization (mPCR/RLB) assay; Bacteria culture
In this study, we defined the population biology of serogroup 6 Streptococcus pneumoniae collected in China and their antibiotic resistance profiles.
The serotypes of 225 S. pneumoniae strains isolated between 1997 and 2011 were identified with the Quellung reaction and serotype-specific PCR. All isolated pneumococci were tested for their sensitivity to 11 kinds of antibiotics with the E-test method or disc diffusion. The sequence types (STs) were analyzed with multilocus sequence typing (MLST).
The frequencies of serotypes and subtypes 6A, 6B-I, 6B-II, 6C, and 6D among the 225 isolates were 46.7% (105/225), 19.6% (44/225), 25.8% (58/225), 6.2% (14/225), and 1.8% (4/225), respectively. Serotype 6E was not found in the serotype 6A isolates, and neither serotype 6F nor 6G was identified in any isolate. MLST analysis revealed 58 STs. The most common STs were ST982 (23.1%), ST90 (14.7%), ST4542 (7.6%), and ST2912 (4.9%). The rates of clonal complex 90 (CC90) and CC386 among the oral-penicillin-nonsusceptible isolates decreased over the years, whereas the rates of CC855 and CC3173 increased. The four CCs had similar penicillin MIC distributions, with a maximum MIC of 2 μg/ml.
This study identified the serotypes/subtypes and CCs/STs of group 6 S. pneumoniae present in China. No salient antibiotic-resistant clones were isolated among the serogroup 6 S. pneumoniae.
Streptococcus pneumoniae; Serotypes; Antibiotic resistance; Children; Epidemiology
Hypoxia preconditioning has been confirmed as an effective strategy to enhance the therapeutic potentials of mesenchymal stem cells (MSCs), such as for myocardial ischemia. However, whether hypoxia preconditioning would produce beneficial effects on MSC-based renal repair has not been demonstrated. In the study, we aimed to determine the feasibility and efficacy of hypoxia preconditioning to enhance MSC-based therapy of acute kidney injury (AKI). MSCs were isolated from human adipose tissues. The paracrine effects of MSCs under normoxia and hypoxia were determined in vitro. Rats of AKI were induced by kidney I/R surgery and randomly divided into three groups: I/R control receiving PBS injection; MSC group receiving normal MSC injection; hypoMSC group receiving hypoxia-preconditioned MSC injection. It was demonstrated in vitro that paracrine effects of MSCs were significantly enhanced, especially angiogenic factors. Dihydroethidium (DHE) staining showed that antioxidative activities of MSCs were significantly enhanced by hypoxia stimulation. Vascularization, apoptosis, and histological injury were all significantly improved in hypoMSC injected group compared with that in control and MSC injected groups. Finally, the renal function was also significantly improved in hypoMSC injected group compared with that in the other two groups as assessed by the serum creatinine and BUN levels.
Infection of macrophages by the human intestinal commensal Enterococcus faecalis generates DNA damage and chromosomal instability in mammalian cells through bystander effects. These effects are characterized by clastogenesis and damage to mitotic spindles in target cells and are mediated, in part, by trans-4-hydroxy-2-nonenal (4-HNE). In this study we investigated the role of cyclooxygenase (COX) and lipoxygenase (LOX) in producing this reactive aldehyde using E. faecalis-infected macrophages and interleukin-10 knockout mice colonized with this commensal. 4-HNE production by E. faecalis-infected macrophages was significantly reduced by COX and LOX inhibitors. The infection of macrophages led to decreased Cox1 and Alox5 expression while COX-2 and 4-HNE increased. Silencing Alox5 and Cox1 with gene-specific siRNAs had no effect on 4-HNE production. In contrast, silencing Cox2 significantly decreased 4-HNE production by E. faecalis-infected macrophages. Depleting intracellular glutathione increased 4-HNE production by these cells. Next, to confirm COX-2 as a source for 4-HNE, we assayed the products generated by recombinant human COX-2 and found 4-HNE in a concentration-dependent manner using arachidonic acid as a substrate. Finally, tissue macrophages in colon biopsies from interleukin-10 knockout mice colonized with E. faecalis were positive for COX-2 by immunohistochemical staining. This was associated with increased staining for 4-HNE-protein adducts in surrounding stroma. These data show that E. faecalis, a human intestinal commensal, can trigger macrophages to produce 4-HNE through COX-2. Importantly, it reinforces the concept of COX-2 as a procarcinogenic enzyme capable of damaging DNA in target cells through bystander effects that contribute to colorectal carcinogenesis.
Cyclooxygenase-2; 4-hydroxy-2-nonenal; carcinogenesis; colorectal cancer; macrophage; commensal
Dendritic cells (DCs), as the most potent professional antigen presenting cells, play a crucial role in both innate and adaptive immune systems. Genomic bacterial DNA mimicked by unmethylated CpG motifs is discovered to possess immunostimulatory effects. CpG-DNA recognized by Toll-like receptor 9 (TLR9) on DCs arouses many immune diseases (such as cancer, viral infection, and autoimmune disorders). In this study we investigated the effects of FC-98 on CpG-induced bone marrow-derived DCs (BMDCs). The results showed that FC-98 significantly inhibited the CpG-induced BMDCs maturation and function by suppressing the expression of surface markers (CD40, CD80, CD86, and MHCII). Moreover, FC-98 downregulated the expression of C-X-C motif chemokine 10 (CXCL-10) both at the mRNA and protein level after CpG induction. Meanwhile, FC-98 markedly affected the migration of BMDCs to T cells without affecting their endocytosis capacity. Furthermore, FC-98 was confirmed to decrease CXCL-10 expression by inhibiting CpG-induced activation of MAPKs (ERK, JNK, and p38) and STAT1 signaling. Overall, these results suggested that FC-98 was a potential molecule in the treatment of CXCL-10-mediated immune diseases.
Streptococcus pneumoniae is a primary cause of bacterial infection in humans. Here, we present the complete genome sequence of S. pneumoniae strain A026, which is a multidrug-resistant strain isolated from cerebrospinal fluid.
Macrophage-induced bystander effects have been implicated as an important mediator of chromosomal instability and colon cancer triggered by Enterococcus faecalis a human intestinal commensal bacteria. There is little understanding about how inflammatory cytokines mediate bystander effects, but questions in this area are important because of the pivotal contributions made by inflammatory processes to cancer initiation and progression. Here we report that the central pro-inflammatory cytokine TNF-α acts as a diffusible mediator of the bystander effects induced by macrophages, an effect caused by a proliferation of macrophages that trigger epithelial cell production of Netrin-1, a neuronal guidance molecule. TNF-α-mediated bystander assays employed a murine co-culture system of primary colonic epithelial cells and E. faecalis-infected macrophages (in vitro), with an IL-10-deficient mouse model of colon cancer that involves long-term colonization with E. faecalis (in vivo). In cell co-cultures, we observed increased expression of the TNF-α receptor Tnfrsf1b and Netrin-1. These effects were blocked by anti-TNF-α antibody or by pretreatment with an inhibitor of NF-κB signaling. RNAi-mediated attenuation of Tnfrsf1b decreased TNF-α-induced netrin-1 production and augmented epithelial cell apoptosis in culture. Extending these observations, colon biopsies from E. faecalis-colonized IL-10−/− mice exhibited crypt hyperplasia and increased staining for macrophages, TNF-α, netrin-1, NF-κB, Tnfrsf1b and the proliferation marker PCNA, also displaying a reduction in epithelial cell apoptosis. Together, our results define a pathway for macrophage-induced bystander effects in which TNF-α triggers TNFRSF1b receptor signaling leading to increased production of Netrin-1, crypt hyperplasia and decreased epithelial cell apoptosis. In elucidating an important commensal-associatedpro-inflammatory mechanism in the intestinal microenvironment, our work highlights the role of Netrin-1 and a specific TNF-α receptor as candidate targets to prevent or treat colorectal cancer.
CRC Model; Bystander effect; TNF-α; Netrin-1; Tnfrsf1b; macrophage; tumorigenesis; colorectal cancer
Intestinal commensal bacteria have recently been shown to trigger macrophages to produce diffusible clastogens (or chromosome-breaking factors) through a bystander effect (BSE) that mediates DNA damage and induces chromosomal instability in neighboring cells. Colon macrophages appear central to colon carcinogenesis and BSE through the expression of tumor necrosis factor-α (TNF-α) and cyclooxygenase-2 (COX-2). The former induces netrin-1, a regulator of intestinal epithelial cell apoptosis, and the latter generates trans-4-hydroxy-2-nonenal (4-HNE), an endogenous mutagen. To test whether colon macrophages are key effectors for BSE, we depleted these cells in interleukin-10 knockout mice colonized with Enterococcus faecalis using encapsulated liposomal clodronate (ELC), a bisphosphonate that causes macrophage apoptosis. We observed that E. faecalis polarizes colon macrophages to an M1 phenotype. In addition, depleting these cells suppressed COX-2 and TNF-α, blocked the formation of 4-HNE protein adducts, and inhibited up-regulation of netrin-1—all markers for BSE. Finally, treatment with ELC prevented colitis, β-catenin activation, and cancer formation. These results show that selected human commensals can polarize colon macrophages to the M1 phenotype and, when activated, serve as the key effector for bacterial-induced BSE. Our findings suggest that depleting M1-polarized macro-phages is a mechanism for the chemopreventive activity of bisphosphonates and that it represents a new strategy for preventing colon cancer induced by intestinal commensals.
Detailed molecular analyses of Clonal Complex 59 (CC59) methicillin-resistant Staphylococcus aureus (MRSA) isolates from children in seven major cities across Mainland China were examined. A total of 110 CC59 isolates from invasive and non-invasive diseases were analyzed by multilocus sequence typing (MLST), Staphylococcus cassette chromosome mec (SCCmec) typing, staphylococcal protein A (spa) typing and pulsed-field gel electrophoresis (PFGE). Antibiotics susceptibilities, carriage of plasmids and 42 virulence genes and the expression of virulence factors were examined. ST59 (101/110, 91.8%) was the predominant sequence type (ST), while single locus variants (SLVs) belonging to ST338 (8/110, 7.3%) and ST375 (1/110, 0.9%) were obtained. Three SCCmec types were found, namely type III (2.7%), type IV (74.5%) and type V (22.7%). Seven spa types including t437, which accounted for 87.3%, were determined. Thirteen PFGE types were obtained. PFGE types A and B were the major types totally accounting for 81.8%. The dominant clone was ST59-t437-IVa (65.5%), followed by ST59-t437-V (14.5%). The positive rate of luks-PV and lukF-PV PVL encoding (pvl) gene was 55.5%. Plasmids were detected in 83.6% (92/110) of the strains. The plasmid size ranging from 23.4 kb to 50 kb was most prevalent which accounted for 83.7% (77/92). A significantly lower expression of hla was found in ST59-t437-IVa compared with ST59-t437-V. Among the 110 cases, 61.8% of the patients were less than 1 year old. A total of 90 cases (81.8%) were community-associated (CA) infections whereas 20 cases (18.2%) were hospital-associated (HA) infections. Out of the 110 patients, 36.4% (40/110) were diagnosed with invasive infectious diseases in which ST59-t437-IVa accounted for 67.5% (27/40). In brief, ST59-t437-IVa was proved as the dominant clone in CC59 MRSA strains. The carriage rate of pvl gene was high. CC59 MRSA could result in CA and HA infections. The majortiy of MRSA infection children were in young age.
Resveratrol (RSV), a natural compound present in the skin and seeds of red grapes, is considered a phytoestrogen and has structural similarity to the synthetic estrogen diethylstilbestrol. RSV inhibits tumor cell growth in estrogen receptor-positive (ER+) and negative (ER-) breast cancer cell lines resulting in cell specific regulation of the G1/S and G2/M stages of the cell cycle. However apoptotic cell death was only observed in ER+ MCF-7 cells. In this study, we designed and synthesized boronic acid derivative of RSV and evaluated their biological effects on ER+ MCF-7 breast cancer cells. The trans-4 analog inhibited the growth of MCF-7 cells and is not a substrate for p-glycoprotein. The trans-4 analog induces G1 cell cycle arrest, which coincides with marked inhibition of G1 cell cycle proteins and a greater pro-apoptotic effect. Finally, the trans-4 analog had no effect on the estrogen-stimulated growth of MCF-7 cells. Our results demonstrate that the trans-4 analog inhibits MCF-7 breast cancer cells by a different mechanism of action than that of RSV (S-phase arrest), and provides a new class of novel boronic acids of RSV that inhibit breast cancer cell growth.
anticancer agent; boronic acid; estrogen receptor positive breast cancer; resveratrol
Inhibitors of histone deacetylases (HDAC) are an important emerging class of drugs for the treatment of cancers. HDAC inhibitors are currently under evaluation in clinical trials as single agents and as sensitizers in combinations with chemotherapies and radiation therapy. Although these drugs have important effects on cancer cell growth and functions, the mechanisms underlying HDAC inhibitor activities remain to be fully defined. By using rational drug design, compound 2, a fluorescent class II HDAC targeting inhibitor, was synthesized and observed to accumulate in the cytoplasmic compartments of treated cells, but not in the nuclei. Furthermore, immunostaining of inhibitor exposed cells for HDAC4 showed accumulation of this enzyme in the cytoplasmic compartment with concomitant increased acetylation of tubulin and nuclear histones. These observations support a mechanism by which nuclear histone acetylation is increased as a result of HDAC4 trapping and sequestration in the cytoplasm after binding to compound 2. The HDAC inhibitor offers potential as a novel theranostic agent, combining diagnostic and therapeutic properties in the same molecule.
To provide guidance for clinical disease prevention and treatment, this study examined the epidemiology, antibiotic susceptibility, and serotype distribution of Streptococcus pneumoniae (S. pneumoniae) associated with invasive pneumococcal diseases (IPDs) among children less than 14 years of age in Shenzhen, China.
Materials and Methods
All the clinical strains were isolated from children less than 14 years old from January 2009 to August 2012. The serotypes and antibiotic resistance of strains of S. pneumoniae were determined using the capsular swelling method and the E-test.
A total of 89 strains were isolated and 87 isolates were included. The five prevailing serotypes were 19F (28.7%), 14 (16.1%), 23F (11.5%), 19A (9.2%) and 6B (6.9%). The most common sequence types (ST) were ST271 (21.8%), ST876 (18.4%), ST320 (8.0%) and ST81 (6.9%) which were mainly related to 19F, 14, 19A and 23F, respectively. The potential coverage by 7-, 10-, and 13-valent pneumococcal conjugate vaccine were 77.0%, 77.0%, and 89.7%, respectively. Among the 87 isolates investigated, 11.5% were resistant to penicillin, and for meningitis isolates, the resistance rate was 100%. Multi-drug resistance (MDR) was exhibited by 49 (56.3%) isolates. Eighty-four isolates were resistance to erythromycin, among which, 56 (66.7%) carried the ermB gene alone and 28 (33.3%) expressed both the ermB and mefA/E genes.
The potential coverage of PCV13 is higher than PCV7 and PCV10 because high rates of serotypes 19A and 6A in Shenzhen. The clinical treatment of IPD needs a higher drug concentration of antibiotics. Continued surveillance of the antimicrobial susceptibility and serotypes distribution of IPD isolates may be necessary.
BACKGROUND & AIMS
Enterococcus faecalis is a human intestinal commensal that produces extracellular superoxide and promotes chromosome instability via macrophage-induced bystander effects. We investigated the ability of 4-hydroxy-2-nonenal (4-HNE)—a diffusible breakdown product of ω-6 polyunsaturated fatty acids—to mediate these effects.
4-HNE was purified from E faecalis-infected macrophages; its genotoxicity was assessed in human colon cancer (HCT116) and primary murine colon epithelial (YAMC) cell lines.
4-HNE induced G2M cell cycle arrest, led to formation γH2AX foci, and disrupted the mitotic spindle in both cell lines. Binucleate tetraploid cells that formed following incubation with 4-HNE were associated with the activation of stathmin and microtubule catastrophe. Silencing glutathione S-transferase α-4, a scavenger of 4-HNE, increased susceptibility of epithelial cells to 4-HNE-induced genotoxicity. Interleukin-10 knockout mice colonized with superoxide-producing E faecalis developed inflammation and colorectal cancer, whereas colonization with a superoxide-deficient strain resulted in inflammation but not cancer. 4-HNE-protein adducts were found in the lamina propria and macrophages in areas of colorectal inflammation.
4-HNE can act as an autochthonous mitotic spindle poison in normal colonic epithelial and colon cancer cells. This finding links the macrophage-induced bystander effects to colorectal carcinogenesis.
CRC model; genetic; cell division; mitosis
Antimicrobial resistance in Streptococcus pneumoniae remains a serious concern worldwide, particularly in Asian countries, despite the introduction of heptavalent pneumococcal conjugate vaccine (PCV7). The Asian Network for Surveillance of Resistant Pathogens (ANSORP) performed a prospective surveillance study of 2,184 S. pneumoniae isolates collected from patients with pneumococcal infections from 60 hospitals in 11 Asian countries from 2008 to 2009. Among nonmeningeal isolates, the prevalence rate of penicillin-nonsusceptible pneumococci (MIC, ≥4 μg/ml) was 4.6% and penicillin resistance (MIC, ≥8 μg/ml) was extremely rare (0.7%). Resistance to erythromycin was very prevalent in the region (72.7%); the highest rates were in China (96.4%), Taiwan (84.9%), and Vietnam (80.7%). Multidrug resistance (MDR) was observed in 59.3% of isolates from Asian countries. Major serotypes were 19F (23.5%), 23F (10.0%), 19A (8.2%), 14 (7.3%), and 6B (7.3%). Overall, 52.5% of isolates showed PCV7 serotypes, ranging from 16.1% in Philippines to 75.1% in Vietnam. Serotypes 19A (8.2%), 3 (6.2%), and 6A (4.2%) were the most prominent non-PCV7 serotypes in the Asian region. Among isolates with serotype 19A, 86.0% and 79.8% showed erythromycin resistance and MDR, respectively. The most remarkable findings about the epidemiology of S. pneumoniae in Asian countries after the introduction of PCV7 were the high prevalence of macrolide resistance and MDR and distinctive increases in serotype 19A.
Eighty group G streptococcal stains were collected from Chinese children. Susceptibility testing was done by a double-dilution and a disk diffusion method. PCR was used to test drug-resistant genes, and the χ2 test and definite probability methods were used to test for statistically significant differences among the three groups. Thirty-four isolates (42.5%) showed resistance to erythromycin. There are differences between the resistance characteristics of group G streptococci from different regions of China.
The activities of the bifunctional folate pathway enzyme dihydrofolate synthase–folylpolyglutamate synthase from Plasmodium falciparum are characterised with respect to their kinetics, substrate specificities and responses to folate analogue inhibitors.
Unusually for a eukaryote, the malaria parasite Plasmodium falciparum expresses dihydrofolate synthase (DHFS) and folylpolyglutamate synthase (FPGS) as a single bifunctional protein. The two activities contribute to the essential pathway of folate biosynthesis and modification. The DHFS activity of recombinant PfDHFS–FPGS exhibited non-standard kinetics at high co-substrate (glutamate and ATP) concentrations, being partially inhibited by increasing concentrations of its principal substrate, dihydropteroate (DHP). Binding of DHP to the catalytic and inhibitory sites exhibited dissociation constants of 0.50 μM and 1.25 μM, respectively. DHFS activity measured under lower co-substrate concentrations, where data fitted the Michaelis–Menten equation, yielded apparent Km values of 0.88 μM for DHP, 22.8 μM for ATP and 5.97 μM for glutamate. Of the substrates tested in FPGS assays, only tetrahydrofolate (THF) was efficiently converted to polyglutamylated forms, exhibiting standard kinetics with an apparent Km of 0.96 μM; dihydrofolate, folate and the folate analogue methotrexate (MTX) were negligibly processed, emphasising the importance of the oxidation state of the pterin moiety. Moreover, MTX inhibited neither DHFS nor FPGS, even at high concentrations. Conversely, two phosphinate analogues of 7,8-dihydrofolate that mimic tetrahedral intermediates formed during DHFS- and FPGS-catalysed glutamylation were powerfully inhibitory. The Ki value of an aryl phosphinate analogue against DHFS was 0.14 μM and for an alkyl phosphinate against FPGS 0.091 μM, with each inhibitor showing a high degree of specificity. This, combined with the absence of DHFS activity in humans, suggests PfDHFS–FPGS might represent a potential new drug target in the previously validated folate pathway of P. falciparum.
Antifolate inhibitor studies; Biphasic kinetics; Folate metabolism; Methotrexate; Phosphinate analogues of folate; Substrate specificity; DHF, dihydrofolate; DHFR, dihydrofolate reductase; DHFS, dihydrofolate synthase; DHP, dihydropteroate; DHPS, dihydropteroate synthase; DTT, dithiothreitol; FPGS, folylpolyglutamate synthase; GTPCH, GTP cyclohydrolase I; HPPK, hydroxymethyldihydropterin pyrophosphokinase; MTX, methotrexate; pAB, para-aminobenzoate; PTPS, pyruvoyltetrahydropterin synthase III; PYR, pyrimethamine; SDX, sulfadoxine; THF, tetrahydrofolate
Quinolone resistance in Enterobacteriaceae results mainly from mutations in type II DNA topoisomerase genes and/or changes in the expression of outer membrane and efflux pumps. Several recent studies have indicated that plasmid-mediated resistance mechanisms also play a significant role in fluoroquinolone resistance, and its prevalence is increasing worldwide. In China, the presence of the qnr gene in the clinical isolates of Enterobacteriaceae has been reported, but this transmissible quinolone resistance gene has not been detected in strains isolated singly from pediatric patients. Because quinolones associated with a variety of adverse side effects on children, they are not authorized for pediatric use. This study therefore aimed to investigate the presence of the qnr gene in clinical isolates of E. coli and K. pneumoniae from pediatric patients in China.
A total 213 of non-repetitive clinical isolates resistant to ciprofloxacin from E. coli and K. pneumoniae were collected from hospitalized patients at five children's hospital in Beijing, Shanghai, Guangzhou, and Chongqing. The isolates were screened for the plasmid-mediated quinolone resistance genes of qnrA, qnrB, and qnrS by PCR. Transferability was examined by conjugation with the sodium azide-resistant E. coli J53. All qnr-positive were analyzed for clonality by enterobacterial repetitive intergenic consensus (ERIC)-PCR.
The study found that 19 ciprofloxacin-resistant clinical isolates of E. coli and K. pneumoniae were positive for the qnr gene, and most of the qnr positive strains were ESBL producers. Conjugation experiments showed that quinolone resitance could be transferred to recipients. Apart from this, different DNA banding patterns were obtained by ERIC-PCR from positive strains, which means that most of them were not clonally related.
This report on transferable fluoroquinolone resistance due to the qnr gene among E. coli and K. pneumoniae strains indicated that plasmid-mediated quinolone resistance has emerged in pediatric patients in China.
A total of 685 clinical Streptococcus pneumoniae isolates from patients with pneumococcal diseases were collected from 14 centers in 11 Asian countries from January 2000 to June 2001. The in vitro susceptibilities of the isolates to 14 antimicrobial agents were determined by the broth microdilution test. Among the isolates tested, 483 (52.4%) were not susceptible to penicillin, 23% were intermediate, and 29.4% were penicillin resistant (MICs ≥ 2 mg/liter). Isolates from Vietnam showed the highest prevalence of penicillin resistance (71.4%), followed by those from Korea (54.8%), Hong Kong (43.2%), and Taiwan (38.6%). The penicillin MICs at which 90% of isolates are inhibited (MIC90s) were 4 mg/liter among isolates from Vietnam, Hong Kong, Korea, and Taiwan. The prevalence of erythromycin resistance was also very high in Vietnam (92.1%), Taiwan (86%), Korea (80.6%), Hong Kong (76.8%), and China (73.9%). The MIC90s of erythromycin were >32 mg/liter among isolates from Korea, Vietnam, China, Taiwan, Singapore, Malaysia, and Hong Kong. Isolates from Hong Kong showed the highest rate of ciprofloxacin resistance (11.8%), followed by isolates from Sri Lanka (9.5%), the Philippines (9.1%), and Korea (6.5%). Multilocus sequence typing showed that the spread of the Taiwan19F clone and the Spain23F clone could be one of the major reasons for the rapid increases in antimicrobial resistance among S. pneumoniae isolates in Asia. Data from the multinational surveillance study clearly documented distinctive increases in the prevalence rates and the levels of antimicrobial resistance among S. pneumoniae isolates in many Asian countries, which are among the highest in the world published to date.