This study focuses on the implementation of modulated modularity clustering (MMC) a new cluster algorithm for the identification of molecular signatures of preeclampsia and intrauterine growth restriction (IUGR), and the identification of affected microRNAs
Eighty-six human placentas from normal (40), growth-restricted (27), and preeclamptic (19) term pregnancies were profiled using Illumina Human-6 Beadarrays. MMC was utilized to generate modules based on similarities in placental transcriptome. Gene Set Enrichment Analysis (GSEA) was used to predict affected microRNAs. Expression levels of these candidate microRNAs were investigated in seventy-one human term placentas as follows: control (29); IUGR (26); and preeclampsia (16).
MMC identified two modules, one representing IUGR placentas and one representing preeclamptic placentas. 326 differentially expressed genes in the module representing IUGR and 889 differentially expressed genes in a module representing preeclampsia were identified. Functional analysis of molecular signatures associated with IUGR identified P13K/AKT, mTOR, p70S6K, apoptosis and IGF-1 signaling as being affected. Analysis of variance of GSEA-predicted microRNAs indicated that miR-194 was significantly down-regulated both in preeclampsia (p=0.0001) and IUGR (p=0.0304), and miR-149 was significantly down-regulated in preeclampsia (p=0.0168).
Implementation of MMC, allowed identification of genes disregulated in IUGR and preeclampsia. The reliability of MMC was validated by comparing to previous linear modeling analysis of preeclamptic placentas.
MMC allowed the elucidation of a molecular signature associated with preeclampsia and a subset of IUGR samples. This allowed the identification of genes, pathways, and microRNAs affected in these diseases.
Placenta; preeclampsia; IUGR; miR-194; miR-149
Aberrant expression of microRNAs plays vital roles in tumor development and progression. As transcription factors (TFs) are the critical components of signaling cascades, specific targeting effects of microRNAs to specific TFs may determine the role of microRNAs in different cancers. In this study, we identified Nuclear Factor I/B (NFIB) as one of the targets of miR-365 which was previously verified as an onco-miR in cutaneous squamous cell carcinoma (CSCC). Down-regulation of NFIB was a general feature in both CSCC cell lines and tumors from patients which show drastically up-regulated miR-365 expression levels. The siRNA-based knockdown of NFIB mimic the carcinogenic transformation of normal cells by ectopically expression of miR-365 which indicates depletion of NFIB is necessary for miR-365 to exert its pro-carcinogenic function. NFIB may represent a functional barrier targeted by miR-365 to the development of CSCC. Further studies also discovered a conserved feedback regulatory circuitry formed by NFIB and miR-365 in CSCC development which may be potentially utilized as therapeutic target to improve the clinical CSCC treatment.
Plants lack a germ line; consequently, during reproduction adult somatic cells within flowers must switch from mitotic proliferation to meiosis. In maize (Zea mays L.) anthers, hypoxic conditions in the developing tassel trigger pre-meiotic competence in the column of pluripotent progenitor cells in the center of anther lobes, and within 24 hr these newly specified germinal cells have patterned their surrounding neighbors to differentiate as the first somatic niche cells. Transcriptomes were analyzed by microarray hybridization in carefully staged whole anthers during initial specification events, after the separation of germinal and somatic lineages, during the subsequent rapid mitotic proliferation phase, and during final pre-meiotic germinal and somatic cell differentiation. Maize anthers exhibit a highly complex transcriptome constituting nearly three-quarters of annotated maize genes, and expression patterns are dynamic. Laser microdissection was applied to begin assigning transcripts to tissue and cell types and for comparison to transcriptomes of mutants defective in cell fate specification. Whole anther proteomes were analyzed at three developmental stages by mass spectrometric peptide sequencing using size-fractionated proteins to evaluate the timing of protein accumulation relative to transcript abundance. New insights include early and sustained expression of meiosis-associated genes (77.5% of well-annotated meiosis genes are constitutively active in 0.15 mm anthers), an extremely large change in transcript abundances and types a few days before meiosis (including a class of 1340 transcripts absent specifically at 0.4 mm), and the relative disparity between transcript abundance and protein abundance at any one developmental stage (based on 1303 protein-to-transcript comparisons).
archesporial cell; cell fate specification; Multiple archesporial cells 1; mac1; pre-meiotic development; genetics of sex
Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has recently been introduced in diagnostic microbiology laboratories for the identification of bacterial and yeast strains isolated from clinical samples. This study aimed to evaluate the accuracy of MALDI-TOF MS in clinical microbiology diagnosis by comparing it with commonly-used VITEK 2 or gene sequencing.
The performances of MALDI-TOF MS and VITEK 2 were compared retrospectively for identifying routine isolates. Discrepancies were analyzed by gene sequencing analysis of the 16S genes.
For 1,025 isolates, classified as 55 species of 25 genera, 1,021 (99.60%) isolates were accurately identified at the genus level, and 957 (93.37%) isolates at the species level by using MALDI-TOF MS. A total of 949 (92.59%) isolates were completely matched by both methods. Both methods found 76 unmatched isolates among which one strain had no definite identification by MALDI-TOF MS and VITEK 2 respectively. However, MALDI-TOF MS made no errors at the genus level while VITEK 2 made 6 (0.58%) errors at the genus level. At the species level, the identification error rates for MALDI-TOF MS and VITEK 2 were 5.56% and 6.24%, respectively.
With a lower identification error rate, MALDI-TOF MS has better performance than VITEK 2 in identifying bacteria found routinely in the clinical laboratory. It is a quick and cost-effective technique, and has the potential to replace conventional phenotype methods in identifying common bacterial isolates in clinical microbiology laboratories.
Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS); VITEK2; bacteria identification
This study investigated the development of tense markers (e.g., past tense –ed) in children with cochlear implants (CIs) over a 3-year span. Nine children who received CIs before 30 months of age participated in this study at three, four, and five years postimplantation. Nine typical 3-, 4-, and 5-year- olds served as control groups. All children participated in a story-retell task. Percent correct of tense marking in the task was computed. Within the groups, percent correct of tense marking changed significantly in children with CIs and in typical children who had more hearing experience. Across the groups, children with CIs were significantly less accurate in tense marking than typical children at four and five years postimplantation. In addition, the performance of tense marking in children with CIs was correlated with their speech perception skills at earlier time points. Errors of tense marking tended to be omission rather than commission errors in typical children as well as in children with CIs. The findings suggested that despite the perceptual and processing constraints, children who received CIs may learn tense marking albeit with a delayed pattern.
Intracavitary brachytherapy (ICBT) is usually applied as boost radiotherapy for superficial residual of nasopharyngeal carcinoma (NPC) after primary extern-beam radiptherapy (ERT). Here, we evaluated the outcome of endoscope-guided interstitial intensity-modulated brachytherapy (IMBT) boost radiation for deep-seated residual NPC.
Two hundred and thirteen patients with residual NPC who were salvaged with brachytherapy boost radiation during 2005–2009 were analyzed retrospectively. Among these patients, 171 patients had superficial residual NPC (≤1 cm below the nasopharyngeal epithelium) were treated with ICBT boost radiation, and interstitial IMBT boost radiation was delivered to 42 patients with deep-seated residual NPC (>1 cm below the nasopharyngeal epithelium). We found that IMBT boost subgroup had a higher ratio of T2b (81.0% VS 34.5%, P<0.001) and stage II (90.5% VS 61.4%, P = 0.001) than that of ICBT boost subgroup. The dosage of external-beam radiotherapy in the nasopharyngeal (63.0±3.8 VS 62.6±4.3 Gray (Gy), P = 0.67) and regional lymph nodes (55.8±5.0 VS 57.5±5.7 Gy, P = 0.11) was comparable in both groups. For brachytherapy, IMBT subgroup had a lower boost radiation dosage than ICBT subgroup (11.0±2.9 VS 14.8±3.2 Gy, P<0.01). Though the IMBT group had deeper residual tumors and received lower boost radiation dosages, both subgroups had the similar 5-year actuarial overall survival rate (IMBT VS ICBT group: 96.8% VS 93.6%, P = 0.87), progression-free survival rate (92.4% VS 86.5%, P = 0.41) and distant metastasis-free survival rate (94.9% VS 92.7%, P = 0.64). Moreover, IMBT boost radiation subgroup had a similar local (97.4% VS 94.4%, P = 0.57) and regional (95.0% VS 97.2%, P = 0.34) control to ICBT subgroup. The acute and late toxicities rates were comparable between the both subgroups.
IMBT boost radiation may be a promising therapeutic selection for deep-seated residual NPC.
NDM-1; Enterobacteriaceae; Providencia rettgeri; Raoultella ornithinolytica; bacteria; China; New Delhi metallo-β-lactamase-1–producing strains
This study provided a yearly record of consonant development for the initial 4 years of cochlear implant (CI) use and established a precedent for using a standardized articulation test, the Goldman–Fristoe Test of Articulation—2 (Goldman, R., & Fristoe, M. . Goldman–Fristoe Test of Articulation—2. Circle Pines, MN: American Guidance Services). The study used CI age as a referent for 32 children who received their CI before 30 months of age. Consonants produced by 70% of the children were listed, as were the most common error types, which were consonant omissions and substitutions. Using consonant repertoire lists and standard scores, the study revealed that children with CIs had acquisition patterns that were similar to their peers when the duration of CI experience was similar to the chronological age norms of typically developing children. The results revealed that CI users need time to coordinate their articulatory organizing principles with the input they receive from their CI. It is appropriate to use length of CI use as a proxy for chronological age during the first 4 years when comparing articulation development with hearing peers.
In the present study, serum samples from 650 goats were collected from five counties between May and June 2012 and antibodies to Toxoplasma gondii were detected by indirect haemagglutination assay; 58 (9%) had antibodies to T. gondii with antibody titres of 1:64 to 1:1024. Seropositive samples were distributed in all five counties: seroprevalences in Kuandian county (15%, 21/139, 95% confidence interval [CI] 9–21%) were statistically different from the four other counties (Gaizhou, Huanren, Xiuyan and Liaoyang), and the seroprevalence difference between Xiuyan county (12%, 15/127, 95% CI 6–17%) and two other counties (Huanren, Liaoyang) was significantly different (P < 0.05). To our knowledge, this is the first report of the seroprevalence of T. gondii exposure in Liaoning cashmere goat in China. Our results indicated that Liaoning cashmere goat could be a potential reservoir for the transmission of T. gondii in Liaoning Province.
Toxoplasma gondii; seroprevalence; Liaoning cashmere goat; indirect haemagglutination assay
Background and Purpose
Treatment outcomes vary greatly in patients with nasopharyngeal carcinoma (NPC). The purpose of this study is to evaluate the influence of radiation and chemotherapy drug action pathway gene polymorphisms on the survival of patients with locoregionally advanced NPC treated with cisplatin- and fluorouracil-based chemoradiotherapy.
Material and Methods
Four hundred twenty-one consecutive patients with locoregionally advanced NPC were prospectively recruited. We utilized a pathway approach and examined 18 polymorphisms in 13 major genes. Polymorphisms were detected using the LDR-PCR technique. Multifactor dimensionality reduction (MDR) analysis was performed to detect potential gene-gene interaction.
After adjustment for clinicopathological characteristics, overall survival was significantly decreased in patients with the MPO rs2243828 CT/CC genotype (HR=2.453, 95% CI, 1.687-3.566, P<0.001). The ERCC1 rs3212986 CC (HR=1.711, 95% CI, 1.135-2.579, P=0.010), MDM2 rs2279744 GT/GG (HR=1.743, 95% CI, 1.086-2.798, P=0.021), MPO rs2243828 CT/CC (HR=3.184, 95% CI, 2.261-4.483, P<0.001) and ABCB1 rs2032582 AT/AA (HR=1.997, 95% CI, 1.086-3.670, P=0.026) genotypes were associated with poor progression-free survival. Prognostic score models based on independent prognostic factors successfully classified patients into low-, intermediate-, and high-risk groups. Furthermore, MDR analysis showed no significant interaction between polymorphisms.
Four single nucleotide polymorphisms were associated with survival in patients with locoregionally advanced NPC treated with cisplatin- and fluorouracil-based chemoradiotherapy. Combining clinical prognostic factors with genetic information was valuable in identifying patients with different risk.
Mesenchymal stem cells (MSCs) have been considered as ideal cells for the treatment of a variety of diseases. However, aging and spontaneous differentiation of MSCs during culture expansion dampen their effectiveness. Previous studies suggest that ex
vivo aging of MSCs is largely caused by epigenetic changes particularly a decline of histone H3 acetylation levels in promoter regions of pluripotent genes due to inappropriate growth environment.
In this study, we examined whether histone deacetylase inhibitor trichostatin A (TSA) could suppress the histone H3 deacetylation thus maintaining the primitive property of MSCs. We found that in regular adherent culture, human MSCs became flatter and larger upon successive passaging, while the expression of pluripotent genes such as Oct4, Sox2, Nanog, Rex-1, CD133 and TERT decreased markedly. Administration of low concentrations of TSA in culture significantly suppressed the morphological changes in MSCs otherwise occurred during culture expansion, increased their proliferation while retaining their cell contact growth inhibition property and multipotent differentiation ability. Moreover, TSA stabilized the expression of the above pluripotent genes and histone H3 acetylation levels in K9 and K14 in promoter regions of Oct4, Sox2 and TERT.
Our results suggest that TSA may serve as an effective culture additive to maintain the primitive feature of MSCs during culture expansion.
Epoxyeicosatrienoic acids (EETs) confer vasoactive and cardioprotective functions. Genetic analysis of the contributions of these short-lived mediators to pathophysiology has been confounded to date by the allelic expansion in rodents of the portion of the genome syntenic to human CYP2J2, a gene encoding one of the principle cytochrome P450 epoxygenases responsible for the formation of EETs in humans. Mice have eight potentially functional genes that could direct the synthesis of epoxygenases with properties similar to those of CYP2J2. As an initial step towards understanding the role of the murine Cyp2j locus, we have created mice bearing a 626-kb deletion spanning the entire region syntenic to CYP2J2, using a combination of homologous and site-directed recombination strategies. A mouse strain in which the locus deletion was complemented by transgenic delivery of BAC sequences encoding human CYP2J2 was also created. Systemic and pulmonary hemodynamic measurements did not differ in wild-type, null, and complemented mice at baseline. However, hypoxic pulmonary vasoconstriction (HPV) during left mainstem bronchus occlusion was impaired and associated with reduced systemic oxygenation in null mice, but not in null mice bearing the human transgene. Administration of an epoxygenase inhibitor to wild-type mice also impaired HPV. These findings demonstrate that Cyp2j gene products regulate the pulmonary vascular response to hypoxia.
In mice and humans, the CYP2J class of cytochrome P450 epoxygenases metabolizes arachidonic acid (AA) to epoxyeicosatrienoic acids (EETs), short-lived mediators with effects on both the pulmonary and systemic vasculature. Genetic dissection of CYP2J function to date has been complicated by allelic expansion in the rodent genome. In this study, the mouse chromosomal locus syntenic to human CYP2J2, containing eight presumed genes and two pseudogenes, was deleted via generation of a recombinant template created by homologous and site-specific recombination steps that joined two precursor bacterial artificial chromosomes (BACs). The Cyp2j null mice were subsequently complemented by transgenic delivery of BAC sequences encoding human CYP2J2. Hypoxic pulmonary vasoconstriction (HPV) and systemic oxygenation during regional alveolar hypoxia were unexpectedly found to be impaired in null mice, but not in null mice bearing the transgenic human allele, suggesting that Cyp2j products contribute to the pulmonary vascular response to hypoxia.
Systemic chemotherapy is the basic palliative treatment for metastatic nasopharyngeal carcinoma (NPC); however, it is not known whether locoregional radiotherapy targeting the primary tumor and regional lymph nodes affects the survival of patients with metastatic NPC. Therefore, we aimed to retrospectively evaluate the benefits of locoregional radiotherapy. A total of 408 patients with metastatic NPC were included in this study. The mortality risks of the patients undergoing supportive treatment and those undergoing chemotherapy were compared with that of patients undergoing locoregional radiotherapy delivered alone or in combination with chemotherapy. Univariate and multivariate analyses were conducted. The contributions of independent factors were assessed after adjustment for covariates with significant prognostic associations (P < 0.05). Both locoregional radiotherapy and systemic chemotherapy were identified as significant independent prognostic factors of overall survival (OS). The mortality risk was similar in the group undergoing locoregional radiotherapy alone and the group undergoing systemic chemotherapy alone [multi-adjusted hazard ratio (HR) = 0.9, P = 0.529]; this risk was 60% lower than that of the group undergoing supportive treatment (HR = 0.4, P = 0.004) and 130% higher than that of the group undergoing both systemic chemotherapy and locoregional radiotherapy (HR = 2.3, P < 0.001). In conclusion, locoregional radiotherapy, particularly when combined with systemic chemotherapy, is associated with improved survival of patients with metastatic NPC.
Nasopharyngeal carcinoma; distant metastases; overall survival; radiotherapy; systemic chemotherapy
To investigate the characteristics and criterion of graft rejection in mice model.
C57BL/6 or BALB/c mice corneal grafts were grafted onto BALB/c hosts. Each group was divided into two subgroups according to the corneal opacity scores 12d after transplantation. The characteristics of opacity and neovascularization were observed. Mice of the 12th, 50th day after transplantation, the grafts biopsy of mice in allogeneic group 1, which opacity score exceed 3, were prepared for histological observation and those restore transparent were endothelial stained.
There was no difference of corneal opacity score on the 7th and 12th day after operation; the histological results had no disparity between syngeneic group and allogeneic group. On the 12th day after surgery, the turbidity curve was apparent in grafts with opacity score < 2. Mononuclear cells were shown in grafts with opacity score reached 3 in allogeneic group 1. Different rejection performance was observed in tissue sections on the 50th day after surgery.
Grafts, opacity score exceeds 3 from the 7th to the 12th day after operation could not be judged as a rejection. We should pay more attention to the variation of grafts opacity since 12d after corneal transplantation.
corneal transplantation; graft survival; experimental study
Rat middle cerebral artery occlusion (MCAO) model is the most commonly used animal model in ischemic stroke studies. In the model, to increase the amount of stem cells or drugs to enter the brain after delivery into the internal carotid artery (ICA), the pterygopalatine artery (PPA) is occluded. However, PPA occlusion is a technically demanding procedure which often causes complications.
In this study, we developed an ICA injection needle to facilitate easy and efficient delivery of stem cells to the ischemic brain through the ICA without the need of PPA occlusion. We injected methylene blue and fluorescence dye DiI-labeled human mesenchymal stem cells (DiI-hMSCs) into the ICA in rats with the ICA injection needle (without PPA ligation) or the conventional micro-injection needle (with PPA ligation) and assessed their distributions.
When methylene blue was injected, evident blue stains were found in the brain of the injection side particularly the middle cerebral artery (MCA)-supplied areas but not in the PPA supplied areas. Similarly, when DiI-hMSCs were injected, the cells largely appeared in the MCA-supplied tissues, which were similar in quantity compared to conventional micro-injection needle injection with PPA occlusion. Moreover, hMSCs injected with the ICA needle or the micro-injection needle similarly improved the functional recovery of the infarcted brain.
Our results indicate that the ICA injection needle is easy to use and efficient in delivering cells to the ischemic brain tissue in rat MCAO model.
Stroke; Acute cerebral infarction
The aim of this study was to explore the correlation of erythropoietin (EPO) with N-terminal pro-B-type natriuretic peptide (NT-proBNP) and high sensitivity C-reactive protein (hs-CRP) in patients with chronic heart failure (CHF) or CHF complicated with anemia, in addition to its correlation with the prognosis of the patient. A total of 217 CHF patients were enrolled in this study. The patients were graded according to the cardiac function criteria of the New York Heart Association (NYHA). The serum EPO, NT-proBNP and hs-CRP levels of the patients were determined. The patients were followed up for ≥24 months. The EPO expression level in patients with NYHA II–IV CHF was significantly higher compared with that in the control group (P<0.05). EPO expression increased with the aggravation of CHF, exhibiting significant differences amongst the various NYHA graded groups (P<0.05). The EPO expression level increased significantly with an increase in NHA grade in addition to the severity of the anemia in the patients with CHF complicated by anemia (P<0.05). In the patients who succumbed (mortality group), the expression level of EPO was significantly higher and the hemoglobin level was significantly lower compared with those of the survival group (P<0.05). The EPO expression levels were elevated in CHF patients and patients with CHF and anemia. The level of expression correlated positively with the severity of CHF as well as that of anemia. Serum EPO measurements were successful in predicting the mortality and re-hospitalization rates of CHF patients at the end point, within two years of follow-up.
heart failure; erythropoietin; N-terminal pro-B-type natriuretic peptide; high sensitivity C-reactive protein; prognosis
Crop productivity is greatly affected by soil salinity; therefore, improvement in salinity tolerance of crops is a major goal in salt-tolerant breeding. The Salt Overly Sensitive (SOS) signal-transduction pathway plays a key role in ion homeostasis and salt tolerance in plants. Here, we report that overexpression of Arabidopsis thaliana SOS1+SOS2+SOS3 genes enhanced salt tolerance in tall fescue. The transgenic plants displayed superior growth and accumulated less Na+ and more K+ in roots after 350 mM NaCl treatment. Moreover, Na+ enflux, K+ influx, and Ca2+ influx were higher in the transgenic plants than in the wild-type plants. The activities of the enzyme superoxide dismutase, peroxidase, catalase, and proline content in the transgenic plants were significantly increased; however, the malondialdehyde content decreased in transgenic plants compared to the controls. These results suggested that co-expression of A. thaliana SOS1+SOS2+SOS3 genes enhanced the salt tolerance in transgenic tall fescue.
Electronic supplementary material
The online version of this article (doi:10.1007/s00709-013-0540-9) contains supplementary material, which is available to authorized users.
SOS pathway genes; Salt stress; Transgenic tall fescue
AIM: To examine transforming growth factor-β1 (TGF-β1) promoter methylation in gastric cancer and to determine if Helicobacter pylori (H. pylori) or interleukin (IL)-1β could induce TGF-β1 hypermethylation in vitro.
METHODS: We examined the frequency and extent of TGF-β1 promoter methylation using methylation-specific PCR in the gastric tissues from 47 gastric cancer patients and 39 non-gastric cancer subjects. H. pylori infection was confirmed by a positive result from either a serological test, histological analysis or C13 urea breath test. GES-1 and MKN-45 cells co-cultured with H. pylori or treated with IL-1β for 12, 24 and 48 h in vitro tested the effects of H. pylori or IL-1β on TGF-β1.
RESULTS: Twenty-four/forty-seven (51%) cases of gastric cancer (GC) tissues showed TGF-β1 promoter methylation, 15/47 (31.9%) cases of matched non-cancerous gastric mucosa tissues from the GC patients, and 11/39 (28%) case of the normal gastric mucosa tissues from non-GC subjects showed TGF-β1 promoter methylation (51% vs 28%, P < 0.05). Significantly higher levels of methylation of TGF-β1 were found in the tumor tissues than in non-tumor tissues from GC patients (0.24 ± 0.06 vs 0.17 ± 0.04, P < 0.05) and normal gastric tissues from non-GC subjects (0.24 ± 0.06 vs 0.15 ± 0.03, P < 0.05). TGF-β1 methylation was found in 48.3% of H. pylori-positive gastric mucosal tissues whereas only 23.1% of H. pylori-negative gastric mucosal tissues showed TGF-β1 methylation (48.3% vs 23.1%, P < 0.05). IL-1β appeared to induce a dose-dependent methylation of TGF-β1 and the strongest methylation was observed in GES-1 cells treated with 2.5 ng/mL of IL-1β for 48 h. Further studies showed that pre-treatment of GES-1 cells with 20 ng/mL IL-1RA for 1 h could partially abolish the effect of IL-1β on TGF-β1 methylation. Infection of GES-1 cells by H. pylori was not found to induce significant TGF-β1 promoter methylation.
CONCLUSION: Our data revealed that TGF-β1 promoter is methylated in GC patients. IL-1β may be an important mediator for H. pylori induced gene methylation during GC development.
Transforming growth factor-β1; Interleukin-1β; Methylation; Helicobacter pylori; Gastric cancer
Neoadjuvant chemotherapy plus radiotherapy is the most common treatment regimen for advanced nasopharyngeal carcinoma (NPC). Whether chronomodulated infusion of chemotherapy can reduce its toxicity is unclear. This study aimed to evaluate the toxic and therapeutic effects of sinusoidal chronomodulated infusion versus flat intermittent infusion of cisplatin (DDP) and 5-fluorouracil (5-FU) followed by radiotherapy in patients with locoregionally advanced NPC. Patients with biopsy-diagnosed untreated stages III and IV NPC (according to the 2002 UICC staging system) were randomized to undergo 2 cycles of sinusoidal chronomodulated infusion (Arm A) or flat intermittent constant rate infusion (Arm B) of DDP and 5-FU followed by radical radiotherapy. Using a “MELODIE” multi-channel programmed pump, the patients were given 12-hour continuous infusions of DDP (20 mg/m2) and 5-FU (750 mg/m2) for 5 days, repeated every 3 weeks for 2 cycles. DDP was administered from 10:00 am to 10:00 pm, and 5-FU was administered from 10:00 pm to 10:00 am each day. Chronomodulated infusion was performed in Arm A, with the peak deliveries of 5-FU at 4:00 am and DDP at 4:00 pm. The patients in Arm B underwent a constant rate of infusion. Radiotherapy was initiated in the fifth week, and both arms were treated with the same radiotherapy techniques and dose fractions. Between June 2004 and June 2006, 125 patients were registered, and 124 were eligible for analysis of response and toxicity. The major toxicity observed during neoadjuvant chemotherapy was neutropenia. The incidence of acute toxicity was similar in both arms. During radiotherapy, the incidence of stomatitis was significantly lower in Arm A than in Arm B (38.1% vs. 59.0%, P = 0.020). No significant differences were observed for other toxicities. The 1-, 3-, and 5-year overall survival rates were 88.9%, 82.4%, and 74.8% for Arm A and 91.8%, 90.2%, and 82.1% for Arm B. The 1-, 3-, and 5-year progression-free survival rates were 91.7%, 88.1%, and 85.2% for Arm A and 100%, 94.5%, and 86.9% for Arm B. The 1-, 3-, and 5-year distant metastasis-free survival rates were 82.5%, 79.1%, and 79.1% for Arm A and 90.2%, 85.2%, and 81.7% for Arm B. Chronochemotherapy significantly reduced stomatitis but was not superior to standard chemotherapy in terms of hematologic toxicities and therapeutic response.
Chronochemotherapy; cisplatin; 5-fluorouracil; nasopharyngeal carcinoma; radiotherapy
Flotillin-2 (FLOT2) has been implicated in several signaling pathways in tumor cells. Our study aimed to investigate the expression pattern and clinicopathological significance of FLOT2 in patients with breast cancer.
The expression level of FLOT2 in normal breast epithelial cells, breast cancer cell lines, and four breast cancer biopsies paired with adjacent noncancerous tissues were quantified using real-time RT-PCR and Western blotting. FLOT2 protein expression was analyzed in 171 archived paraffin-embedded breast cancer samples using immunohistochemistry (IHC). Statistical analyses were performed to evaluate the clinicopathological significance of FLOT2 expression.
FLOT2 was significantly upregulated in breast cancer cell lines and tissue samples compared with normal cells and adjacent noncancerous breast tissues, respectively. IHC analysis revealed high expression levels of FLOT2 in 82 of 171 (48.0%) breast cancer specimens. Statistical analysis revealed that FLOT2 expression was significantly correlated with clinical stage (P < 0.001), T classification (P < 0.001), M classification (P < 0.001), histological differentiation (P = 0.005) and ErbB2 expression (P = 0.003). Patients with higher levels of FLOT2 expression had a shorter overall survival duration than patients with lower FLOT2 expression levels. Multivariate analysis suggested that FLOT2 expression was an independent prognostic marker for survival in patients with breast cancer.
The current results demonstrated that high FLOT2 protein expression was associated with poor outcomes in patients with breast cancer. FLOT2 could be used as a prognostic biomarker for breast cancer progression.
FLOT2; Breast cancer; Prognosis; Biomarker
In order to obtain a better molecular understanding of recurrent urinary tract infection (RUTI), we collected 75 cases with repeatedly occurring uncomplicated UTI. The genetic relationships among uropathogenic Escherichia coli (UPEC) isolates were analyzed by pulsed-field gel electrophoresis. While 39 (52%) of the RUTI cases were defined as “persistence” of the same strain as the primary infecting strain, 36 (48%) were characterized by “reinfection” with a new strain that is different from the primary strain. We then examined the antimicrobial susceptibilities and phylogenetic backgrounds of 39 persistence and 86 reinfection UPEC isolates, and screened 44 virulence factor (VF) genes. We found that isolates had significant differences in the following: placement in phylogenetic group B2 (41% versus 21%; P = 0.0193) and the presence of adhesin genes iha (49% versus 28%; P = 0.0233) and papG allele I′ (51% versus 24%; P = 0.003), iron uptake genes fyuA (85% versus 58%; P = 0.0037), irp-2 (87% versus 65%; P = 0.0109), and iutA (87% versus 58%; P = 0.0014), and an aggregate VF score (median, 11 versus 9; P = 0.0030). In addition, 41% of persistence strains harbored three adhesin genes simultaneously, whereas 22% of reinfection isolates did (P = 0.0289). Moreover, 59% versus 29% (P = 0.0014) of persistence and reinfection isolates contained seven types of iron uptake genes. Taken together, the antimicrobial susceptibilities of UPEC isolates had little effect on the RUTI. Compared with reinfection strains, persistence UPEC isolates exhibited higher VF scores and carried more VF genes than may be involved in the development and progression of RUTI.
In this study, we sequenced and phylogenetic analyses of the VP2 genes from twelve canine parvovirus (CPV) strains obtained from eleven domestic dogs and a giant panda (Ailuropoda melanoleuca) in China. A novel canine parvovirus (CPV) was detected from the giant panda in China.
Nucleotide and phylogenetic analysis of the capsid protein VP2 gene classified the CPV as a new CPV-2a type. Substitution of Gln for Arg at the conserved 370 residue in CPV presents an unusual variation in the new CPV-2a amino acid sequence of the giant panda and is further evidence for the continuing evolution of the virus.
These findings extend the knowledge on CPV molecular epidemiology of particular relevance to wild carnivores.
Canine parvovirus; VP2 gene; Point mutation; Phylogenetic analysis; Giant panda
Catalase (CAT) breaks down H2O2 into H2O and O2 to protects cells from oxidative damage. However, its translational potential is limited because exogenous CAT cannot enter living cells automatically. This study is aimed to investigate if PEP-1-CAT fusion protein can effectively protect cardiomyocytes from oxidative stress due to hypoxia/reoxygenation (H/R)-induced injury.
H9c2 cardomyocytes were pretreated with catalase (CAT) or PEP-1-CAT fusion protein followed by culturing in a hypoxia and re-oxygenation condition. Cell apoptosis were measured by Annexin V and PI double staining and Flow cytometry. Intracellular superoxide anion level was determined, and mitochondrial membrane potential was measured. Expression of apoptosis-related proteins including Bcl-2, Bax, Caspase-3, PARP, p38 and phospho-p38 was analyzed by western blotting.
PEP-1-CAT protected H9c2 from H/R-induced morphological alteration and reduced the release of lactate dehydrogenase (LDH) and malondialdehyde content. Superoxide anion production was also decreased. In addition, PEP-1-CAT inhibited H9c2 apoptosis and blocked the expression of apoptosis stimulator Bax while increased the expression of Bcl-2, leading to an increased mitochondrial membrane potential. Mechanistically, PEP-1-CAT inhibited p38 MAPK while activating PI3K/Akt and Erk1/2 signaling pathways, resulting in blockade of Bcl2/Bax/mitochondrial apoptotic pathway.
Our study has revealed a novel mechanism by which PEP-1-CAT protects cardiomyocyte from H/R-induced injury. PEP-1-CAT blocks Bcl2/Bax/mitochondrial apoptotic pathway by inhibiting p38 MAPK while activating PI3K/Akt and Erk1/2 signaling pathways.
Cell-penetrating peptide; PEP-1; Catalase; Cardiomyocyte; Apoptosis; MAPK
Caveolae are cholesterol and sphingolipids rich subcellular domains on plasma membrane. Caveolae contain a variety of signaling proteins which provide platforms for signaling transduction. In addition to enriched with cholesterol and sphingolipids, caveolae also contain a variety of fatty acids. It has been well-established that acylation of protein plays a pivotal role in subcellular location including targeting to caveolae. However, the fatty acid compositions of caveolae and the type of acylation of caveolar proteins remain largely unknown. In this study, we investigated the fatty acids in caveolae and caveolin-1 bound fatty acids.
Caveolae were isolated from Chinese hamster ovary (CHO) cells. The caveolar fatty acids were extracted with Folch reagent, methyl esterificated with BF3, and analyzed by gas chromatograph-mass spectrometer (GC/MS). The caveolin-1bound fatty acids were immunoprecipitated by anti-caveolin-1 IgG and analyzed with GC/MS.
In contrast to the whole CHO cell lysate which contained a variety of fatty acids, caveolae mainly contained three types of fatty acids, 0.48 µg palmitic acid, 0.61 µg stearic acid and 0.83 µg oleic acid/caveolae preparation/5×107 cells. Unexpectedly, GC/MS analysis indicated that caveolin-1 was not acylated by myristic acid; instead, it was acylated by palmitic acid and stearic acid.
Caveolae contained a special set of fatty acids, highly enriched with saturated fatty acids, and caveolin-1 was acylated by palmitic acid and stearic acid. The unique fatty acid compositions of caveolae and acylation of caveolin-1 may be important for caveolae formation and for maintaining the function of caveolae.
Canine distemper virus (CDV) infects a variety of carnivores, including wild and domestic Canidae. In this study, we sequenced and phylogenetic analyses of the hemagglutinin (H) genes from eight canine distemper virus (CDV) isolates obtained from seven raccoon dogs (Nyctereutes procyonoides) and a giant panda (Ailuropoda melanoleuca) in China.
Phylogenetic analysis of the partial hemagglutinin gene sequences showed close clustering for geographic lineages, clearly distinct from vaccine strains and other wild-type foreign CDV strains, all the CDV strains were characterized as Asia-1 genotype and were highly similar to each other (91.5-99.8% nt and 94.4-99.8% aa). The giant panda and raccoon dogs all were 549Y on the HA protein in this study, irrespective of the host species.
These findings enhance our knowledge of the genetic characteristics of Chinese CDV isolates, and may facilitate the development of effective strategies for monitoring and controlling CDV for wild canids and non-cainds in China.
Canine distemper virus; Haemagglutinin (H) gene; Genotype; Phylogenetic analysis