Germination is a biological process important to plant development and agricultural production. Barley and rice diverged 50 million years ago, but share a similar germination process. To gain insight into the conservation of their underlying gene regulatory programs, we compared transcriptomes of barley and rice at start, middle and end points of germination, and revealed that germination regulated barley and rice genes (BRs) diverged significantly in expression patterns and/or protein sequences. However, BRs with higher protein sequence similarity tended to have more conserved expression patterns. We identified and characterized 316 sets of conserved barley and rice genes (cBRs) with high similarity in both protein sequences and expression patterns, and provided a comprehensive depiction of the transcriptional regulatory program conserved in barley and rice germination at gene, pathway and systems levels. The cBRs encoded proteins involved in a variety of biological pathways and had a wide range of expression patterns. The cBRs encoding key regulatory components in signaling pathways often had diverse expression patterns. Early germination up-regulation of cell wall metabolic pathway and peroxidases, and late germination up-regulation of chromatin structure and remodeling pathways were conserved in both barley and rice. Protein sequence and expression pattern of a gene change quickly if it is not subjected to a functional constraint. Preserving germination-regulated expression patterns and protein sequences of those cBRs for 50 million years strongly suggests that the cBRs are functionally significant and equivalent in germination, and contribute to the ancient characteristics of germination preserved in barley and rice. The functional significance and equivalence of the cBR genes predicted here can serve as a foundation to further characterize their biological functions and facilitate bridging rice and barley germination research with greater confidence.
We describe the development and validation of the PROMIS Sexual Function and Satisfaction (PROMIS SexFS) measures version 1.0 for cancer populations.
To develop a customizable self-report measure of sexual function and satisfaction as part of the U.S. National Institutes of Health PROMIS® Network.
Our multidisciplinary working group followed a comprehensive protocol for developing psychometrically robust patient reported outcome (PRO) measures including qualitative (scale development) and quantitative (psychometric evaluation) development. We performed an extensive literature review, conducted 16 focus groups with cancer patients and multiple discussions with clinicians, and evaluated candidate items in cognitive testing with patients. We administered items to 819 cancer patients. Items were calibrated using item response theory and evaluated for reliability and validity.
Main Outcome Measures
The PROMIS Sexual Function and Satisfaction (PROMIS SexFS) measures version 1.0 include 79 items in 11 domains: interest in sexual activity, lubrication, vaginal discomfort, erectile function, global satisfaction with sex life, orgasm, anal discomfort, therapeutic aids, sexual activities, interfering factors, and screener questions.
In addition to content validity (patients indicate that items cover important aspects of their experiences) and face validity (patients indicate that items measure sexual function and satisfaction), the measure shows evidence for discriminant validity (domains discriminate between groups expected to be different), convergent validity (strong correlations between scores on PROMIS and scores on conceptually-similar older measures of sexual function), as well as favorable test-retest reliability among people not expected to change (inter-class correlations from 2 administrations of the instrument, 1 month apart).
The PROMIS SexFS offers researchers a reliable and valid set of tools to measure self-reported sexual function and satisfaction among diverse men and women. The measures are customizable; researchers can select the relevant domains and items comprising those domains for their study.
patient-reported outcome measures; sexual function; satisfaction; cancer; quality of life; male and female sexual dysfunction
Tacrolimus is a widely used immunosuppressive drug for preventing the rejection of solid organ transplants. The efficacy of tacrolimus shows considerable variability, which might be related to genetic variation among recipients. We conducted a retrospective study of 240 Chinese renal transplant recipients receiving tacrolimus as immunosuppressive drug. The retrospective data of all patients were collected for 40 days after transplantation. Seventeen SNPs of CYP3A5, CYP3A4, COMT, IL-10 and POR were identified by the SNaPshot assay. Tacrolimus blood concentrations were obtained on days 1–3, days 6–8 and days 12–14 after transplantation, as well as during the period of the predefined therapeutic concentration range. Kruskal–Wallis test was used to examine the effect of genetic variation on the tacrolimus concentration/dose ratio (C0/D) at different time points. Chi-square test was used to compare the proportions of patients who achieved the target C0 range in the different genotypic groups at weeks 1, 2, 3 and 4 after transplantation. After correction for multiple testing, there was a significant association of C0/D with CYP3A5*3, CYP3A4*1G and CYP3A4 rs4646437 T>C at different time points after transplantation. The proportion of patients in the IL-10 rs1800871-TT group who achieved the target C0 range was greater (p = 0.004) compared to the IL-10 rs1800871-CT and IL-10 rs1800871-CC groups at week 3 after transplantation. CYP3A5*3, CYP3A4 *1G, CYP3A4 rs4646437 T>C and IL-10 rs1800871 C>T might be potential polymorphisms affecting the interindividual variability in tacrolimus metabolism among Chinese renal transplant recipients.
The structure-activity relationship study of a diphenylpropanamide series of RORγ selective modulators is reported. Compounds were screened using chimeric receptor Gal4 DNA-binding domain (DBD)-NR ligand binding domain cotransfection assay in a two-step format. Three different regions of the scaffold were modified to assess the effects on repression of RORγ transcriptional activity and potency. The lead compound 1 exhibits modest mouse pharmacokinetics and an acceptable in vitro profile which makes it a suitable in vivo probe to interrogate the functions of RORγ in animal models of disease.
Despite advances in radiation therapy, chemotherapy, and newly developed molecular targeting therapies, long-term survival after resection for patients with NSCLC remains less than 50%. We investigated factors predicting postoperative locoregional recurrences and distant metastases in patients with clinical stage I non-small-cell lung cancer (NSCLC) after surgical resection.
All patients with clinical stage I NSCLC, who underwent surgical resection between January 2002 and June 2006, were reviewed retrospectively. Multiple logistic regression analyses were used to identify independent risk factors for patients with locoregional recurrences and distant metastases.
A total of 261 patients were eligible. Overall survival was significant related to locoregional recurrences (P = 0.03) and distant metastases (P <0.001). There were significant differences of locoregional recurrence in tumor differentiation (P = 0.032) and advanced pathological stage (P = 0.002). In the group of distant metastases, there were significant differences in tumor differentiation (P = 0.035), lymphovascular space invasion (P = 0.031). Among the relationship between pattern of distant metastasis and clinicopathologic variables in patients with clinical stage I NSCLC, SUVmax (P = 0.02) and tumor size (P = 0.001) had significant differences. According to multiple logistic regression analysis, tumor differentiation is the only risk factor of postoperative outcome for locoregional recurrence and serum CEA (>3.5 ng/mL) is the predictor of distant metastasis.
Tumor differentiation and serum CEA were predictors of postoperative relapse for clinical stage I NSCLC after surgical resection. Risk factors of postoperative recurrence in patients with clinical stage I NSCLC may enable us to optimize the patient selection for postoperative adjuvant therapies or neoadjuvant treatment before surgery.
Non-small cell lung cancer; Locoregional recurrence; Distant metastasis; Carcinoembryonic antigen
XB130 has been reported to be expressed by various types of cells such as thyroid cancer and esophageal cancer cells, and it promotes the proliferation and invasion of thyroid cancer cells. Our previous study demonstrated that XB130 is also expressed in gastric cancer (GC), and that its expression is associated with the prognosis, but the role of XB130 in GC has not been well characterized.
In this study, we investigated the influence of XB130 on gastric tumorigenesis and metastasis in vivo and in vitro using the MTT assay, clonogenic assay, BrdU incorporation assay, 3D culture, immunohistochemistry and immunofluorescence. Western blot analysis was also performed to identify the potential mechanisms involved.
The proliferation, migration, and invasion of SGC7901 and MNK45 gastric adenocarcinoma cell lines were all significantly inhibited by knockdown of XB130 using small hairpin RNA. In a xenograft model, tumor growth was markedly inhibited after shXB130-transfected GC cells were implanted into nude mice. After XB130 knockdown, GC cells showed a more epithelial-like phenotype, suggesting an inhibition of the epithelial-mesenchymal transition (EMT) process. In addition, silencing of XB130 reduced the expression of p-Akt/Akt, upregulated expression of epithelial markers including E-cadherin, α-catenin and β-catenin, and downregulated mesenchymal markers including fibronectin and vimentin. Expression of oncoproteins related to tumor metastasis, such as MMP2, MMP9, and CD44, was also significantly reduced.
These findings indicate that XB130 enhances cell motility and invasiveness by modulating the EMT-like process, while silencing XB130 in GC suppresses tumorigenesis and metastasis, suggesting that it may be a potential therapeutic target.
Gastric cancer; Adaptor protein; Oncogene; Epithelial-mesenchymal transition-like
Retrospective studies have demonstrated that nearly 50% of patients with ovarian cancer with normal cancer antigen 125 (CA125) levels have persistent disease; however, prospectively distinguishing between patients is currently impossible. Here, we demonstrate that for one patient, with the first reported fibroblast growth factor receptor 2 (FGFR2) fusion transcript in ovarian cancer, circulating tumor DNA (ctDNA) is a more sensitive and specific biomarker than CA125, and it can also inform on a candidate therapeutic. For a 4-year period, during which the patient underwent primary debulking surgery and chemotherapy, tumor recurrences, and multiple chemotherapeutic regimens, blood samples were longitudinally collected and stored. Whereas postsurgical CA125 levels were elevated only three times for 28 measurements, the FGFR2 fusion ctDNA biomarker was readily detectable by quantitative real-time reverse transcription-polymerase chain reaction (PCR) in all of these same blood samples and in the tumor recurrences. Given the persistence of the FGFR2 fusion, we treated tumor cells derived from this patient and others with the FGFR2 inhibitor BGJ398. Only tumor cells derived from this patient were sensitive to FGFR2 inhibitor treatment. Using the same methodologic approach, we demonstrate in a second patient with a different fusion that PCR and agarose gel electrophoresis can also be used to identify tumor-specific DNA in the circulation. Taken together, we demonstrate that a relatively inexpensive, PCR-based ctDNA surveillance assay can outperform CA125 in identifying occult disease.
This research was designed to investigate the effects of low pressure radio-frequency (RF) oxygen plasma treatment (OPT) on the surface of commercially pure titanium (CP-Ti) and Ti6Al4V. Surface topography, elemental composition, water contact angle, cell viability, and cell morphology were surveyed to evaluate the biocompatibility of titanium samples with different lengths of OP treating time.
Materials and Methods
CP-Ti and Ti6Al4V discs were both classified into 4 groups: untreated, treated with OP generated by using oxygen (99.98%) for 5, 10, and 30 min, respectively. After OPT on CP-Ti and Ti6Al4V samples, scanning probe microscopy, X-ray photoelectron spectrometry (XPS), and contact angle tests were conducted to determine the surface topography, elemental composition and hydrophilicity, respectively. The change of surface morphology was further studied using sputtered titanium on silicon wafers. 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay and F-actin immunofluorescence stain were performed to investigate the viability and spreading behavior of cultivated MG-63 cells on the samples.
The surface roughness was most prominent after 5 min OPT in both CP-Ti and Ti6Al4V, and the surface morphology of sputtered Ti sharpened after the 5 min treatment. From the XPS results, the intensity of Ti°, Ti2+, and Ti3+ of the samples’ surface decreased indicating the oxidation of titanium after OPT. The water contact angles of both CP-Ti and Ti6Al4V were increased after 5 min OPT. The results of MTT assay demonstrated MG-63 cells proliferated best on the 5 min OP treated titanium sample. The F-actin immunofluorescence stain revealed the cultivated cell number of 5 min treated CP-Ti/Ti6Al4V was greater than other groups and most of the cultivated cells were spindle-shaped.
Low pressure RF oxygen plasma modified both the composition and the morphology of titanium samples’ surface. The CP-Ti/Ti6Al4V treated with 5 min OPT displayed the roughest surface, sharpest surface profile and best biocompatibility.
Protein functions are largely affected by their conformations. This is exemplified in proteins containing modular domains. However, the evolutionary dynamics that define and adapt the conformation of such modular proteins remain elusive. Here we show that cis-interactions between the C-terminal phosphotyrosines and SH2 domain within the protein tyrosine phosphatase Shp2 can be tuned by an adaptor protein, Grb2. The competitiveness of two phosphotyrosines, namely pY542 and pY580, for cis-interaction with the same SH2 domain is governed by an antagonistic combination of contextual amino acid sequence and position of the phosphotyrosines. Specifically, pY580 with the combination of a favorable position and an adverse sequence has an overall advantage over pY542. Swapping the sequences of pY542 and pY580 results in one dominant form of cis-interaction and subsequently inhibits the trans-regulation by Grb2. Thus, the antagonistic combination of sequence and position may serve as a basic design principle for proteins with tunable conformations.
STAT3 activation is frequently detected in breast cancer and this pathway has emerged as an attractive molecular target for cancer treatment. Recent experimental evidence suggests ALDH-positive (ALDH+), or cell surface molecule CD44-positive (CD44+) but CD24-negative (CD24−) breast cancer cells have cancer stem cell properties. However, the role of STAT3 signaling in ALDH+ and ALDH+/CD44+/CD24− subpopulations of breast cancer cells is unknown.
Methods and Results
We examined STAT3 activation in ALDH+ and ALDH+/CD44+/CD24− subpopulations of breast cancer cells by sorting with flow cytometer. We observed ALDH-positive (ALDH+) cells expressed higher levels of phosphorylated STAT3 compared to ALDH-negative (ALDH−) cells. There was a significant correlation between the nuclear staining of phosphorylated STAT3 and the expression of ALDH1 in breast cancer tissues. These results suggest that STAT3 is activated in ALDH+ subpopulations of breast cancer cells. STAT3 inhibitors Stattic and LLL12 inhibited STAT3 phosphorylation, reduced the ALDH+ subpopulation, inhibited breast cancer stem-like cell viability, and retarded tumorisphere-forming capacity in vitro. Similar inhibition of STAT3 phosphorylation, and breast cancer stem cell viability were observed using STAT3 ShRNA. In addition, LLL12 inhibited STAT3 downstream target gene expression and induced apoptosis in ALDH+ subpopulations of breast cancer cells. Furthermore, LLL12 inhibited STAT3 phosphorylation and tumor cell proliferation, induced apoptosis, and suppressed tumor growth in xenograft and mammary fat pad mouse models from ALDH+ breast cancer cells. Similar in vitro and tumor growth in vivo results were obtained when ALDH+ cells were further selected for the stem cell markers CD44+ and CD24−.
These studies demonstrate an important role for STAT3 signaling in ALDH+ and ALDH+/CD44+/CD24− subpopulations of breast cancer cells which may have cancer stem cell properties and suggest that pharmacologic inhibition of STAT3 represents an effective strategy to selectively target the cancer stem cell-like subpopulation.
5-Hydroxymethylcytosine (5-hmC) is a newly discovered modified form of cytosine that has been suspected to be an important epigenetic modification in neurodevelopment. While DNA methylation dynamics have already been implicated during neurodevelopment, little is known about hydroxymethylation in this process. Here, we report DNA hydroxymethylation dynamics during cerebellum development in the human brain. Overall, we find a positive correlation between 5-hmC levels and cerebellum development. Genome-wide profiling reveals that 5-hmC is highly enriched on specific gene regions including exons and especially the untranslated regions (UTRs), but it is depleted on introns and intergenic regions. Furthermore, we have identified fetus-specific and adult-specific differentially hydroxymethylated regions (DhMRs), most of which overlap with genes and CpG island shores. Surprisingly, during development, DhMRs are highly enriched in genes encoding mRNAs that can be regulated by fragile X mental retardation protein (FMRP), some of which are disrupted in autism, as well as in many known autism genes. Our results suggest that 5-hmC-mediated epigenetic regulation may broadly impact the development of the human brain, and its dysregulation could contribute to the molecular pathogenesis of neurodevelopmental disorders.
Accession number: Sequencing data have been deposited to GEO with accession number GSE40539.
MiRNAs are small, noncoding RNA molecules that act as posttranscriptional regulators of gene expression and function as important regulators in cancer-related processes. The miR-19a is overexpressed in various cancers and has been causally related to cellular proliferation and growth. To determine whether miR-19a plays a role in laryngeal squamous cell carcinoma (LSCC), we used quantitative real time PCR to detect miR-19a expression in LSCC tissues. We found that miR-19a is overexpressed in LSCC and correlated with neck nodal metastasis, poor differentiation and advanced stage. Statistical analysis suggests that higher level of miR-19a was associated with reduced overall survival. In vitro functional study showed that inhibition of miR-19a by antisense oligonucleotides (ASO) led to apoptosis and reduction of cell proliferation in LSCC cells. Furthermore, growth of LSCC xenograft tumors was significantly suppressed by repeated injection of ASO-miR-19a lentivirus. The TUNEL stain and transmission electron microscopy also detected increased apoptotic cells in ASO-miR-19a treated LSCC xenografts. In addition, both realtime PCR and western blot showed ASO-miR-19a can upregulate TIMP-2 expression and this suggests miR-19a is related with TIMP-2 pathway in LSCC cells. Taken together, these data suggest that miR-19a plays an oncogenic role in the progression of LSCC, and may serve as a biomarker or therapeutic target for patients with LSCC.
miR-19a; LSCC; TIMP-2; apoptosis
To determine whether patients' expectations of benefit in early-phase oncology trials depend on how patients are queried and to explore whether expectations are associated with patient characteristics.
Patients and Methods
Participants were 171 patients in phase I or II oncology trials in the United States. After providing informed consent for a trial but before receiving the investigational therapy, participants answered questions about expectations of benefit. We randomly assigned participants to one of three groups corresponding to three queries about expectations: frequency type, belief type, or both. Main outcomes were differences in expectations by question type and the extent to which expectations were associated with demographic characteristics, numeracy, dispositional optimism, religiousness/spirituality, understanding of research, and other measures.
The belief-type group had a higher mean expectation of benefit (64.4 of 100) than the combination group (51.6; P = .01) and the frequency-type group (43.1; P < .001). Mean expectations in the combination and frequency groups were not significantly different (P = .06). Belief-type expectations were associated with a preference for nonquantitative information (r = −0.19; 95% CI, −0.19 to −0.36), knowledge about research (r = −0.21; 95% CI, −0.38 to −0.03), dispositional optimism (r = 0.20; 95% CI, 0.01 to 0.37), and spirituality (r = 0.22; 95% CI, 0.03 to 0.38). Frequency-type expectations were associated with knowledge about clinical research (r = −0.27; 95% CI, −0.27 to −0.51).
In early-phase oncology trials, patients' reported expectations of benefit differed according to how patients were queried and were associated with patient characteristics. These findings have implications for how informed consent is obtained and assessed.
Prehypertension has been associated with adverse cerebrovascular events and brain damage. The aims of this study were to investigate i) whether short- and long-term treatments with losartan or amlodipine for prehypertension were able to prevent blood pressure (BP)-linked brain damage, and ii) whether there is a difference in the effectiveness of treatment with losartan and amlodipine in protecting BP-linked brain damage. In the present study, prehypertensive treatment with losartan and amlodipine (6 and 16 weeks treatment with each drug) was performed on 4-week-old stroke-prone spontaneously hypertensive rats (SHRSP). The results showed that long-term (16 weeks) treatment with losartan is the most effective in lowering systolic blood pressure in the long term (up to 40 weeks follow-up). Additionally, compared with the amlodipine treatment groups, the short- and long-term losartan treatments protected SHRSP from stroke and improved their brains structurally and functionally more effectively, with the long-term treatment having more benefits. Mechanistically, the short- and long-term treatments with losartan reduced the activity of the local renin-angiotensin-aldosterone system (RAAS) in a time-dependent manner and more effectively than their respective counterpart amlodipine treatment group mainly by decreasing AT1R levels and increasing AT2R levels in the cerebral cortex. By contrast, the amlodipine treatment groups inhibited brain cell apoptosis more effectively as compared with the losartan treatment groups mainly through the suppression of local oxidative stress. Taken together, the results suggest that long-term losartan treatment for prehypertension effectively protects SHRSP from stroke-induced brain damage, and this protection is associated with reduced local RAAS activity than with brain cell apoptosis. Thus, the AT1R receptor blocker losartan is a good candidate drug that may be used in the clinic for long-term treatment on prehypertensive populations in order to prevent BP-linked brain damage.
prehypertension; losartan; amlodipine; stroke-prone spontaneously hypertensive rats
Dysfunctional mitochondria participate in the progression of chronic kidney disease (CKD). Pirfenidone is a newly identified anti-fibrotic drug. However, its mechanism remains unclear. Mitochondrial dysfunction is an early event that occurs prior to the onset of renal fibrosis. In this context, we investigated the protective effect of pirfenidone on mitochondria and its relevance to apoptosis and oxidative stress in renal proximal tubular cells. A remnant kidney rat model was established. Human renal proximal tubular epithelial cells (HK2) using rotenone, a mitochondrial respiratory chain complex Ι inhibitor were further investigated in vitro to examine the mitochondrial protective effect of pirfenidone. Pirfenidone protected mitochondrial structures and functions by stabilizing the mitochondrial membrane potential, maintaining ATP production and improving the mitochondrial DNA (mtDNA) copy number. Pirfenidone decreased tubular cell apoptosis by inhibiting the mitochondrial apoptotic signaling pathway. Pirfenidone also reduced oxidative stress by enhancing manganese superoxide dismutase (Mn-SOD) and inhibiting intracellular reactive oxygen species (ROS) generation, which suggested that the anti-oxidant effects occurred at least partially via the mitochondrial pathway. Pirfenidone may be effective prior to the onset of renal fibrosis because this drug exerts its anti-fibrotic effect by protection of mitochondria in renal proximal tubular cells.
Streptococcal toxic shock syndrome (STSS) is an uncommon but life-threatening disease caused by Streptococcus pyogenes.
To understand the clinical and molecular characteristics of STSS, we analyzed clinical data and explored the emm types, superantigen genes, and pulsed-field gel electrophoresis of causative S. pyogenes isolates obtained between 2005 and 2012.
In total, 53 patients with STSS were included in this study. The median age of the patients was 57 years (range: 9–83 years), and 81.1% were male. The most prevalent underlying disease was diabetes mellitus (45.3%). Skin and soft-tissue infection accounted for 86.8% of STSS. The overall mortality rate was 32.1%. Underlying diseases had no statistical impact on mortality. A total of 19 different emm types were identified. The most prevalent emm type was emm102 (18.9%), followed by emm11 (17%), emm1 (11.3%), emm87 (9.4%), and emm89 (7.5%). There was no statistically significant association between emm type and a fatal outcome. Among the superantigen genes, speB was the most frequently detected one (92.5%), followed by smeZ (90.6%), speG (81.1%), speC (39.6%), and speF (39.6%). The majority of emm102 strains were found to have speB, speC, speG, and smeZ. The presence of speG was negatively associated with a fatal outcome (P = 0.045).
Our surveillance revealed the emergence of uncommon emm types, particularly emm102, causing STSS in southern Taiwan. Characterization of clinical, epidemiological, and molecular characteristics of STSS will improve our understanding of this life-threatening disease.
Heterogeneity within pluripotent stem cell (PSC) populations is indicative of dynamic changes that occur when cells drift between different states. Although the role of metastability in PSCs is unclear, it appears to reflect heterogeneity in cell signaling. Using the Fucci cell-cycle indicator system, we show that elevated expression of developmental regulators in G1 is a major determinant of heterogeneity in human embryonic stem cells. Although signaling pathways remain active throughout the cell cycle, their contribution to heterogeneous gene expression is restricted to G1. Surprisingly, we identify dramatic changes in the levels of global 5-hydroxymethylcytosine, an unanticipated source of epigenetic heterogeneity that is tightly linked to cell-cycle progression and the expression of developmental regulators. When we evaluated gene expression in differentiating cells, we found that cell-cycle regulation of developmental regulators was maintained during lineage specification. Cell-cycle regulation of developmentally regulated transcription factors is therefore an inherent feature of the mechanisms underpinning differentiation.
•Embryonic stem cells are lineage primed in G1•Transcription of developmentally regulated genes is cell-cycle regulated•5hmC is cell-cycle regulated•Stem cells initiate differentiation from G1
Pluripotent stem cell heterogeneity has been attributed to stochastic variations in signaling pathways across the population. Using Fucci cell-cycle reporters, Dalton and colleagues show that stem cell “lineage priming” in G1 is associated with cell-cycle-dependent changes in the transcription of developmentally regulated genes. Moreover, these changes are paralleled by levels of the epigenetic mark 5-hydroxymethylcytosine. These findings identify the cell cycle as major source of heterogeneity in human pluripotent stem cells.
Terrestrial mud volcanism represents the prominent surface geological feature, where fluids and hydrocarbons are discharged along deeply rooted structures in tectonically active regimes. Terrestrial mud volcanoes (MVs) directly emit the major gas phase, methane, into the atmosphere, making them important sources of greenhouse gases over geological time. Quantification of methane emission would require detailed insights into the capacity and efficiency of microbial metabolisms either consuming or producing methane in the subsurface, and establishment of the linkage between these methane-related metabolisms and other microbial or abiotic processes. Here we conducted geochemical, microbiological and genetic analyses of sediments, gases, and pore and surface fluids to characterize fluid processes, community assemblages, functions and activities in a methane-emitting MV of southwestern Taiwan. Multiple lines of evidence suggest that aerobic/anaerobic methane oxidation, sulfate reduction and methanogenesis are active and compartmentalized into discrete, stratified niches, resembling those in marine settings. Surface evaporation and oxidation of sulfide minerals are required to account for the enhanced levels of sulfate that fuels subsurface sulfate reduction and anaerobic methanotrophy. Methane flux generated by in situ methanogenesis appears to alter the isotopic compositions and abundances of thermogenic methane migrating from deep sources, and to exceed the capacity of microbial consumption. This metabolic stratification is sustained by chemical disequilibria induced by the mixing between upward, anoxic, methane-rich fluids and downward, oxic, sulfate-rich fluids.
metabolic stratification; terrestrial mud volcano; sulfate-to-methane transition zone; methanogenesis; 16S rRNA gene clone library; metagenome
Spontaneous 46,XX primary ovarian insufficiency (POI), also known as ‘premature menopause’ or ‘premature ovarian failure’, refers to ovarian dysfunction that results in a range of abnormalities, from infertility to early menopause as the end stage. The most common known genetic cause of POI is the expansion of a CGG repeat to 55–199 copies (premutation) in the 5′ untranslated region in the X-linked fragile X mental retardation 1 (FMR1) gene. POI associated with the FMR1 premutation is referred to as fragile X-associated POI (FXPOI). Here, we characterize a mouse model carrying the human FMR1 premutation allele and show that FMR1 premutation RNA can cause a reduction in the number of growing follicles in ovaries and is sufficient to impair female fertility. Alterations in selective serum hormone levels, including FSH, LH and 17β-estradiol, are seen in this mouse model, which mimics findings in humans. In addition, we also find that LH-induced ovulation-related gene expression is specifically altered. Finally, we show that the FMR1 premutation allele can lead to reduced phosphorylation of Akt and mTOR proteins. These results together suggest that FMR1 premutation RNA could cause the POI associated with FMR1 premutation carriers, and the Akt/mTOR pathway may serve as a therapeutic target for FXPOI.
Background. The suppression of Helicobacter pylori (H. pylori) decreases H. pylori-related diseases. The probiotics have an inhibitory effect on H. pylori. Aim. We investigated the effects of long-term use of yogurt on H. pylori based on Mongolian gerbils' model. Materials and Methods. Yogurt (containing a supplement of Lactobacillus acidophilus, Bifidobacterium lactis, etc.) was used. Forty-six gerbils were divided into five groups. All groups were inoculated with H. pylori for 5 to 8 weeks. The yogurt was given as follows: Group (Gr.) A: from 1st to 4th week; Gr. B from 5th to 8th week; Gr. C: from 17th week to sacrifice; Gr. D: from 5th week to sacrifice. Gerbils were sacrificed on the 52nd week. Histology was evaluated according to the Sydney system. Results. The positive rates of H. pylori were 60% (Gr. A), 75% (Gr. B), 67% (Gr. C), 44% (Gr. D), and 100% (Gr. E). Gr. D showed lower inflammatory score. Only Gr. E (60%) had intestinal metaplasia. Gr. D showed higher IL-10 and lower TNF-α expression than Gr. E. Conclusion. Long-term intake of yogurt could decrease H. pylori infection. The long-term use of yogurt would be an alternative strategy to manage H. pylori infection.
Q fever is serologically cross-reactive with other intracellular microorganisms. However, studies of the serological status of Mycoplasma pneumoniae and Chlamydophila pneumoniae during Q fever are rare. We conducted a retrospective serological study of M. pneumoniae and C. pneumoniae by enzyme-linked immunosorbent assay (ELISA), a method widely used in clinical practice, in 102 cases of acute Q fever, 39 cases of scrub typhus, and 14 cases of murine typhus. The seropositive (57.8%, 7.7%, and 0%, p<0.001) and seroconversion rates (50.6%, 8.8%, and 0%, p<0.001) of M. pneumoniae IgM, but not M. pneumoniae IgG and C. pneumoniae IgG/IgM, in acute Q fever were significantly higher than in scrub typhus and murine typhus. Another ELISA kit also revealed a high seropositivity (49.5%) and seroconversion rate (33.3%) of M. pneumoniae IgM in acute Q fever. The temporal and age distributions of patients with positive M. pneumoniae IgM were not typical of M. pneumoniae pneumonia. Comparing acute Q fever patients who were positive for M. pneumoniae IgM (59 cases) with those who were negative (43 cases), the demographic characteristics and underlying diseases were not different. In addition, the clinical manifestations associated with atypical pneumonia, including headache (71.2% vs. 81.4%, p=0.255), sore throat (8.5% vs. 16.3%, p=0.351), cough (35.6% vs. 23.3%, p=0.199), and chest x-ray suggesting pneumonia (19.3% vs. 9.5%, p=0.258), were unchanged between the two groups. Clinicians should be aware of the high seroprevalence of M. pneumoniae IgM in acute Q fever, particularly with ELISA kits, which can lead to misdiagnosis, overestimations of the prevalence of M. pneumoniae pneumonia, and underestimations of the true prevalence of Q fever pneumonia.
Inflammation; Mammalian target of rapamycin; LDL receptor pathway; Atherosclerosis
Accumulation of microtubule-associated protein tau has been observed in the brain of aging and tauopathies. Tau was observed in microglia, but its role is not illustrated. By immunofluorescence staining and the fractal dimension value assay in the present study, we observed that microglia were activated in the brains of rats and mice during aging, simultaneously, the immunoreactivities of total tau and the phosphorylated tau were significantly enhanced in the activated microglia. Furtherly by transient transfection of tau40 (human 2N/4R tau) into the cultured rat microglia, we demonstrated that expression of tau40 increased the level of Iba1, indicating activation of microglia. Moreover, expression of tau40 significantly enhanced the membranous localization of the phosphorylated tau at Ser396 in microglia possibly by a mechanism involving protein phosphatase 2A, extracellular signal-regulated kinase and glycogen synthase kinase-3β. It was also found that expression of tau40 promoted microglial migration and phagocytosis, but not proliferation. And we observed increased secretion of several cytokines, including interleukin (IL)-1β, IL-6, IL-10, tumor necrosis factor-α and nitric oxide after the expression of tau40. These data suggest a novel role of human 2N/4R tau in microglial activation.
Aromatic essential oils extracted from fresh fruits of Litsea cubeba (Lour.) Pers., have diverse medical and economic values. The dominant components in these essential oils are monoterpenes and sesquiterpenes. Understanding the molecular mechanisms of terpenoid biosynthesis is essential for improving the yield and quality of terpenes. However, the 40 available L. cubeba nucleotide sequences in the public databases are insufficient for studying the molecular mechanisms. Thus, high-throughput transcriptome sequencing of L. cubeba is necessary to generate large quantities of transcript sequences for the purpose of gene discovery, especially terpenoid biosynthesis related genes.
Using Illumina paired-end sequencing, approximately 23.5 million high-quality reads were generated. De
novo assembly yielded 68,648 unigenes with an average length of 834 bp. A total of 38,439 (56%) unigenes were annotated for their functions, and 35,732 and 25,806 unigenes could be aligned to the GO and COG database, respectively. By searching against the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG), 16,130 unigenes were assigned to 297 KEGG pathways, and 61 unigenes, which contained the mevalonate and 2-C-methyl-D-erythritol 4-phosphate pathways, could be related to terpenoid backbone biosynthesis. Of the 12,963 unigenes, 285 were annotated to the terpenoid pathways using the PlantCyc database. Additionally, 14 terpene synthase genes were identified from the transcriptome. The expression patterns of the 16 genes related to terpenoid biosynthesis were analyzed by RT-qPCR to explore their putative functions.
RNA sequencing was effective in identifying a large quantity of sequence information. To our knowledge, this study is the first exploration of the L. cubeba transcriptome, and the substantial amount of transcripts obtained will accelerate the understanding of the molecular mechanisms of essential oils biosynthesis. The results may help improve future genetic and genomics studies on the molecular mechanisms behind the chemical composition of essential oils in L. cubeba fruits.
Leptospira-induced macrophage death has been confirmed to play a crucial role in pathogenesis of leptospirosis, a worldwide zoonotic infectious disease. Intracellular free Ca2+ concentration ([Ca2+]i) elevation induced by infection can cause cell death, but [Ca2+]i changes and high [Ca2+]i-induced death of macrophages due to infection of Leptospira have not been previously reported.
We first used a Ca2+-specific fluorescence probe to confirm that the infection of L. interrogans strain Lai triggered a significant increase of [Ca2+]i in mouse J774A.1 or human THP-1 macrophages. Laser confocal microscopic examination showed that the [Ca2+]i elevation was caused by both extracellular Ca2+ influx through the purinergic receptor, P2X7, and Ca2+ release from the endoplasmic reticulum, as seen by suppression of [Ca2+]i elevation when receptor-gated calcium channels were blocked or P2X7 was depleted. The LB361 gene product of the spirochete exhibited phosphatidylinositol phospholipase C (L-PI-PLC) activity to hydrolyze phosphatidylinositol-4,5-bisphosphate (PIP2) into inositol-1,4,5-trisphosphate (IP3), which in turn induces intracellular Ca2+ release from endoplasmic reticulum, with the Km of 199 µM and Kcat of 8.566E-5 S-1. Secretion of L-PI-PLC from the spirochete into supernatants of leptospire-macrophage co-cultures and cytosol of infected macrophages was also observed by Western Blot assay. Lower [Ca2+]i elevation was induced by infection with a LB361-deficient leptospiral mutant, whereas transfection of the LB361 gene caused a mild increase in [Ca2+]i. Moreover, PI-PLCs (PI-PLC-β3 and PI-PLC-γ1) of the two macrophages were activated by phosphorylation during infection. Flow cytometric detection demonstrated that high [Ca2+]i increases induced apoptosis and necrosis of macrophages, while mild [Ca2+]i elevation only caused apoptosis.
This study demonstrated that L. interrogans infection induced [Ca2+]i elevation through extracellular Ca2+ influx and intracellular Ca2+ release cause macrophage apoptosis and necrosis, and the LB361 gene product was shown to be a novel PI-PLC of L. interrogans responsible for the [Ca2+]i elevation.