The MeltPro TB/INH assay, recently approved by the Chinese Food and Drug Administration, is a closed-tube, dual-color, melting curve analysis-based, real-time PCR test specially designed to detect 30 isoniazid (INH) resistance mutations in katG position 315 (katG 315), the inhA promoter (positions −17 to −8), inhA position 94, and the ahpC promoter (positions −44 to −30 and −15 to 3) of Mycobacterium tuberculosis. Here we evaluated both the analytical performance and clinical performance of this assay. Analytical studies with corresponding panels demonstrated that the accuracy for detection of different mutation types (10 wild-type samples and 12 mutant type samples), the limit of detection (2 × 103 to 2 × 104 bacilli/ml), reproducibility (standard deviation [SD], <0.4°C), and the lowest heteroresistance level (40%) all met the parameters preset by the kit. The assay could be run on five types of real-time PCR machines, with the shortest running time (105 min) obtained with the LightCycler 480 II. Clinical studies enrolled 1,096 clinical isolates collected from three geographically different tuberculosis centers, including 437 INH-resistant isolates and 659 INH-susceptible isolates characterized by traditional drug susceptibility testing on Löwenstein-Jensen solid medium. The clinical sensitivity and specificity of the MeltPro TB/INH assay were 90.8% and 96.4%, respectively. DNA sequencing analysis showed that, except for the 5 mutants outside the detection range of the MeltPro assay, a concordance rate between the two methods of 99.1% (457/461) was obtained. Among the 26 mutation types detected, katG S315T (AGC→ACC), inhA −15C→T, katG S315N (AGC→AAC), and ahpC promoter −10C→T accounted for more than 90%. Overall, the MeltPro TB/INH assay represents a reliable and rapid tool for the detection of INH resistance in clinical isolates.
Ataxia telangiectasia mutated (ATM) gene is critical in the process of recognizing and repairing DNA lesions and is related to invasion and metastasis of malignancy. The incidence rate of papillary thyroid cancer (PTC) has increased for several decades and is higher in females than males. In this study, we want to investigate whether ATM polymorphisms are associated with gender-specific metastasis of PTC. 358 PTC patients in Northern China, including 109 males and 249 females, were included in our study. Four ATM single nucleotide polymorphisms (SNPs) were genotyped using Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-TOF-MS). Association between genotypes and the gender-specific risk of metastasis was assessed by odds ratios (OR) and 95% confidence intervals (CI) under the unconditional logistic regression analysis. Significant associations were observed between rs189037 and metastasis of PTC in females under different models of inheritance (codominant model: OR = 0.15, 95% CI 0.04–0.56, P = 0.01 for GA versus GG and OR = 0.08, 95% CI 0.01–0.74, P = 0.03 for AA versus GG, resp.; dominant model: OR = 0.49, 95% CI 0.25–0.98, P = 0.04; overdominant model: OR = 0.47, 95% CI 0.25–0.89, P = 0.02). However, no association remained significant after Bonferroni correction. Our findings suggest a possible association between ATM rs189037 polymorphisms and metastasis in female PTCs.
A dam ant colony optimization (D-ACO) analysis of the overall stability of high arch dams on complicated foundations is presented in this paper. A modified ant colony optimization (ACO) model is proposed for obtaining dam concrete and rock mechanical parameters. A typical dam parameter feedback problem is proposed for nonlinear back-analysis numerical model based on field monitoring deformation and ACO. The basic principle of the proposed model is the establishment of the objective function of optimizing real concrete and rock mechanical parameter. The feedback analysis is then implemented with a modified ant colony algorithm. The algorithm performance is satisfactory, and the accuracy is verified. The m groups of feedback parameters, used to run a nonlinear FEM code, and the displacement and stress distribution are discussed. A feedback analysis of the deformation of the Lijiaxia arch dam and based on the modified ant colony optimization method is also conducted. By considering various material parameters obtained using different analysis methods, comparative analyses were conducted on dam displacements, stress distribution characteristics, and overall dam stability. The comparison results show that the proposal model can effectively solve for feedback multiple parameters of dam concrete and rock material and basically satisfy assessment requirements for geotechnical structural engineering discipline.
Diabetes mellitus (DM) is an incurable metabolic disease constituting a major threat to human health. Insulin-producing cells (IPCs) differentiated from mesenchymal stem cells (MSCs) hold great promise in the treatment of DM. The development of an efficient IPC induction system is a crucial step for the clinical application of IPCs for DM. Laminin 411 is a key component of the basement membrane and is involved in the regulation of cell differentiation; however, little is known about a role of laminin 411 in the regulation of IPC differentiation from human MSCs.
MSCs were isolated from human umbilical cord (UC-MSCs) and expanded in an in vitro culture system. UC-MSCs were then cultured in the IPC induction and differentiation medium in the presence of laminin 411. Flow cytometry, Quantitative realtime PCR, immunofluorescence staining, ELISA, Western blotting and other techniques were applied to determine IPC generation, insulin expression and related mechanisms. To evaluate potential therapeutic efficacy of IPCs induced from UC-MSCs, a type-1 diabetes (T1DM) rat model was generated using streptozotocin. Blood glucose, insulin levels, and survival of rats were monitored periodically following intravenous injection of the tested cells.
Laminin 411 markedly induced the expression of the genes Foxa2 and Sox17, markers for pancreatic precursor cells, efficiently induced IPC differentiation from MSCs, and up-regulated insulin expression at both mRNA and protein levels. Furthermore, the expression of the genes known to govern insulin expression including Pdx1 and Ngn3 was markedly induced by laminin 411, which suggests that Pdx1 and Ngn3 signaling pathways are involved in laminin 411 induced-insulin expression machinery. More importantly, administration of laminin 411-induced IPCs rapidly and significantly down-regulated fasting blood glucose levels, significantly reduced the HbA1c concentration and markedly improved the symptoms and survival of T1DM rats.
Our results demonstrate that laminin 411 acts as a potent differentiation inducer of IPCs from UC-MSCs via the Pdx1 and Ngn3 signaling pathways. Moreover, transfusion of laminin 411 induced-IPCs more efficiently improves symptoms and survival of T1DM rats. These novel finding highlights a potential clinical application of laminin 411 induced-IPCs in the treatment of T1DM, which calls for further studies.
Laminin 411; Mesenchymal stem cell; Insulin-producing cell
Soybeans and other pulses contain oligosaccharides which may cause intestinal disturbances such as flatulence. This study was undertaken to investigate α-galactosidase-producing probiotics added to frozen foods which can survive warming treatments used in thawing and consumption of the pulses. The maximum α-galactosidase activity (1.26 U/mg protein) was found in Bifidobacterium breve S46. Lactobacillus casei had the highest α-galactosidase thermostability among the various strains, with D values of 35, 29, and 9.3 minutes at 50°C, 55°C, and 60°C, respectively. The enzyme activity was less affected than viable cells by heating. However, the D values of two bacterial enzymes were lower than those of three commercial α-galactosidase-containing products. Freshly grown cells and their enzymes were more stable than the rehydrated cultures and their enzymes. Practical Application. Enzymes and cultures can be added to foods in order to enhance the digestibility of carbohydrates in the gastrointestinal tract. However since many foods are warmed, it is important that the thermostability of the enzymes be assessed. This paper provides data on the stability of α-galactosidase, which could potentially be added to food matrices containing stachyose or raffinose, such as beans.
The use of mesenchymal stromal cells (MSCs) for treatment of bacterial infections, including systemic processes like sepsis, is an evolving field of investigation. The present study was designed to investigate the potential use of MSCs, harvested from compact bone, and their interactions with the innate immune system, during polymicrobial sepsis induced by cecal ligation and puncture (CLP). We also wanted to elucidate the role of endogenous HO-1 in MSCs during a systemic bacterial infection. MSCs harvested from the bones of heme oxygenase (HO)-1 deficient (−/−) and wild-type (+/+) mice improved the survival of HO-1−/− and HO-1+/+ recipient mice when administered after the onset of polymicrobial sepsis induced by cecal ligation and puncture (CLP), compared with the administration of fibroblast control cells. The MSCs, originating from compact bone in mice, enhanced the ability of neutrophils to phagocytize bacteria in vitro and in vivo, and to promote bacterial clearance in the peritoneum and blood after CLP. Moreover, after depleting neutrophils in recipient mice, the beneficial effects of MSCs were entirely lost, demonstrating the importance of neutrophils for this MSC response. MSCs also decreased multiple organ injury in susceptible HO-1−/− mice, when administered after the onset of sepsis. Taken together, these data demonstrate that the beneficial effects of treatment with MSCs after the onset of polymicrobial sepsis is not dependent on endogenous HO-1 expression, and that neutrophils are crucial for this therapeutic response.
polymicrobial sepsis; mesenchymal stromal cells; compact bone; neutrophils; bacterial clearance; cytoprotective
Periprocedural myocardial infarction (PMI) may occur in approximately 5% to 30% of patients undergoing percutaneous coronary intervention. Whether the morphology of coronary plaque calcium affects the occurrence of PMI is unknown.
Materials and Methods
A total of 616 subjects with stable angina and normal baseline cardiac troponin I levels who had undergone computed tomography angiography (CTA) were referred to elective percutaneous coronary intervention. The morphology of coronary calcium was determined by CTA analysis. PMI was defined as an elevation in 24-h post-procedural cardiac troponin I levels of > 5 times the upper limit of normal with either symptoms of myocardial ischemia, new ischemic electrocardiographic changes, or documented complications during the procedure. Logistic regression was performed to identify the effect of the morphology of coronary calcium on the occurrence of PMI.
According to the presence or morphology of coronary calcium as shown by CTA, 210 subjects were grouped in the heavy calcification group, 258 in the mild calcification group, 40 in the spotty calcification group and 108 in the control group. The dissection rate was significantly higher in the heavy calcification group than in the control group (7.1 % vs. 1.9%, p = 0.03). The occurrence of PMI in the heavy calcification group was significantly higher than that in the control group (OR 4.38, 95% CI 1.80–10.65, p = 0.001). After multivariate adjustment, the risk of PMI still remained significantly higher in the heavy calcification group than in the control group (OR 4.04, 95% CI 1.50–10.89, p = 0.003).
The morphology of coronary calcium determined by CTA may help to predict the subsequent occurrence of PMI. A large amount of coronary calcium may be predictive of PMI.
The aim of this study was to elucidate the prevalence of transmitted drug-resistant (TDR) mutations and reverse transcriptase (RT) thumb subdomain polymorphisms in CRF01_AE and CRF07_BC virus among newly diagnosed, therapy-naive HIV-1 patients in Guangdong Province, China. One hundred and sixty-four samples were collected in the Guangzhou Eighth People's Hospital. The entire protease gene and 300 codons of the entry part of the reverse transcriptase were amplified and sequenced. Furthermore, genotypic drug resistance, polymorphisms, and their phylogeny were analyzed. According to eligibility criteria, seven samples were excluded, and 119 of 157 (75.8%) samples (84 CRF01_AE and 35 CRF07_BC) were amplified and sequenced successfully. The prevalence of TDR identified in the present study was 6.7% [8/119, 95% confidence interval (CI) 1.8–11.6%]. Three major resistance mutations, K103N, M184V, and Y188L, each of which caused more than one drug resistance, appeared in only two patients; the prevalence [1.7 % (2/119)] was relatively low. Until now, this is the first observation of the five newly identified accessory mutations, V35T, K43E, V60I, K122E, and E203D, and seven thumb subdomain polymorphisms, A272P, K277R, K281R, T286A, E291D, V292I, and I293V, in the RT gene in China. These findings provide useful information for guidance on the antiretroviral therapy (ART) policy in China where therapeutic options are still limited.
Infectious bronchitis virus (IBV), a prototype of the Coronaviridae family, is an economically important causative agent of infectious bronchitis in chickens and causes an acute and highly contagious upper respiratory tract infections that may lead to nephritis. However, the molecular antiviral mechanisms of chickens to IBV infection remain poorly understood. In this study, we conducted global gene expression profiling of chicken kidney tissue after nephropathogenic IBV infection to better understand the interactions between host and virus.
IBV infection contributed to differential expression of 1777 genes, of which 876 were up-regulated and 901 down-regulated in the kidney compared to those of control chickens and 103 associated with immune and inflammatory responses may play important roles in the host defense response during IBV infection. Twelve of the altered immune-related genes were confirmed by real-time RT-PCR. Gene ontology category, KEGG pathway, and gene interaction networks (STRING analysis) were analyzed to identify relationships among differentially expressed genes involved in signal transduction, cell adhesion, immune responses, apoptosis regulation, positive regulation of the I-kappaB kinase/NF-kappaB cascade and response to cytokine stimulus. Most of these genes were related and formed a large network, in which IL6, STAT1, MYD88, IRF1 and NFKB2 were key genes.
Our results provided comprehensive knowledge regarding the host transcriptional response to IBV infection in chicken kidney tissues, thereby providing insight into IBV pathogenesis, particularly the involvement of innate immune pathway genes associated with IBV infection.
Infectious bronchitis virus; Kidney; Microarray; Transcriptome
Mutations 1 295 228 C>T and 1 295 250 C>T (termed C228T and C250T respectively), corresponding to −124 C>T and −146 C>T from the translation start site in the promoter of the telomerase reverse transcriptase (TERT) gene, have recently been reported in human cancers, but not in thyroid cancers yet. We explored these mutations in thyroid cancers by genomic sequencing of a large number of primary tumor samples. We found the C228T mutation in 0 of 85 (0.0%) benign thyroid tumors, 30 of 257 (11.7%) papillary thyroid cancers (PTC), 9 of 79 (11.4%) follicular thyroid cancers (FTC), 3 of 8 (37.5%) poorly differentiated thyroid cancers (PDTC), 23 of 54 (42.6%) anaplastic thyroid cancers (ATC), and 8 of 12 (66.7%) thyroid cancer cell lines. The C250T mutation was uncommon, but mutually exclusive with the C228T mutation, and the two mutations were collectively found in 11 of 79 (13.9%) FTC, 25 of 54 (46.3%) ATC, and 11 of 12 (91.7%) thyroid cancer cell lines. Among PTC variants, the C228T mutation was found in 4 of 13 (30.8%) tall-cell PTC (TCPTC), 23 of 187 (12.3%) conventional PTC, and 2 of 56 (3.6%) follicular variant PTC samples. No TERT mutation was found in 16 medullary thyroid cancer samples. The C228T mutation was associated with the BRAF V600E mutation in PTC, being present in 19 of 104 (18.3%) BRAF mutation-positive PTC vs 11 of 153 (7.2%) the BRAF mutation-negative PTC samples (P=0.0094). Conversely, BRAF mutation was found in 19 of 30 (63.3%) C228T mutation-positive PTC vs 85 of 227 (37.4%) C228T mutation-negative PTC samples (P=0.0094). We thus for the first time, to our knowledge, demonstrate TERT promoter mutations in thyroid cancers, that are particularly prevalent in the aggressive thyroid cancers TCPTC, PDTC, ATC and BRAF mutation-positive PTC, revealing a novel genetic background for thyroid cancers.
TERT promoter mutations; thyroid cancers; BRAF V600E mutation; telomerase reverse transcriptase; thyroid tumorigenesis
Glioblastomas are the most aggressive forms of primary brain tumors due to their tendency to invade surrounding healthy brain tissues, rendering them largely incurable. The water channel protein, Aquaporin-4 (AQP4) is a key molecule for maintaining water and ion homeostasis in the central nervous system and has recently been reported with cell survival except for its well-known function in brain edema. An increased AQP4 expression has been demonstrated in glioblastoma multiforme (GBM), suggesting it is also involved in malignant brain tumors. In this study, we show that siRNA-mediated down regulation of AQP4 induced glioblastoma cell apoptosis in vitro and in vivo. We further show that several apoptotic key proteins, Cytochrome C, Bcl-2 and Bad are involved in AQP4 signaling pathways. Our results indicate that AQP4 may serve as an anti-apoptosis target for therapy of glioblastoma.
The present study investigates the varied spatial distribution of syphilis cases in Shenzhen, China, and explores the individual-, neighbourhood- and district-level factors affecting the distribution.
This study uses spatial analysis and multi-level generalised estimating equations to explore the spatial distribution of reported syphilis cases among individuals in Shenzhen, Guangdong Province, China. The spatial distribution of primary/secondary and latent cases was investigated using the Moran’s I-statistic. Primary/secondary syphilis cases were compared with all syphilis cases using a three-level model with individual (n=6496), neighbourhood (n=55) and district (n=6) levels.
A total of 6496 syphilis cases were reported in 2009 with 35.8% primary and secondary syphilis cases. Both primary/secondary syphilis cases (Moran’s I value=0.33, p<0.01) and latent syphilis cases (Moran’s I value=0.19, p<0.01) showed significant spatial clustering at the neighbourhood level. Adjusting for the number of reporting hospitals, the best model found that the following characteristics were associated with primary/secondary syphilis infection: individuals who are younger in age (p=0.003), male (p<0.001), migrant labourers (p=0.047) and those who live in districts with a higher gross domestic product (p<0.001).
There is substantial clustering of primary and secondary syphilis cases at the neighbourhood level in Shenzhen, suggesting the need for greater STD health service provision in these clustered neighbourhoods. Spatially targeted syphilis control measures may be useful to optimise testing, treatment and partner services.
This Letter proposes a new strategy of a three-dimensional (3D) scanning pipeline to achieve complete 3D digitization of complex objects in a real scene. This strategy consists of a one-dimensional array of optical 3D sensors combined with an automatically controlled turntable. An efficient calibration method for the sensor array is presented to guarantee the accuracy of the 3D measurement. Furthermore, an automatic registration technique is also proposed for aligning multiple range images taken from sensor array. Experiment results are also presented to demonstrate the proposed approach.
Acyl homoserine lactone (AHL)-based quorum sensing commonly refers to cell density-dependent regulatory mechanisms found in bacteria. However, beyond bacteria, this cell-to-cell communication mechanism is poorly understood. Here we show that a methanogenic archaeon, Methanosaeta harundinacea 6Ac, encodes an active quorum sensing system that is used to regulate cell assembly and carbon metabolic flux. The methanogen 6Ac showed a cell density-dependent physiology transition, which was related to the AHL present in the spent culture and the filI gene-encoded AHL synthase. Through extensive chemical analyses, a new class of carboxylated AHLs synthesized by FilI protein was identified. These carboxylated AHLs facilitated the transition from a short cell to filamentous growth, with an altered carbon metabolic flux that favoured the conversion of acetate to methane and a reduced yield in cellular biomass. The transcriptomes of the filaments and the short cell forms differed with gene expression profiles consistent with the physiology. In the filaments, genes encoding the initial enzymes in the methanogenesis pathway were upregulated, whereas those for cellular carbon assimilation were downregulated. A luxI–luxR ortholog filI–filR was present in the genome of strain 6Ac. The carboxylated AHLs were also detected in other methanogen cultures and putative filI orthologs were identified in other methanogenic genomes as well. This discovery of AHL-based quorum sensing systems in methanogenic archaea implies that quorum sensing mechanisms are universal among prokaryotes.
carboxylated acyl homoserine lactones; filI-encoded AHL synthase; methanogenic archaea; physiology transition; quorum sensing
To understand better the risk of tuberculosis transmission with increasing delay in tuberculosis treatment, we undertook a retrospective cohort study in Shenzhen, China.
All pulmonary tuberculosis cases in the Shenzhen tuberculosis surveillance database from 1993–2010 were included. Sputum smear positivity and presence of pulmonary cavity were used as proxies for risk of tuberculosis transmission.
Among 48,441pulmonary tuberculosis cases, 70% presented with symptoms of pulmonary TB, 62% were sputum smear positive, and 21% had a pulmonary cavity on chest x-ray. 95.3% of patients self-presented for evaluation of illness after a median 58 days of delay after symptoms began. The proportion presenting sputum smear positive (p<0.001) and with a pulmonary cavity (p<0.001) increased significantly with increasing duration of delay.
Delayed diagnosis and treatment of tuberculosis is associated with a significantly increased risk of pulmonary sputum smear positivity and pulmonary cavity. To decrease risk of transmission, treatment delay needs to be reduced further.
Background & Aims
Acute-on-chronic liver failure (ACLF) is one of the most deadly, prevalent, and costly diseases in Asia. However, no prognostic model has been developed that is based specifically on data gathered from Asian patients with ACLF. The aim of the present study was to quantify the survival time of ACLF among Asians and to develop a prognostic model to estimate the probability of death related to ACLF.
We conducted a retrospective observational cohort study to analyze clinical data from 857 patients with ACLF/pre-ACLF who did not undergo liver transplantation. Kaplan–Meier and Cox proportional hazards regression model were used to estimate survival rates and survival affected factors. The area under the receiver operating characteristic curve (auROC) was used to evaluate the performance of the models for predicting early mortality.
The mortality rates among patients with pre-ACLF at 12 weeks and 24 weeks after diagnosis were 30.5% and 33.2%, respectively. The mortality rates among patients with early-stage ACLF at 12 weeks and 24 weeks after diagnosis were 33.9% and 37.1%, respectively. The difference in survival between pre-ACLF patients and patients in the early stage of ACLF was not statistically significant. The prognostic model identified 5 independent factors significantly associated with survival among patients with ACLF and pre-ACLF: the model for end-stage liver disease (MELD) score; age, hepatic encephalopathy; triglyceride level and platelet count.
The findings of the present study suggest that the Chinese diagnostic criteria of ACLF might be broadened, thus enabling implementation of a novel model to predict ACLF-related death after comprehensive medical treatment.
Coastal line is now polluted by many kinds of sewage including heavy metals discharged by intensive human activities. Cadmium is a nonessential heavy metal for organisms and can cause many kinds of adverse effect on the organisms. Suaeda salsa, a pioneer halophyte in intertidal zone of the Bohai coast, was proved to have cadmium-tolerant capacity. Given that, S. salsa was suggested as a potential coastal bio-indicator plant for cadmium contamination in the intertidal zone. Therefore, it is essential to investigate the responsive mechanism of S. salsa to cadmium since few studies focus on this subject till now. In the present study, six genes were selected to investigate the variation profiles of mRNA expression by fluorescent real-time quantitative PCR, including those involved in myo-inositol synthesis, redox reaction, salt-tolerant reaction. Results showed that cadmium exposure significantly modulate the mRNA expressions of MIPS, Nhx1, CAT2, GST, Prx Q genes. It suggested that cadmium exposure exerted an oxidative stress on S. salsa, disturbed Na+ homeostasis across membranes and interfered with the metabolism of inositol. In addition, CAT2 gene could be used as a gene marker in S. salsa to indicate cadmium pollution.
Suaeda salsa; qRT-PCR; Gene expression; Cadmium
As an environmental contaminant, mercury is of great concern due to its high risk to environmental and human health. The halophyte Suaeda salsa is the dominant plant in the intertidal zones of the Yellow River Delta (YRD) where has been contaminated by mercury in some places. This study aimed at evaluating the chronic effects of mercury (Hg2+, 20 µg L−1) and the influence of an environmentally relevant salinity (NaCl, 500 mM) on mercury-induced effects in S. salsa. A total of 43 protein spots with significant changes were identified in response to Hg2+, salinity and combined Hg2+ and salinity. These proteins can be categorized into diverse functional classes, related to metabolic processes, photosynthesis, stress response, protein fate, energy metabolism, signaling pathways and immunosuppression. Metabolic responses demonstrated that Hg2+ could disturb protein and energy metabolisms in S. salsa co-exposed with or without salinity. In addition, both antagonistic and synergistic effects between Hg2+ and salinity were confirmed by differential levels of proteins (magnesium-chelatase and ribulose-l,5-bisphosphate carboxylase/oxygenase) and metabolites (valine, malonate, asparagine, glycine, fructose and glucose) in S. salsa. These findings suggest that a combination of proteomics and metabolomics can provide insightful information of environmental contaminant-induced effects in plants at molecular levels.
A sinusoidal modulation scheme is described for selective heteronuclear polarization transfer between two dilute spins in double cross polarization magic-angle-spinning nuclear magnetic resonance spectroscopy. During the second N → C cross polarization, the 13C RF amplitude is modulated sinusoidally while the 15N RF amplitude is tangent. This modulation induces an effective spin-lock field in two selective frequency bands in either side of the 13C RF carrier frequency, allowing for simultaneous polarization transfers from 15N to 13C in those two selective frequency bands. It is shown by experiments and simulations that this sinusoidal modulation allows one to selectively polarize from 15N to its covalently bonded 13Cα and 13C′ carbons in neighboring peptide planes simultaneously, which is useful for establishing the back-bone connectivity between two sequential residues in protein structural elucidation. The selectivity and efficiency were experimentally demonstrated on a uniformly 13C,15N-labeled β1 immunoglobulin binding domain of protein G (GB1).
Double cross polarization (DCP); NCA/NCO; Solid-state MAS NMR; Heteronuclear polarization transfer
The search for a strategy to provide temporary liver support and salvage the patients with acute-on-chronic liver failure (ACLF) remains an important issue. This study was designed to evaluate the experience in artificial liver support system (ALSS) combined with liver transplantation (LT) in the treatment of ACLF.
One hundred and seventy one patients with HBV related ACLF undergoing LT between January 2001 and December 2009 were included. Of the 171 patients, 115 received 247 sessions of plasma exchange-centered ALSS treatment prior to LT (ALSS-LT group) and the other 56 received emergency LT (LT group). The MELD score were 31±6 and 30±7 in ALSS-LT group and LT group. ALSS treatment resulted in improvement of liver function and better tolerance to LT. The average level of serum total bilirubin before LT was lower than that before the first time of ALSS treatment. The median waiting time for a donor liver was 12 days (2–226 days) from the first run of ALSS treatment to LT. Compared to LT group, the beneficial influences of ALSS on intraoperative blood loss and endotracheal intubation time were also observed in ALSS-LT group. The 1-year and 5-year survival rates in the ALSS-LT group and LT group were 79.2% and 83%, 69.7% and 78.6%.
Plasma exchange-centered ALSS is beneficial in salvaging patients with ACLF when a donor liver is not available. The consequential LT is the fundamental treatment modality to rescue these patients and lead to a similar survival rate as those patients receiving emergency transplantation.
Recent studies demonstrate that UHRF1 is required for DNA methylation maintenance by targeting DNMT1 to DNA replication foci, presumably through its unique hemi-methylated DNA-binding activity and interaction with DNMT1. UHRF2, another member of the UHRF family proteins, is highly similar to UHRF1 in both sequence and structure, raising questions about its role in DNA methylation. In this study, we demonstrate that, like UHRF1, UHRF2 also binds preferentially to methylated histone H3 lysine 9 (H3K9) through its conserved tudor domain and hemi-methylated DNA through the SET and Ring associated domain. Like UHRF1, UHRF2 is enriched in pericentric heterochromatin. The heterochromatin localization depends to large extent on its methylated H3K9-binding activity and to less extent on its methylated DNA-binding activity. Coimmunoprecipitation experiments demonstrate that both UHRF1 and UHRF2 interact with DNMT1, DNMT3a, DNMT3b and G9a. Despite all these conserved functions, we find that UHRF2 is not able to rescue the DNA methylation defect in Uhrf1 null mouse embryonic stem cells. This can be attributed to the inability for UHRF2 to recruit DNMT1 to replication foci during S phase of the cell cycle. Indeed, we find that while UHRF1 interacts with DNMT1 in an S phase-dependent manner in cells, UHRF2 does not. Thus, our study demonstrates that UHRF2 and UHRF1 are not functionally redundant in DNA methylation maintenance and reveals the cell-cycle-dependent interaction between UHRF1 and DNMT1 as a key regulatory mechanism targeting DNMT1 for DNA methylation.
UHRF2; UHRF1; DNA methylation; histone methylation; DNMT1
Post-weaning multisystemic wasting syndrome (PMWS) associated with PCV2 is one of the most costly diseases currently faced by the swine industry. The development of effective vaccines against PCV2 infection has been accepted as an important strategy in the prophylaxis of PMWS.
In the present study, a PK-15 cell-adapted formalin-inactivated prototype vaccine candidate was prepared using a strain of PCV2 from China. Inactivation of the virus was accomplished using a standard formalin inactivation protocol. The protective properties of the inactivated PCV2 vaccine were evaluated in piglets. Ten 28-day-old pigs were randomly assigned to two groups, each with five. Group 1 was vaccinated intramuscularly with the inactivated virus preparation; Group 2 received sterile PBS as a placebo. By 28 days post-vaccination (DPV), Groups 1 and 2 were challenged intranasally and intramuscularly with 5 × 107 TCID50 of a virulent PCV2 isolate.
The vaccinated pigs seroconverted to PCV2 and had high levels of serum antibodies to PCV2 at 28 days after vaccination, whereas the control pigs remained seronegative. No significant signs of clinical disease were recorded following the challenge with PCV2, but moderate amounts of PCV2 antigen were detected in most lymphoid organs of the control pigs. PCV2 was detected in two out of the five vaccinated pigs. Furthermore, pathological lesions and viremia were milder in the vaccinated group.
The obtained results indicate that the inactivated PCV2 virus vaccine with an oil adjuvant induce an immunological response in pigs that appears to provide protection from infection with PCV2. The vaccine, therefore, may have the potential to serve as a vaccine aimed to protect pigs from developing PMWS.
Porcine circovirus type 2; Post-weaning multisystemic wasting syndrome; Single-dose immunization; Formalin-inactivated vaccine
Sepsis remains the leading cause of death in critically ill patients despite modern advances in critical care. Intestinal barrier dysfunction may lead to secondary bacterial translocation and the development of the multiple organ dysfunction syndrome during sepsis. Cyclooxygenase-2 (COX-2) is highly upregulated in the intestine during sepsis and we hypothesized that it may be critical in the maintenance of intestinal epithelial barrier function during peritonitis-induced polymicrobial sepsis. COX-2−/− and COX-2+/+ BALB/c mice underwent cecal ligation and puncture (CLP) or sham surgery. Mice chimeric for COX-2 were derived by bone marrow transplantation and underwent CLP. C2BBe1 cells, an intestinal epithelial cell line, were treated with the COX-2 inhibitor NS-398, PGD2, or vehicle and stimulated with cytokines. COX-2−/− mice developed exaggerated bacteremia and increased mortality compared with COX-2+/+ mice following CLP. Mice chimeric for COX-2 exhibited the recipient phenotype suggesting that epithelial COX-2 expression in the ileum attenuates bacteremia following CLP. Absence of COX-2 significantly increased epithelial permeability of the ileum and reduced expression of the tight junction proteins zonula occludens-1 (ZO-1), occludin, and claudin-1 in the ileum following CLP. Furthermore, PGD2 attenuated cytokine-induced hyperpermeability and ZO-1 downregulation in NS-398-treated C2BBe1 cells. Our findings reveal that absence of COX-2 is associated with enhanced intestinal epithelial permeability and leads to exaggerated bacterial translocation and increased mortality during peritonitis-induced sepsis. Taken together, our results suggest that epithelial expression of COX-2 in the ileum is a critical modulator of tight junction protein expression and intestinal barrier function during sepsis.
Nucleotide-binding oligomerization domain protein 2 (NOD2) stimulates diverse inflammatory responses resulting in differential cellular phenotypes. To identify the role of NOD2 in vascular arterial obstructive diseases, we investigated the expression and pathophysiological role of NOD2 in a vascular injury model of neointimal hyperplasia.
Methods and Results
We first analyzed for neointimal hyperplasia following femoral artery injury in NOD2+/+ and NOD2−/− mice. NOD2−/− mice showed a 2.86-fold increase in neointimal formation that was mainly composed of SM α-actin positive cells. NOD2 was expressed in vascular smooth muscle cells (VSMCs) and NOD2−/− VSMCs showed increased cell proliferation in response to mitogenic stimuli, PDGF-BB or fetal bovine serum (FBS), compared with NOD2+/+ VSMCs. Furthermore, NOD2 deficiency markedly promoted VSMCs migration in response to PDGF-BB and this increased cell migration was attenuated by a PI3 kinase inhibitor. However, PKC and JNK inhibitors exerted negligible effects. Moreover, muramyl dipeptide-stimulated NOD2 prevented PDGF-BB-induced VSMCs migration.
Functional NOD2 is expressed in VSMCs, and NOD2 deficiency promoted VSMCs proliferation, migration, and neointimal formation after vascular injury. These results provide evidence for the involvement of NOD2 in vascular homeostasis and tissue injury, serving as a potential molecular target in the modulation of arteriosclerotic vascular disease.