A 62-year-old woman was admitted to our hospital in 2011 because of recurrent abdominal pain, nausea and constipation for six months. Computed tomography enterography (CTE) showed tortuous thread-like calcifications in the ileocolic vein and right colic vein, while colonoscopy revealed purple-blue mucosa extending from the cecum to the splenic flexure. Based on the results of these tests, the patient was diagnosed with idiopathic mesenteric phlebosclerosis (IMP). She had a history of Chinese medical liquor intake for one and a half years and her symptoms subsided after conservative treatment. In 2013, a 63-year-old male patient who presented with recurrent lower right abdominal pain, bloating, melena and diarrhea for fifteen months was admitted to our institution. Colonoscopy and CTE led to the diagnosis of IMP. He also used Chinese medical liquor for approximately 12 years. The patient underwent total colectomy and the postoperative course was uneventful. We searched for previously published reports on similar cases and analyzed the clinical data of 50 cases identified in PubMed. As some of these patients admitted use of Chinese medicines, we hypothesize that Chinese medicines may play a role in the pathogenesis of IMP.
Idiopathic mesenteric phlebosclerosis; Chinese medical liquor; Ileus; Colonoscopy; Computed tomography enterography
AIM: To assess the relationship between the P268S, JW1 and N852S polymorphisms and Crohn’s disease (CD) susceptibility in Zhuang patients in Guangxi, China.
METHODS: Intestinal tissues from 102 Zhuang [48 CD and 54 ulcerative colitis (UC)] and 100 Han (50 CD and 50 UC) unrelated patients with inflammatory bowel disease and 72 Zhuang and 78 Han unrelated healthy individuals were collected in the Guangxi Zhuang Autonomous Region from January 2009 to March 2013. Genomic DNA was extracted using the phenol chloroform method. The P268S, JW1 and N852S polymorphisms were amplified using polymerase chain reaction (PCR), detected by restriction fragment length polymorphism (RFLP), and verified by gene sequencing.
RESULTS: Heterozygous mutation of P268S in the NOD2/CARD15 gene was detected in 10 CD cases (six Zhuang and four Han), two Han UC cases, and one Zhuang healthy control, and P268S was strongly associated with the Chinese Zhuang and Han CD populations (P = 0.016 and 0.022, respectively). No homozygous mutant P268S was detected in any of the groups. No significant difference was found in P268S genotype and allele frequencies between UC and control groups (P > 0.05). Patients with CD who carried P268S were likely to be ≤ 40 years of age (P = 0.040), but were not significantly different with regard to race, lesion site, complications, and other clinical features (P > 0.05). Neither JW1 nor N852S polymorphisms of the NOD2/CARD15 gene were found in any of the subjects (P > 0.05).
CONCLUSION: P268S polymorphism may be associated with CD susceptibility in the Zhuang population in the Guangxi Zhuang Autonomous Region, China. In contrast, JW1 and N852S polymorphisms may not be related to CD susceptibility in these patients.
Crohn’s disease; NOD2/CARD15; Single nucleotide polymorphisms
Zinc (Zn) deficiency and water scarcity are major challenges in rice (Oryza sativa L.) under an intensive rice production system. This study aims to investigate the impact of water-saving management and different Zn fertilization source (ZnSO4 and Zn-EDTA) regimes on grain yield and Zn accumulation in rice grain. Different water managements, continuous flooding (CF), and alternate wetting and drying (AWD) were applied during the rice growing season. Compared with CF, the AWD regime significantly increased grain yield and Zn concentrations in both brown rice and polished rice. Grain yield of genotypes (Nipponbare and Jiaxing27), on the average, was increased by 11.4%, and grain Zn concentration by 3.9% when compared with those under a CF regime. Zn fertilization significantly increased Zn density in polished rice, with a more pronounced effect of ZnSO4 being observed as compared with Zn-EDTA, especially under an AWD regime. Decreased phytic acid content and molar ratio of phytic acid to Zn were also noted in rice grains with Zn fertilization. The above results demonstrated that water management of AWD combined with ZnSO4 fertilization was an effective agricultural practice to elevate grain yield and increase Zn accumulation and bioavailability in rice grains.
Rice; Alternate wetting and drying; Soil fertilization; Zinc
The relationship between resistin and non-alcoholic steatohepatitis (NASH) is not clear, some studies claimed that serum resistin levels were associated with neither the presence of NASH nor its severity, others declared that serum resistin was related with inflammation and fibrosis in NASH. Our animal study verified that the distribution of resistin in the liver is correlated with inflammation in NASH. However, there is no pertinent study in humans.
Thirty patients with NASH, 28 simple steatosis, and 43 controls were recruited. Blood was collected for resistin, liver chemistries, fasting insulin and some metabolic parameters. Liver histology was scored according to NAFLD activity scoring system. Hepatic resistin expression was examined by real-time polymerase chain reaction, immunohistochemistry. Resistin protein expression was confirmed by western blotting in 13 patients with concomitant NAFLD and gallstone.
Serum resistin was significantly elevated in both NASH and simple steatotic subjects compared with controls (all P < 0.05). Hepatic resistin was significantly increased in NASH patients in both mRNA and protein levels than those in simple steatosis and control subjects (all P < 0.05). Both serum and hepatic resistin had a correlation with obesity, but not with insulin resistance. The distribution of resistin positive cells was predominantly in perisinusoidal cells (such as Kupffer cells and hepatic stellate cells) in human NASH. Multivariate analysis revealed that waist-hip ratio, higher serum triglyceride, and hyperresistinemia were independent factors related to higher grade of steatosis; whereas hepatic resistin and serum cytokeratin predict NASH and severity of liver fibrosis.
Hepatic resistin overexpression in NASH patients is associated with the severity of liver inflammation and fibrosis. Liver-derived resistin may be involved in the pathogenesis of human NASH.
Resistin; Nonalcoholic steatohepatitis; Nonalcoholic fatty liver disease; Inflammation; Adipokine
Background: Maternal obesity has adverse effects on oocyte quality, embryo development, and the health of the offspring.
Objectives: To understand the underlying mechanisms responsible for the negative effects of maternal obesity, we investigated the DNA methylation status of several imprinted genes and metabolism-related genes.
Methods: Using a high-fat-diet (HFD)-induced mouse model of obesity, we analyzed the DNA methylation of several imprinted genes and metabolism-related genes in oocytes from control and obese dams and in oocytes and liver from their offspring. Analysis was performed using combined bisulfite restriction analysis (COBRA) and bisulfite sequencing.
Results: DNA methylation of imprinted genes in oocytes was not altered in either obese dams or their offspring; however, DNA methylation of metabolism-related genes was changed. In oocytes of obese mice, the DNA methylation level of the leptin (Lep) promoter was significantly increased and that of the Ppar-α promoter was reduced. Increased methylation of Lep and decreased methylation of Ppar-α was also observed in the liver of female offspring from dams fed the high-fat diet (OHFD). mRNA expression of Lep and Ppar-α was also significantly altered in the liver of these OHFD. In OHFD oocytes, the DNA methylation level of Ppar-α promoter was increased.
Conclusions: Our results indicate that DNA methylation patterns of several metabolism-related genes are changed not only in oocytes of obese mice but also in oocytes and liver of their offspring. These data may contribute to the understanding of adverse effects of maternal obesity on reproduction and health of the offspring.
Citation: Ge ZJ, Luo SM, Lin F, Liang QX, Huang L, Wei YC, Hou Y, Han ZM, Schatten H, Sun QY. 2014. DNA methylation in oocytes and liver of female mice and their offspring: effects of high-fat-diet–induced obesity. Environ Health Perspect 122:159–164; http://dx.doi.org/10.1289/ehp.1307047
Observational studies and small clinical trials suggested that hormone replacement therapy (HRT) decreases risk of cognitive loss and Alzheimer’s disease (AD) in postmenopausal women and may have value in primary prevention.
A clinical trial was designed to determine if HRT delays AD or memory loss. This report describes the rationale and original design of the trial and details extensive modifications that were required to respond to unanticipated findings that emerged from other studies during the course of the trial.
The trial was designed as a multi-center, placebo-controlled primary prevention trial for women 65 years of age or older with a family history of dementia. Recruitment from local sites was supplemented by centralized efforts to use names of Medicare beneficiaries. Inclusion criteria included good general health and intact memory functioning. Participants were randomized to HRT or placebo in a 1:1 ratio. Assignment was stratified by hysterectomy status and site. The primary outcomes were incident AD and memory decline on neuropsychological testing.
Enrollment began in March 1998. In response to the Women’s Health Initiative (WHI) May 2002 report of increased incidence of heart disease, stroke, pulmonary embolism, and breast cancer among women randomized to HRT, participants were re-consented with a revised consent form. Procedural modifications, including discontinuation of study medication and a modification of the planned primary outcome based on a final enrollment below the target enrollment (N = 477), were enacted in response to the subsequent WHI Memory Study report of increased risk of dementia and poorer cognitive function with HRT. The mean length of treatment exposure prior to discontinuation was 2.14 years. Participants’ mean age at baseline was 72.8; mean education was 14.2 years. Minority participation was 19% and 34% had a hysterectomy. The study continues to follow these participants for a total of 5 years blind to the original medication assignment.
Results reported from the WHI during the course of this study mandated extensive procedural modifications, including discontinuing recruitment before completion and halting study medication. Alternative strategies for study redesign that were considered are discussed.
Maternal diabetes mellitus not only has severe deleterious effects on fetal development, but also it affects transmission to the next generation. However, the underlying mechanisms for these effects are still not clear.
We investigated the methylation patterns and expressions of the imprinted genes Peg3, Snrpn, and H19 in mid-gestational placental tissues and on the whole fetus utilizing the streptozotocin (STZ)-induced hyperglycemic mouse model for quantitative analysis of methylation by PCR and quantitative real-time PCR. The protein expression of Peg3 was evaluated by Western blot.
We found that the expression of H19 was significantly increased, while the expression of Peg3 was significantly decreased in dpc10.5 placentas of diabetic mice. We further found that the methylation level of Peg3 was increased and that of H19 was reduced in dpc10.5 placentas of diabetic mice. When pronuclear embryos of normal females were transferred to normal/diabetic (NN/ND) pseudopregnant females, the methylation and expression of Peg3 in placentas was also clearly altered in the ND group compared to the NN group. However, when the pronuclear embryos of diabetic female were transferred to normal pesudopregnant female mice (DN), the methylation and expression of Peg3 and H19 in dpc10.5 placentas was similar between the two groups.
We suggest that the effects of maternal diabetes on imprinted genes may primarily be caused by the adverse uterus environment.
Maternal diabetes milieu; DNA imprinting; Placenta
Dysthymia is a form of chronic mild depression that has a complex relationship with major depressive disorder (MDD). Here we investigate the role of environmental risk factors, including stressful life events and parenting style, in patients with both MDD and dysthymia. We ask whether these risk factors act in the same way in MDD with and without dysthymia.
We examined the clinical features in 5,950 Han Chinese women with MDD between 30–60 years of age across China. We confirmed earlier results by replicating prior analyses in 3,950 new MDD cases. There were no significant differences between the two data sets. We identified sixteen stressful life events that significantly increase the risk of dysthymia, given the presence of MDD. Low parental warmth, from either mother or father, increases the risk of dysthymia. Highly threatening but short-lived threats (such as rape) are more specific for MDD than dysthymia. While for MDD more severe life events show the largest odds ratio versus controls, this was not seen for cases of MDD with or without dysthymia.
There are increased rates of stressful life events in MDD with dysthymia, but the impact of life events on susceptibility to dysthymia with MDD differs from that seen for MDD alone. The pattern does not fit a simple dose-response relationship, suggesting that there are moderating factors involved in the relationship between environmental precipitants and the onset of dysthymia. It is possible that severe life events in childhood events index a general susceptibility to chronic depression, rather than acting specifically as risk factors for dysthymia.
Evidence supporting the impact of therapeutic zinc supplementation on the duration and severity of diarrhea among children under five is largely derived from studies conducted in South Asia. China experiences a substantial portion of the global burden of diarrhea, but the impact of zinc treatment among children under five has not been well documented by previously published systematic reviews on the topic. We therefore conducted a systematic literature review, which included an exhaustive search of the Chinese literature, in an effort to update previously published estimates of the effect of therapeutic zinc. We conducted systematic literature searches in various databases, including the China National Knowledge Infrastructure (CNKI), and abstracted relevant data from studies meeting our inclusion and exclusion criteria. We used STATA 12.0 to pool select outcomes and to generate estimates of percentage difference and relative risk comparing outcomes between zinc and control groups. We identified 89 Chinese and 15 non-Chinese studies for the review, including studies in 10 countries from all WHO geographic regions, and analyzed a total of 18,822 diarrhea cases (9469 zinc and 9353 control). None of the included Chinese studies had previously been included in published pooled effect estimates. Chinese and non-Chinese studies reported the effect of therapeutic zinc supplementation on decreased episode duration, stool output, stool frequency, hospitalization duration and proportion of episodes lasting beyond three and seven days. Pooling Chinese and non-Chinese studies yielded an overall 26% (95% CI: 20%−32%) reduction in the estimated relative risk of diarrhea lasting beyond three days among zinc-treated children. Studies conducted in and outside China report reductions in morbidity as a result of oral therapeutic zinc supplementation for acute diarrhea among children under five years of age. The WHO recommendation for zinc treatment of diarrhea episodes should be supported in all low- and middle-income countries.
zinc; children; global health; China
Altered chloride homeostasis has been thought to be a risk factor for several brain disorders, while less attention has been paid to its role in liver disease. We aimed to analyze the involvement and possible mechanisms of altered chloride homeostasis of GABAergic neurons within the substantia nigra pars reticulata (SNr) in the motor deficit observed in a model of encephalopathy caused by acute liver failure, by using glutamic acid decarboxylase 67 - green fluorescent protein knock-in transgenic mice.
Alterations in intracellular chloride concentration in GABAergic neurons within the SNr and changes in the expression of two dominant chloride homeostasis-regulating genes, KCC2 and NKCC1, were evaluated in mice with hypolocomotion due to hepatic encephalopathy (HE). The effects of pharmacological blockade and/or activation of KCC2 and NKCC1 functions with their specific inhibitors and/or activators on the motor activity were assessed.
In our mouse model of acute liver injury, chloride imaging indicated an increase in local intracellular chloride concentration in SNr GABAergic neurons. In addition, the mRNA and protein levels of KCC2 were reduced, particularly on neuronal cell membranes; in contrast, NKCC1 expression remained unaffected. Furthermore, blockage of KCC2 reduced motor activity in the normal mice and led to a further deteriorated hypolocomotion in HE mice. Blockade of NKCC1 was not able to normalize motor activity in mice with liver failure.
Our data suggest that altered chloride homeostasis is likely involved in the pathophysiology of hypolocomotion following HE. Drugs aimed at restoring normal chloride homeostasis would be a potential treatment for hepatic failure.
Foot-and-mouth disease virus (FMDV) causes a highly contagious infection in cloven-hoofed animals. The format of FMD virus-like particles (VLP) as a non-replicating particulate vaccine candidate is a promising alternative to conventional inactivated FMDV vaccines. In this study, we explored a prokaryotic system to express and assemble the FMD VLP and validated the potential of VLP as an FMDV vaccine candidate. VLP composed entirely of FMDV (Asia1/Jiangsu/China/2005) capsid proteins (VP0, VP1 and VP3) were simultaneously produced as SUMO fusion proteins by an improved SUMO fusion protein system in E. coli. Proteolytic removal of the SUMO moiety from the fusion proteins resulted in the assembly of VLP with size and shape resembling the authentic FMDV. Immunization of guinea pigs, swine and cattle with FMD VLP by intramuscular inoculation stimulated the FMDV-specific antibody response, neutralizing antibody response, T-cell proliferation response and secretion of cytokine IFN-γ. In addition, immunization with one dose of the VLP resulted in complete protection of these animals from homologous FMDV challenge. The 50% protection dose (PD50) of FMD VLP in cattle is up to 6.34. These results suggest that FMD VLP expressed in E. coli are an effective vaccine in guinea pigs, swine and cattle and support further development of these VLP as a vaccine candidate for protection against FMDV.
Despite the abundance of studies on γ-aminobutyric acid (GABA) ergic neuron distribution in the mouse developing spinal cord, no investigation has been devoted so far to their birthdates. In order to determine the spinal neurogenesis of a specific phenotype, the GABAergic neurons in the spinal cord, we injected bromodeoxyuridine (BrdU) at different developmental stages of the glutamic acid decarboxylase (GAD)67-green fluorescent protein (GFP) knock-in mice. We thus used GFP to mark GABAergic neurons and labeled newly born cells with the S-phase marker BrdU at different embryonic stages. Distribution of GABAergic neurons labeled with BrdU was then studied in spinal cord sections of 60-day-old mice. Our birthdating studies revealed that GABAergic neurogenesis was present at embryonic day 10.5 (E10.5). Since then, the generation of GABAergic neurons significantly increased, and reached a peak at E11.5. Two waves for the co-localization of GABA and BrdU in the spinal cord were seen at E11.5 and E13.5 in the present study. The vast majority of GABAergic neurons were generated before E14.5. Thereafter, GABA-positive neuron generation decreased drastically. The present results showed that the birthdates of GABAergic neurons in each lamina were different. The peaks of GABAergic neurogenesis in lamina II were at E11.5 and E13.5, while in lamina I and III, they were at E13.5 and E12.5, respectively. The present results suggest that the birthdates of GABAergic neurons vary in different lamina and follow a specific temporal sequence. This will provide valuable information for future functional studies.
GABAergic neuron; BrdU; birthdate; spinal cord; mouse
Objective. This study was to evaluate the effect of riluzole on methylmercury- (MeHg-) induced oxidative stress, through promotion of glutathione (GSH) synthesis by activating of glutamate transporters (GluTs) in rat cerebral cortex. Methods. Eighty rats were randomly assigned to four groups, control group, riluzole alone group, MeHg alone group, and riluzole + MeHg group. The neurotoxicity of MeHg was observed by measuring mercury (Hg) absorption, pathological changes, and cell apoptosis of cortex. Oxidative stress was evaluated via determining reactive oxygen species (ROS), 8-hydroxy-2-deoxyguanosine (8-OHdG), malondialdehyde (MDAs), carbonyl, sulfydryl, and GSH in cortex. Glutamate (Glu) transport was studied by measuring Glu, glutamine (Gln), mRNA, and protein of glutamate/aspartate transporter (GLAST) and glutamate transporter-1 (GLT-1). Result. (1) MeHg induced Hg accumulation, pathological injury, and apoptosis of cortex; (2) MeHg increased ROS, 8-OHdG, MDA, and carbonyl, and inhibited sulfydryl and GSH; (3) MeHg elevated Glu, decreased Gln, and downregulated GLAST and GLT-1 mRNA expression and protein levels; (4) riluzole antagonized MeHg-induced downregulation of GLAST and GLT-1 function and expression, GSH depletion, oxidative stress, pathological injury, and apoptosis obviously. Conclusion. Data indicate that MeHg administration induced oxidative stress in cortex and that riluzole could antagonize this situation through elevation of GSH synthesis by activating of GluTs.
Melastatin-like transient receptor potential channel 2 (TRPM2) is an oxidant-sensitive and cationic non-selective channel that is expressed in mammalian vascular endothelium. Here we investigated the functional role of TRPM2 channels in hydrogen peroxide (H2O2)-induced cytosolic Ca2+ ([Ca2+]i) elavation, whole-cell current increase, and apoptotic cell death in murine heart microvessel endothelial cell line H5V. A TRPM2 blocking antibody (TM2E3), which targets the E3 region near the ion permeation pore of TRPM2, was developed. Treatment of H5V cells with TM2E3 reduced the [Ca2+]i rise and whole-cell current change in response to H2O2. Suppressing TRPM2 expression using TRPM2-specific short hairpin RNA (shRNA) had similar inhibitory effect. H2O2-induced apoptotic cell death in H5V cells was examined using MTT assay, DNA ladder formation analysis, and DAPI-based nuclear DNA condensation assay. Based on these assays, TM2E3 and TRPM2-specific shRNA both showed protective effect against H2O2-induced apoptotic cell death. TM2E3 and TRPM2-specific shRNA also protect the cells from tumor necrosis factor (TNF)-α-induced cell death in MTT assay. In contrast, overexpression of TRPM2 in H5V cells resulted in an increased response in [Ca2+]i and whole-cell currents to H2O2. TRPM2 overexpression also aggravated the H2O2-induced apoptotic cell death. Downstream pathways following TRPM2 activation was examined. Results showed that TRPM2 activity stimulated caspase-8, caspase-9 and caspase-3. These findings strongly suggest that TRPM2 channel mediates cellular Ca2+ overload in response to H2O2 and contribute to oxidant-induced apoptotic cell death in vascular endothelial cells. Down-regulating endogenous TRPM2 could be a means to protect the vascular endothelial cells from apoptotic cell death.
Low-dose arginine vasopressin (AVP) has been proposed as an adjunctive vasopressor for the treatment of advanced vasodilatory shock. However, its effects on renal, hepatic, and intestinal dysfunction during sepsis remain controversial.
Fecal peritonitis was induced in 20 anesthetized, invasively monitored, mechanically ventilated female pigs. Following the time point of septic shock (defined as mean artery pressure (MAP) ≤65 mmHg), animals were randomly assigned to the following groups (n = 10): 1) a norepinephrine group with MAP between 65 and 75 mmHg; and 2) an AVP group with a constant infusion rate of 0.5 mU.kg-1.min-1.
MAP, pulmonary capillary wedge pressure, hematocrit, TNF-α, IL-6, and IL-10 were similar in the two groups during the 28-h observation period. Infusion of AVP was associated with lower total norepinephrine and fluid requirements. There was a statistically significant improvement in renal function as assessed by increased urine output and renal blood flow, and decreased serum creatinine, in the AVP group when compared with the norepinephrine group (P < 0.05). Histological analyses of the intestine, liver, and kidney showed similar light microscopical appearance of the two groups. Apoptotic cells in the liver were significantly fewer in the AVP group when compared with the norepinephrine group (P < 0.05).
An adjunctive AVP to norepinephrine infusion exhibits a favorable impact on renal function without deleterious effects on the liver and intestine in a porcine model of experimental sepsis when compared with norepinephrine infusion alone.
Blood flow; intestine; kidney; liver; sepsis; vasopressin
Porcine circovirus type 2 (PCV2) is a primary etiological agent of post-weaning multi-systemic wasting syndrome (PMWS), which is a disease of increasing importance to the pig industry worldwide. Hollow mesoporous silica nanoparticles (HMSNs) have gained increasing interest for use in vaccines.
To study the potential of HMSNs for use as a protein delivery system or vaccine carriers. HMSNs were synthesized by a sol–gel/emulsion(oil-in-water/ethanol) method, purified PCV2 GST-ORF2-E protein was loaded into HMSNs, and the resulting HMSN/protein mixture was injected into mice. The uptake and release profiles of protein by HMSNs in vitro were investigated. PCV2 GST-ORF2-E specific antibodies and secretion of IFN-γ were detected by enzyme-linked immunosorbent assays, spleen lymphocyte proliferation was measured by the MTS method, and the percentage of CD4+ and CD8+ were determined by flow cytometry.
HMSNs were found to yield better binding capacities and delivery profiles of proteins; the specific immune response induced by PCV2 GST-ORF2-E was maintained for a relatively long period of time after immunization with the HMSN/protein complex.
The findings suggest that HMSNs are good protein carriers and have high potential for use in future applications in therapeutic drug delivery.
Hollow mesoporous silica nanoparticles (HMSNs); Porcine circovirus type 2 (PCV2): ORF2; Delivery; Immunization; Mice
A seventy-six-year-old lady with poor vision of the left eye due to previous retinal detachment presented with acute visual loss of her right eye secondary to central retinal artery occlusion. Clinical examination showed a pale right optic disc, macular edema, and a cherry red spot. Optical coherence tomography done four hours after onset showed right acute cystoid macular edema and diffuse inner retinal thickening. Subsequent treatment with intravenous carbonic anhydrase inhibitor resulted in some visual improvement. Central retinal artery occlusion has been known to produce diffuse intraretinal edema instead of cystoids changes. We would like to discuss a case of acute cystoid macular edema in acute central retinal artery occlusion.
The objective of this study was to investigate the use of cationized Pleurotus eryngii polysaccharide (CPEPS) as a nonviral gene delivery vehicle to transfer plasmid DNA encoding transforming growth factor beta-1 (pTGF-β1) into mesenchymal stem cells (MSCs) in vitro. Crude P. eryngii polysaccharide was purified, and then cationized by grafting spermine onto the backbone of the polysaccharide. Agarose gel electrophoresis, transmission electron microscopy, and a Nano Sense Zetasizer (Malvern Instruments, Malvern, UK) were used to characterize the CPEPS-pTGF-β1 nanoparticles. The findings of cytotoxicity analysis showed that when the nanoparticles were formulated with a CPEPS/pTGF-β1 weight ratio ≥ 10:1, a greater gel retardation effect was observed during agarose gel electrophoresis. The CPEPS-pTGF-β1 nanoparticles with a weight ratio of 20:1, respectively, possessed an average particle size of 80.8 nm in diameter and a zeta potential of +17.4 ± 0.1 mV. Significantly, these CPEPS-pTGF-β1 nanoparticles showed lower cytotoxicity and higher transfection efficiency than both polyethylenimine (25 kDa) (P = 0.006, Student’s t-test) and LipofectamineTM 2000 (P = 0.002, Student’s t-test). Additionally, the messenger RNA expression level of TGF-β1 in MSCs transfected with CPEPS-pTGF-β1 nanoparticles was significantly higher than that of free plasmid DNA-transfected MSCs and slightly elevated compared with that of Lipofectamine 2000-transfected MSCs. Flow cytometry analysis demonstrated that 92.38% of MSCs were arrested in the G1 phase after being transfected with CPEPS-pTGF-β1 nanoparticles, indicating a tendency toward differentiation. In summary, the findings of this study suggest that the CPEPS-pTGF-β1 nanoparticles prepared in this work exhibited excellent transfection efficiency and low toxicity. Therefore, they could be developed into a promising nonviral vector for gene delivery in vitro.
nonviral gene vector; transfection; plasmid DNA; spermine
Mast cells (MCs) deficient rats (Ws/Ws) were used to investigate the roles of MCs in visceral hyperalgesia. Ws/Ws and wild control (+/+) rats were exposed to T. spiralis or submitted to acute cold restraint stress (ACRS). Levels of proteinase-activated receptor 2 (PAR2) and nerve growth factor (NGF) were determined by immunoblots and RT-PCR analysis, and the putative signal pathways including phosphorylated extracellular-regulated kinase (pERK1/2) and transient receptor potential vanilloid receptor 1 (TRPV1) were further identified. Visceral hyperalgesia triggered by ACRS was observed only in +/+ rats. The increased expression of PAR2 and NGF was observed only in +/+ rats induced by T. spiralis and ACRS. The activation of pERK1/2 induced by ACRS occurred only in +/+ rats. However, a significant increase of TRPV1 induced by T. spiralis and ACRS was observed only in +/+ rats. The activation of PAR2 and NGF via both TRPV1 and pERK1/2 signal pathway is dependent on MCs in ACRS-induced visceral hyperalgesia rats.
The purpose of this study was to develop a sustained drug-release model for water-soluble drugs using silica nanoparticles.
Hollow-type mesoporous silica nanoparticles (HMSNs) were prepared using Na2CO3 solution as the dissolution medium for the first time. The water-soluble compound, silybin meglumine, was used as the model drug. The Wagner–Nelson method was used to calculate the in vivo absorption fraction.
The results of transmission electron microscopy and nitrogen adsorption revealed that the empty HMSNs had uniformly distributed particles of size 50–100 nm, a spherical appearance, a large specific surface area (385.89 ± 1.12 m2/g), and ultralow mean pore size (2.74 nm). The highly porous structure allowed a large drug-loading rate (58.91% ± 0.39%). In 0.08 M Na2CO3 solution, silybin meglumine-loaded HMSNs could achieve highly efficacious and long-term sustained release for 72 hours in vitro. The results of in vitro–in vivo correlation revealed that HMSNs in 0.08 M Na2CO3 solution had a correlation coefficient R2 value of 0.9931, while those of artificial gastric juice and artificial intestinal juice were only 0.9287 and 0.7689, respectively.
The findings of in vitro–in vivo correlation indicate that HMSNs together with Na2CO3 solution could achieve an excellent linear relationship between in vitro dissolution and in vivo absorption for 72 hours, leading to a promising model for sustained release of water-soluble drugs.
hollow-type mesoporous silica nanoparticle; silybin meglumine; in vitro dissolution; in vivo absorption; in vitro-in vivo correlation
AIM: To assess the safety of Bifidobacterium longum (B. longum) JDM301 based on complete genome sequences.
METHODS: The complete genome sequences of JDM301 were determined using the GS 20 system. Putative virulence factors, putative antibiotic resistance genes and genes encoding enzymes responsible for harmful metabolites were identified by blast with virulence factors database, antibiotic resistance genes database and genes associated with harmful metabolites in previous reports. Minimum inhibitory concentration of 16 common antimicrobial agents was evaluated by E-test.
RESULTS: JDM301 was shown to contain 36 genes associated with antibiotic resistance, 5 enzymes related to harmful metabolites and 162 nonspecific virulence factors mainly associated with transcriptional regulation, adhesion, sugar and amino acid transport. B. longum JDM301 was intrinsically resistant to ciprofloxacin, amikacin, gentamicin and streptomycin and susceptible to vancomycin, amoxicillin, cephalothin, chloramphenicol, erythromycin, ampicillin, cefotaxime, rifampicin, imipenem and trimethoprim-sulphamethoxazol. JDM301 was moderately resistant to bacitracin, while an earlier study showed that bifidobacteria were susceptible to this antibiotic. A tetracycline resistance gene with the risk of transfer was found in JDM301, which needs to be experimentally validated.
CONCLUSION: The safety assessment of JDM301 using information derived from complete bacterial genome will contribute to a wider and deeper insight into the safety of probiotic bacteria.
Bifidobacterium longum; Safety assessment; Genome; Antibiotic resistance; Harmful metabolite; Virulence factor
Coxiella burnetii is the etiological agent of human Q fever. In this study, adaptive transfer of mouse bone marrow–derived dendritic cells (BMDCs) stimulated with C. burnetii antigen, phase I whole-cell antigen (PIAg), lipopolysaccharide (LPS)–removed PIAg (PIIAg), protein antigen Com1, or SecB significantly reduced coxiella burden in recipient mice compared with control mice. Mice that received PIIAg-pulsed BMDCs displayed substantially lower coxiella burden than recipient mice of PIAg-pulsed BMDCs after C burnetii challenge. The protection offered by the antigen-activated BMDCs was correlated with the increased proliferation of helper T (TH) TH1 CD4+ cells, preferential development of TH17 cells, and impaired expansion of regulatory T lymphocytes. Our results suggest that PIIAg is far superior to PIAg in activating BMDCs to confer protection against C. burnetii in vivo, whereas Com1 and SecB are protective antigens because Com1- or SecB-pulsed BMDCs confer partial protection.