MicroRNA-9 promotes the proliferation, migration, and invasion of breast cancer cells via down-regulating FOXO1.

PURPOSE: The objective of the study was to investigate the role of microRNA-9 (miR-9) targeting forkhead box O1 (FOXO1) in the proliferation, migration, and invasion of breast cancer cells. METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to determine the expressions of miR-9 and FOXO1 mRNA in breast cancer tissues, normal breast tissues, breast cancer cell lines, and normal breast epithelial cells. After the up-regulation of miR-9 expression, qRT-PCR and Western blotting were used to determine the expression of FOXO1. The luciferase reporter gene assay was used to validate the target gene. The CCK-8 assay, scratch-wound healing assay, and Transwell invasion assay were used to investigate the changes in the proliferation, migration, and invasion of breast cancer cells, respectively. RESULTS: MicroRNA-9 expression was significantly up-regulated in breast cancer tissues and breast cancer cell lines when compared with normal breast tissues and normal breast epithelial cells (both P < 0.05). FOXO1 mRNA and protein expressions were substantially down-regulated in breast cancer tissues and breast cancer cell lines when compared with normal breast tissues and normal breast epithelial cells (both P < 0.05). There can be a negative correlation between miR-9 and FOXO1 mRNA in breast cancer. Luciferase reporter gene assay indicated that miR-9 can down-regulate FOXO1 expression at a post-transcriptional level through binding specifically to FOXO1 3'UTR. The results of CCK-8 assay, scratch-wound healing assay, and Transwell invasion assay revealed that the inhibition of miR-9 can suppress MCF7 cell proliferation, migration, and invasion. Additionally, the expression of miR-9 increased significantly whilst that of FOXO1 decreased substantially as the disease progressed (P < 0.05). CONCLUSIONS: Our study provides evidence that miR-9 can promote the proliferation, migration, and invasion of breast cancer cells via down-regulating FOXO1.

Peripheral Blood Gene Expression Changes Associated With Primary Graft Dysfunction After Lung Transplantation.

Recipient responses to primary graft dysfunction (PGD) after lung transplantation may have important implications to the fate of the allograft. We therefore evaluated longitudinal differences in peripheral blood gene expression in subjects with PGD. RNA expression was measured throughout the first transplant year in 106 subjects enrolled in the Clinical Trials in Organ Transplantation-03 study using a panel of 100 hypothesis-driven genes. PGD was defined as grade 3 in the first 72 posttransplant hours. Eighteen genes were differentially expressed over the first year based on PGD development, with significant representation from innate and adaptive immunity genes, with most differences identified very early after transplant. Sixteen genes were overexpressed in the blood of patients with PGD compared to those without PGD within 7 days of allograft reperfusion, with most transcripts encoding innate immune/inflammasome-related proteins, including genes previously associated with PGD. Thirteen genes were underexpressed in patients with PGD compared to those without PGD within 7 days of transplant, highlighted by T cell and adaptive immune regulation genes. Differences in gene expression present within 2 h of reperfusion and persist for days after transplant. Future investigation will focus on the long-term implications of these gene expression differences on the outcome of the allograft.

The Perioperative Lung Transplant Virome: Torque Teno Viruses Are Elevated in Donor Lungs and Show Divergent Dynamics in Primary Graft Dysfunction.

Primary graft dysfunction (PGD) is a principal cause of early morbidity and mortality after lung transplantation, but its pathogenic mechanisms are not fully clarified. To date, studies using standard clinical assays have not linked microbial factors to PGD. We previously used comprehensive metagenomic methods to characterize viruses in lung allografts >1 mo after transplant and found that levels of Anellovirus, mainly torque teno viruses (TTVs), were significantly higher than in nontransplanted healthy controls. We used quantitative polymerase chain reaction to analyze TTV and shotgun metagenomics to characterize full viral communities in acellular bronchoalveolar lavage from donor organs and postreperfusion allografts in PGD and non-PGD lung transplant recipient pairs. Unexpectedly, TTV DNA levels were elevated 100-fold in donor lungs compared with healthy adults (p = 0.0026). Although absolute TTV levels did not differ by PGD status, PGD cases showed a smaller increase in TTV levels from before to after transplant than did control recipients (p = 0.041). Metagenomic sequencing revealed mainly TTV and bacteriophages of respiratory tract bacteria, but no viral taxa distinguished PGD cases from controls. These findings suggest that conditions associated with brain death promote TTV replication and that greater immune activation or tissue injury associated with PGD may restrict TTV abundance in the lung.

Effects of dietary fermented garlic on the growth performance, relative organ weights, intestinal morphology, cecal microflora and serum characteristics of broiler chickens

The present study was conducted to evaluate the effects of feeding broilers with garlic fermented by Leuconostoc citreum SK2556. A total of 250 male broiler chicks was randomly housed into 25 floor pens. Five dietary treatments with five replicates of 10 chicks each (n=50 chicks/treatment). A corn and soybean meal based diet was used as the control diet (NC). The experimental diets were formulated by mixing the basal diet either with antibiotics (10 ppm; PC) or fermented garlic (FG) at the concentrations of 0.1% (FG1), 0.3% (FG3) or 0.5% (FG5) in diets. Daily weight gain, feed intake, and feed:gain ratio were not affected by any of the dietary treatments. Average daily gain on day 21 linearly increased (p= 0.024) with increasing FG levels. The relative weight of the bursa of Fabricius showed a progressive decline with increasing the FG levels. Jejunal villus height was not influenced by dietary treatments. Villus width linearly decreased as FG levels increased (p= 0.17). Jejunal crypt depth was significantly lower (p < 0.05) in the FG1 and FG3 groups compared with the NC group. Villus height: crypt depth ratio linearly increased (p= 0.018) with increasing FG levels. The population of cecal microflora was not altered by dietary treatments. Broiler chickens fed the FG5 diet exhibited (p < 0.05) higher blood levels of total protein and cholesterol compared with those fed the NC diet. Collectively, the results show that dietary FG marginally affected growth performance, especially during the first days rearing, improved intestinal morphology, and altered blood characteristics of broiler chickens.

Effects of dietary fermented garlic on the growth performance, relative organ weights, intestinal morphology, cecal microflora and serum characteristics of broiler chickens

The present study was conducted to evaluate the effects of feeding broilers with garlic fermented by Leuconostoc citreum SK2556. A total of 250 male broiler chicks was randomly housed into 25 floor pens. Five dietary treatments with five replicates of 10 chicks each (n=50 chicks/treatment). A corn and soybean meal based diet was used as the control diet (NC). The experimental diets were formulated by mixing the basal diet either with antibiotics (10 ppm; PC) or fermented garlic (FG) at the concentrations of 0.1% (FG1), 0.3% (FG3) or 0.5% (FG5) in diets. Daily weight gain, feed intake, and feed:gain ratio were not affected by any of the dietary treatments. Average daily gain on day 21 linearly increased (p= 0.024) with increasing FG levels. The relative weight of the bursa of Fabricius showed a progressive decline with increasing the FG levels. Jejunal villus height was not influenced by dietary treatments. Villus width linearly decreased as FG levels increased (p= 0.17). Jejunal crypt depth was significantly lower (p < 0.05) in the FG1 and FG3 groups compared with the NC group. Villus height: crypt depth ratio linearly increased (p= 0.018) with increasing FG levels. The population of cecal microflora was not altered by dietary treatments. Broiler chickens fed the FG5 diet exhibited (p < 0.05) higher blood levels of total protein and cholesterol compared with those fed the NC diet. Collectively, the results show that dietary FG marginally affected growth performance, especially during the first days rearing, improved intestinal morphology, and altered blood characteristics of broiler chickens.(AU)

Quantitative candidate gene association studies of metabolic traits in Han Chinese type 2 diabetes patients.

Recent genome-wide association studies have identified many loci associated with type 2 diabetes mellitus (T2DM), hyperuricemia, and obesity in various ethnic populations. However, quantitative traits have been less well investigated in Han Chinese T2DM populations. We investigated the association between candidate gene single nucleotide polymorphisms (SNPs) and metabolic syndrome-related quantitative traits in Han Chinese T2DM subjects. Unrelated Han Chinese T2DM patients (1975) were recruited. Eighty-six SNPs were genotyped and tested for association with quantitative traits including lipid profiles, blood pressure, body mass index (BMI), serum uric acid (SUA), glycated hemoglobin (HbA1c), plasma glucose [fasting plasma glucose (FPG)], plasma glucose 120 min post-OGTT (P2PG; OGTT = oral glucose tolerance test), and insulin resistance-related traits. We found that CAMTA1, ABI2, VHL, KAT2B, PKHD1, ESR1, TOX, SLC30A8, SFI1, and MYH9 polymorphisms were associated with HbA1c, FPG, and/or P2PG; GCK, HHEX, TCF7L2, KCNQ1, and TBX5 polymorphisms were associated with insulin resistance-related traits; ABCG2, SLC2A9, and PKHD1 polymorphisms were associated with SUA; CAMTA1, VHL, KAT2B, PON1, NUB1, SLITRK5, SMAD3, FTO, FANCA, and PCSK2 polymorphisms were associated with blood lipid traits; CAMTA1, SPAG16, TOX, KCNQ1, ACACB, and MYH9 polymorphisms were associated with blood pressure; and UBE2E3, SPAG16, SLC2A9, CDKAL1, CDKN2A/B, TCF7L2, SMAD3, and PNPLA3 polymorphisms were associated with BMI (all P values <0.05). Some of the candidate genes were associated with metabolic and anthropometric traits in T2DM in Han Chinese. Although none of these associations reached genome-wide significance (P < 5 x 10(-8)), genes and loci identified in this study are worthy of further replication and investigation.

MYH9 gene polymorphisms may be associated with cerebrovascular blood flow in patients with type 2 diabetes.

Genetic factors play an important role in type 2 diabetes (T2D) complications. Alteration of cerebrovascular blood flow (CBF) is a direct result of cerebrovascular diseases. However, few studies have reported the role of genetics on CBF in patients with T2D. We investigated whether single-nucleotide polymorphisms (SNPs) in metabolic disease genes are associated with CBF in patients with T2D. CBF velocities of CBF were measured in 337 Han Chinese patients with T2D using transcranial Doppler sonography, with 54 cerebrovascular blood flow parameters documented. Fifty-two SNPs from 31 metabolic disease candidate genes were genotyped in patients. Quantitative allelic association and haplotype analyses were performed for candidate gene SNPs and CBF phenotypes. Spearman correlation was used to determine the relationship between CBF parameters and basic clinical characteristics, particularly, body mass index, lipids, fibrinogen, and GHbA1c. MYH9 gene SNPs (rs875726 and rs735853) may be associated with the peak velocity of the right-middle cerebral artery. SNPs rs875726, rs2009930, and rs375246 of the MYH9 gene may be associated with the mean velocity of the right-anterior and posterior cerebral artery. The haplotype G-C-A (rs2239782-rs3752462- rs2269532) of MYH9 may be associated with CBF. MYH9 gene polymorphisms may be associated with multiple CBF phenotypes in Chinese patients with T2D. However, whether MYH9 is a candidate gene for cerebrovascular diseases in Chinese patients with T2D remains unknown.

Establishment of cytochrome P450 3A4 and glutathione S-transferase A1-transfected human hepatoma cell line and functional analysis.

This study aimed to enhance the drug metabolism function of the human hepatoma cell line C3A and to explore the related significance for patients with severe liver disease. The important liver phase I and phase II drug metabolism enzymes, cytochrome P450 3A4 (CYP 3A4) and glutathione S-transferase A1 (GST A1), were constructed into a double expression vector and then transfected into C3A cells. Furthermore, in order to increase the expression of CYP 3A4 and GST A1, they were optimized according to human optimal codons. Another double-expression vector, pBudCE4.1-optimized CYP 3A4-optimized GST A1, was constructed and then transfected into C3A to establish a stable cell line. The drug metabolism function of C3A was evaluated. Sequence determination and analysis results showed that the recombinant plasmid pBudCE4.1-CYP 3A4-GST A1 met the application standard and its transfection was successful. The expression and activity of CYP 3A4 and GST A1 in unoptimized C3A cells were higher than those in blank C3A cells. Unoptimized C3A had a better drug metabolism function. Although some C3A cells transfected with pBudCE4.1-optimized CYP 3A4-optimized GST A1 survived, they grew slowly, and were therefore not applicable in clinical practice. Unoptimized C3A is superior to blank C3A in drug metabolism, and could be applied in the bioartificial liver support system as a new material.

Early detection of human immunodeficiency virus on dried blood spot specimens: sensitivity across serial specimens. Women and Infants Transmission Study Group.

OBJECTIVE: To evaluate the use of dried blood spot (DBS) specimens and the early diagnostic value of the polymerase chain reaction (PCR) for detection of the human immunodeficiency virus (HIV) in DBS specimens collected at predefined age intervals from a large cohort of U.S. infants at risk of congenital or perinatal HIV infection. DESIGN: We assayed available DBS specimens (n = 272) obtained during the first 4 months of life from 144 infants (41 infected, 103 uninfected) born to HIV-infected mothers enrolled in the Women and Infants Transmission Study. The DBS PCR results were compared with infant HIV infection status, PCR on liquid blood, and viral culture results. Analyses also included sensitivity and specificity of assay as related to the age of the infant when the specimen was obtained. RESULTS: The DBS specimen PCR results were concordant with results from liquid blood specimens and with results from viral culture. The DBS PCR was highly specific for all age groups. Sensitivity in detecting HIV infection status rapidly increased during the first month of life, from 19% (5/26) by 1 week to 96% (25/26) by 1 month of age. Specimens obtained on the day of birth or the next day were the least likely to have detectable HIV DNA. CONCLUSIONS: The PCR assay of DBS specimens is a reliable tool for the early diagnosis of HIV infection and has important advantage over that of liquid blood DNA PCR and viral culture. These advantages include a lower volume of blood required for testing, increased safety, and ease of storage or transport of specimens. Thus DBS PCR is a useful test for clinical and epidemiologic tracking of infants at risk of HIV infection.

Lower limb bypass for intermittent Claudication; is it really worth the risk

Intra-inguinal bypass for intermittent claudication is a safe option with low morbidity and mortality (AU)