Effects of iodine methionine on boar sperm quality during liquid storage at 17°C.

Microbial environment is one of the important factors that affect the quality of preserved semen. Iodine methionine (IM), participating in the production and activation of metabolic enzymes, is a new type of amino acid chelate. To date, there has been no report to evaluate the effects of IM on boar semen preservation at 17°C. This study was designed to investigate the effects of IM on boar sperm quality and reproductive performance during liquid storage at 17°C and its antibacterial effect. Semen samples collected from six Yorkshire boars were diluted with basic liquid containing different concentrations of IM (0, 20, 40, 80, 160 and 320 µM). Subsequently, sperm motility, plasma membrane integrity and acrosome integrity were determined. After 6 days of preservation, the difference in microbial composition between control group and 80 µM IM group was compared using 16S rDNA sequencing, and the effects of IM on reproductive performance were also compared and analysed between the two groups. The results demonstrated that 20, 40 and 80 µM IM improved boar sperm motility, plasma membrane integrity and acrosome integrity. 80 µM IM was the optimum concentration. Conversely, 160 and 320 µM IM resulted in deleterious consequences to boar sperm quality compared to the control group and other treatment groups (p < .05). After 6 days of preservation, sperm motility, plasma membrane integrity and acrosome integrity were 56.0%, 51.8% and 59.4%, respectively. There was no significant difference in non-return rate between the two groups (p > .05). But the litter size of 80 µM IM group was significantly higher than that of control group (p < .05). 80 µM IM inhibited proliferation of the phylum Proteobacteria and the genus Staphylococcus as well as Pseudomonas (p < .05). Further studies are required to understand the antibacterial mechanism of IM in liquid-preserved boar semen.

Association between oral health habits and dental caries among children in Pyongyang, Democratic People's Republic of Korea.

AIM: To evaluate the self-reported oral health habits and their association with the occurrence of dental caries among children in Pyongyang, Democratic People's Republic of Korea (DPRK), after 6 years of activities under the auspices of the Children's Oral Health Promotion Programme (COHPP). METHODS: The data were collected in September 2013 in two of the most central districts of Pyongyang City, DPRK. The sample consisted of 492 children aged 10 and 13 years who had participated in the COHPP for 6 years. The children filled in a self-completed, structured questionnaire on oral health habits and were examined clinically by a dentist. The differences in mean (SD) number of decayed primary (dt) and permanent teeth (DT) and their sum (dt + DT) subdivided according to genders, age groups, districts and self-reported oral health habits were evaluated using Mann-Whitney U-test. The associations between self-reported oral health habits and the occurrence of dental caries were evaluated with chi-square test and logistic regression analyses. RESULTS: The school-aged children commonly reported healthy oral hygiene habits but sweet snacks were commonly used. The occurrence of dental caries associated statistically significantly with the frequency of sweet snacking (p=0.011) but not with the frequency of tooth brushing (p=0.725) or the use of water for thirst instead of sugary beverages (p=0.189). CONCLUSION: A more effective promotion of healthy dietary habits with innovative approaches and close collaboration with different social actors will be needed in future.

Comparison of two preventive interventions on dental caries among children in Democratic People's Republic of Korea.

OBJECTIVES: The aim was to compare the change in dental caries status in two different intervention groups of the Children's Oral Health Promotion Programme (COHPP). METHODS: A longitudinal study among 500 children who had participated into the COHPP for 6 years was conducted in Pyongyang, Democratic People's Republic of Korea (DPRK). Children in Group I received intensified school-based intervention and were clinically examined at the age of 7 years in 2007 (n = 250), 10 years in 2010 (n = 250) and 13 years in 2013 (n = 242). Children in Group II (n = 250) joined the programme at the age of 4 years in kindergarten in 2007, were provided with early preschool-based intervention and were clinically examined at the age of 7 years in 2010 and 10 years in 2013. RESULTS: Both the prevalence and the mean number of dt + DT decreased significantly in both groups during the follow-up. This was due to decrease in the number of dt, whereas the number of DT remained relatively constant. Poisson regression showed that the association between the group status and the change in the number of dt + DT was statistically significant when adjusted for gender but disappeared when the school was included in the analysis. CONCLUSIONS: The decrease in dental caries may be partly due to the exfoliation of deciduous teeth and dental treatment received. However, the study gave some reference emphasizing the early starting of the prevention.

PU.1 antisense lncRNA against its mRNA translation promotes adipogenesis in porcine preadipocytes.

Antisense long non-coding RNAs (AS lncRNAs) play important roles in refined regulation of animal gene expression. However, their functions and molecular mechanisms for domestic animal adipogenesis are largely unknown. Here, we found a novel AS lncRNA transcribed from the porcine PU.1 gene (also known as SPI1) by strand-specific RT-PCR. Results showed that PU.1 AS lncRNA was expressed and generally lower than the level of PU.1 mRNA in porcine subcutaneous adipose, heart, liver, spleen, lympha, skeletal muscle and kidney tissues. We further found that the levels of PU.1 mRNA and PU.1 protein were significantly lower in subcutaneous and intermuscular adipose than in mesenteric and greater omentum adipose, whereas the levels of PU.1 AS lncRNA showed no difference in porcine adipose tissues from four different parts of the body. During porcine adipogenesis, levels of PU.1 mRNA increased at day 2 and then gradually decreased. Meanwhile, PU.1 AS lncRNA exhibited an expression trend similar to PU.1 mRNA but sharply decreased after day 2. Interestingly, PU.1 protein level rose during differentiation. In addition, at day 6 after differentiation, knockdown of endogenous PU.1 promoted adipogenesis, whereas knockdown of endogenous PU.1 AS lncRNA had the opposite effect. Moreover, peroxisome proliferator-activated receptor gamma (PPARG) and fatty acid synthase (FASN) were significantly upregulated in the PU.1 shRNA treatment group (P < 0.05), whereas they were downregulated in the PU.1 AS shRNA treatment group (P < 0.05). Adipose triglyceride lipase [ATGL; also known as patatin-like phospholipase domain containing 2 (PNPLA2)] and hormone-sensitive lipase [HSL; also known as lipase, hormone-sensitive (LIPE)] contrasted with PPARG and FASN. Finally, the PU.1 mRNA/PU.1 AS lncRNA duplex was detected by an endogenous ribonuclease protection assay combined with RT-PCR. Based on the above results, we suggest that PU.1 AS lncRNA (vs. its mRNA translation) promotes adipogenesis through the formation of a sense-antisense RNA duplex with PU.1 mRNA.

Effects of bovine serum albumin on boar sperm quality during liquid storage at 17°C.

This study aimed to investigate the effects of bovine serum albumin (BSA) on boar sperm quality during liquid storage at 17°C. Boar semen samples were collected and diluted with Modena containing different concentrations (0, 1, 2, 3, 4, 5 and 6 g/l) of BSA, and sperm motility, plasma membrane integrity, acrosome integrity, total antioxidative capacity (T-AOC) activity and malondialdehyde (MDA) content were measured and analysed. The results showed that Modena supplemented with 3, 4 and 5 g/l BSA could improve boar sperm motility, effective survival time and plasma membrane integrity (p < 0.05), decrease MDA content (p < 0.05), while no statistical difference was observed for sperm acrosome integrity and T-AOC activity among these three groups (p > 0.05). The semen sample diluted with Modena containing 4 g/l BSA could achieve optimum effect, and sperm survival time was 7.5 days. After 7 days preservation, sperm motility, plasma membrane integrity and acrosome integrity were 54%, 49% and 78%, respectively. T-AOC activity and MDA content were 1.03 U/ml and 17.5 nmol/ml, respectively. In conclusion, Modena supplemented with BSA reduced the oxidative stress and improved the sperm quality of boar semen during liquid storage at 17°C, and 4 g/l BSA was the optimum concentration. Further studies are required to obtain more concrete results on the determination of antioxidant capacities of BSA in liquid preserved boar semen.

Effect of functional nuclear factor-kappaB genetic polymorphisms on hepatitis B virus persistence and their interactions with viral mutations on the risk of hepatocellular carcinoma.

BACKGROUND: Nonresolving inflammation and viral mutations are important in hepatitis B virus (HBV)-induced hepatocarcinogenesis. However, the effects of genetic polymorphisms affecting nuclear factor-kappaB (NF-κB) on HBV persistence and generation of hepatocellular carcinoma (HCC)-related HBV mutations remain unknown. PATIENTS AND METHODS: rs28362491 (NFKB1 -94Ins > Del), rs2233406 (NFKBIA -826C > T), rs3138053 (NFKBIA -881A > G), and rs696 (NFKBIA +2758G > A) were genotyped in 1342 healthy controls, 327 HBV-clearance subjects, and 3976 HBV-positive subjects including 1495 HCC patients, using quantitative PCR. HBV mutations were determined by sequencing. The NFKBIA promoter activity was assessed by transient transfection. Multiplicative interactions of the polymorphisms and viral mutations were assessed by multivariate logistic regression. RESULTS: Compared with HBV-clearance subjects, rs2233406 (CT versus CC) and rs3138053 (AG or AG + GG versus AA) significantly decreased HBV persistence, especially in the genotype B HBV-infected subjects. In the genotype C HBV-infected subjects, rs2233406 variant genotypes were significantly associated with an increased risk of HCC [CT versus CC: age-, gender-adjusted odds ratio (AOR), 1.33; 95% confidence interval (CI) 1.01-1.75 in training set and AOR, 1.59; 95% CI 1.01-2.52 in validation set] compared with HCC-free HBV-infected subjects and significantly increased the frequencies of HCC-related HBV mutations (A1762T/G1764A, T1753V, preS1 start codon mutation, and preS deletion); rs28362491 (Del/Del or Ins/Del + Del/Del versus Ins/Ins) significantly increased the frequency of A1762T/G1764A and reduced the frequency of preS2 start codon mutation. The variant genotypes impaired NFKBIA promoter activity in hepatic cells. The interaction of rs2233406 variant genotypes (CT + TT versus CC) with A1762T/G1764A significantly increased HCC risk in genotype C HBV-infected subjects, with AOR of 2.61 (95% CI 1.09-6.26). CONCLUSION: Genetic polymorphisms improving NF-κB activity contribute to genotype B HBV clearance. The rs2233406 variant genotypes significantly increase HCC risk, possibly via facilitating immune selection of the HBV mutations. The host-virus interactions are important in identifying HBV-infected subjects who are more likely to develop HCC.

Correlation of forkhead box transcription factor O1 and myosin heavy chain isoforms in porcine skeletal muscle.

We examined the expression of myosin heavy chain (MyHC) isoforms and forkhead box transcription factor O1 (FoxO1) in porcine soleus and extensor digitorum longus (EDL) muscles to clarify the correlation of FoxO1 and the relative abundance of transcripts of MyHC isoforms. Soleus muscle was found to be redder than EDL muscles in pigs, and immunohistochemical fast MyHC staining showed more oxidative type I fibers compared to EDL. qRT-PCR quantification of MyHC isoforms I, IIa, IIx, and IIb showed that expression of MyHC I and MyHC IIa mRNAs was much higher, whereas expression of MyHC IIx and MyHC IIb mRNAs was much lower in porcine soleus muscle compared to EDL muscle. Expression of FoxO1 mRNA and p-FoxO1 protein was significantly more abundant in porcine soleus muscle compared to EDL muscle. The expression of phosphorylated FoxO1 (p-FoxO1) was positively correlated with the expression of MyHC I (R = 0.9747, P < 0.01) and negatively correlated with the expression of MyHC IIx (R = -0.9963, P < 0.01) and MyHC IIb (R = -0.9834, P < 0.01). Taken together, these results suggested that FoxO1 may play a pivotal role in the determination of muscle fiber type.

Characteristics of intrahepatic cholangiocarcinoma in patients with hepatitis B virus infection: clinicopathologic study of resected tumours.

Recent efforts suggest an aetiological role of hepatitis B virus (HBV) infection in intrahepatic cholangiocarcinoma (ICC). The purpose of this study was to clarify the clinicopathologic characteristics and surgical outcomes of patients with HBV-associated ICC. All patients with chronic HBV infection were identified from a database of patients with ICC that underwent surgical resection between 1 January 2005 and 31 December 2006. Their clinicopathologic and survival characteristics were compared with ICC patients without chronic HBV infection. The age of the HBV-associated ICC patients tend to be younger than that of ICC patients without chronic HBV infection. HBV-associated ICC patients tend to have higher abnormal α-fetoprotein levels and lower abnormal serum carbohydrate antigen19-9 (CA19-9), r-glutamyltransferase (r-GT) and alkaline phosphatase levels. The pathologic features of the resected specimens revealed that HBV-associated ICC patients tended to be of the mass-forming type have a lower prevalence of lymphatic involvement and poorer tumour differentiation, and a higher prevalence of capsule formation and liver cirrhosis. Patients with HBV-associated ICC had a significantly better survival than patients without chronic HBV infection. The clinicopathological features of HBV-associated ICC patients showed significant differences from ICC patients without HBV infection. These tumours are characterized by the mass-forming growth pattern and appeared to have a more favourable prognosis.

Transcriptome profile analysis of porcine adipose tissue by high-throughput sequencing.

Novel high-throughput deep sequencing technology has dramatically changed the study of the functional complexity of transcriptomes. Here, we report the first use of this technology for analysing the wide range of transcriptional changes in porcine adipose tissue from different breeds and different growth phases in a model of obesity. Digital gene expression (DGE) data sets were instrumental to verifying the predicted gene models. We obtained a sequencing depth of over 3 million tags per sample (lean, obese-a and obese-b). Tag mapping indicated expression of more than 76% of all genes represented in three transcript databases. We found the expression level of 1596 genes was significantly different between lean and obese-a library (P < 0.01). Among them, we found 84 genes expressed only in the obese-a library and 95 genes expressed only in the lean library. Moreover, the expression of 4403 genes was found to be remarkably different between the obese-a and obese-b library (P < 0.01); 642 of these were only expressed in obese-a, and 618 were only expressed in obese-b. When mapping to genes, it was found that the sense transcripts account for 67.42%, 68.65% and 66.61% of all clean tags in the lean, obese-a and obese-b libraries respectively. By comparison, the ratio of sense to antisense mapping of the total number of tags was approximately 6:1 for all libraries. This suggests that transcriptional regulation on the sense strand has a major role in adipose deposition, although a high number of antisense mapping events were also detected. We anticipated more than 20,000 different novel tags to be localized to the porcine genome. Among them, 799 different clean tags with a copy number of more than 1000 were detected. In conclusion, our deep sequencing analysis revealed a high degree of transcriptional complexity in the regulatory mechanisms of adipogenesis and resulted in the discovery and validation of new gene products in porcine adipose tissue.

PGC-1ß modulates the expression of genes involved in mitochondrial function and adipogenesis during preadipocyte differentiation.

This study determined the expression of genes involved in mitochondrial function and adipogenesis at mRNA and protein levels by transfecting rat differentiating preadipocytes with siRNA/Lipofectamine complex and pcDNA-PGC-1ß (peroxisome proliferator-activated receptor-γ coactivator-1ß)/Lipofectamine complex, respectively, to further elucidate the role of PGC-1ß in white preadipocyte differentiation. The results showed that the transfection of PGC-1ß siRNA inhibited the expressions of mitochondrial genes malate dehydrogenase, carnitine palmitoyltransferase 1, nuclear respiratory factor 1, ATP synthesis, adipocyte differentiation key transcription factor peroxisome proliferator-activated receptor-γ, sterol regulatory element binding protein 1c and fatty acid synthetase, whereas the triglyceride synthesis was retarded (p < 0.05). Furthermore, overexpression of PGC-1ß up-regulated the expressions of adipogenic and mitochondrial biosynthetic marker genes and promoted triglyceride accumulation during 3T3-L1 adipocyte differentiation. These observations suggest that PGC-1ß modulates the expression of mitochondrial function and adipogenesis-related genes and affects white preadipocyte differentiation.

Upregulation of KCNMA1 facilitates the reversal effect of verapamil on the chemoresistance to cisplatin of esophageal squamous cell carcinoma cells.

OBJECTIVE: This study aimed to investigate the reversal effect of verapamil (VER) on the chemoresistance to cisplatin of esophageal squamous cell carcinoma (ESCC) cells. PATIENTS AND METHODS: The reversal effect of VER on cisplatin resistance in ESCC cells was evaluated via CCK-8 assay, colony formation assessment, and flow cytometry. The key genes that mediate this effect were screened via high-throughput transcriptome se¬quencing. The mRNA and protein expression levels of potassium calcium-activated channel subfamily M alpha 1 (KCNMA1) in ESCC cells were examined via quantitative real-time PCR and Western blot analysis, respectively. The protein expressions of KCNMA1 in tissue samples from patients with either positive or negative responses to the therapeutic regimen of VER were determined via immunohistochemistry assay. Cell models with KCNMA1 knockdown and overexpression were es¬tablished to examine the role of KCNMA1 in mediating the reversal effect of VER on the chemoresistance to cisplatin of ESCC cells. RESULTS: Results revealed that VER significantly decreased the 50% inhibitory concentration of cisplatin, inhibited colony formation, and induced apoptosis in ESCC cells. The curative effects of VER combined with chemotherapeutic drugs in KCNMA1-positive patients were better than those in KCNMA1-negative patients. KCNMA1 upregulation enhanced the reversal effect of VER on the chemoresistance to cisplatin of ESCC cells. CONCLUSIONS: KCNMA1 facilitated the reversal effect of VER on cisplatin resistance in ESCC cells.

Next generation tools for high-throughput promoter and expression analysis employing single-copy knock-ins at the Hprt1 locus.

We have engineered a set of useful tools that facilitate targeted single copy knock-in (KI) at the hypoxanthine guanine phosphoribosyl transferase 1 (Hprt1) locus. We employed fine scale mapping to delineate the precise breakpoint location at the Hprt1(b-m3) locus allowing allele specific PCR assays to be established. Our suite of tools contains four targeting expression vectors and a complementing series of embryonic stem cell lines. Two of these vectors encode enhanced green fluorescent protein (EGFP) driven by the human cytomegalovirus immediate-early enhancer/modified chicken beta-actin (CAG) promoter, whereas the other two permit flexible combinations of a chosen promoter combined with a reporter and/or gene of choice. We have validated our tools as part of the Pleiades Promoter Project (http://www.pleiades.org), with the generation of brain-specific EGFP positive germline mouse strains.

Thermal effect on piezoelectric stick-slip actuator systems.

The piezoelectric stick-slip (PZT-SS) actuator is known to achieve motion with a theoretically unlimited range yet high resolution (several nanometers). In this type of actuator, friction plays an active role in producing a meaningful stick-slip motion. However, friction is a source of heat which may cause significant temperature rise, affecting the dynamic performance of the actuator. Our study aimed to measure temperature rise in the stick-slip motion and to understand whether such a rise could significantly affect the displacement of the stick-slip motion. In this study, a temperature measurement system was developed using the off-the-shelf components, with which the temperature rise up to 0.436 degrees C was successfully measured on a proprietary PZT-SS actuator. The experiment further shows that the temperature rise affects the displacement of the actuator when operating voltage is at the low end (approximately 6 V). Therefore, one of the design recommendations for such an actuator system is that the operating voltage should be at the high end (approximately 30 V). The study also measured the temperature rise (approximately 0.263 degrees C after the system worked for 6300 s) at the friction interface due to the piezoelectric element which is a part of the whole PZT-SS actuator. This means that temperature rise is due to both the friction at two interacting surface and the operation of the piezoelectric element.

Acute and chronic cold exposure differentially affects the browning of porcine white adipose tissue.

Piglets are characteristically cold intolerant and thus susceptible to high mortality. However, browning of white adipose tissue (WAT) can induce non-shivering thermogenesis as a potential strategy to facilitate the animal's response to cold. Whether cold exposure can induce browning of subcutaneous WAT (sWAT) in piglets in a similar manner as it can in humans remains largely unknown. In this study, piglets were exposed to acute cold (4°C, 10 h) or chronic cold exposure (8°C, 15 days), and the genes and proteins of uncoupling protein 1 (UCP1)-dependent and independent thermogenesis, mitochondrial biogenesis, lipogenic and lipolytic processes were analysed. Interestingly, acute cold exposure induced browning of porcine sWAT, smaller adipocytes and the upregulated expression of UCP1, PGC1α, PGC1ß, C/EBPß, Cidea, UCP3, CKMT1 and PM20D1. Conversely, chronic cold exposure impaired the browning process, reduced mitochondrial numbers and the expression of browning markers, including UCP1, PGC1α and PRDM16. The present study demonstrated that acute cold exposure (but not chronic cold exposure) induces porcine sWAT browning. Thus, browning of porcine sWAT could be a novel strategy to balance the body temperature of piglets, and thus could be protective against cold exposure.

The expression and function of miRNA-106 in pediatric osteosarcoma.

OBJECTIVE: This study is to investigate the expression and biological function of miRNA-106b in osteosarcoma. PATIENTS AND METHODS: Freshly resected osteosarcoma tissues and the corresponding para-carcinoma tissues were collected from 54 patients. Then miR-106b level in the carcinoma tissues was detected by real-time PCR. To test the function of miR-106b, osteosarcoma cell line U2-OS was transfected with miR-106b inhibitor in vitro. Then, the proliferation, cell cycle, invasion and metastasis of U2-OS cells were detected by CCK-8 assay, flow cytometry, and transwell respectively. The PI3K/AKT signaling pathway activity after treatment was detected by Western blot. RESULTS: miR-106b level was significantly higher in osteosarcoma, and its expression was positively correlated with the lung metastasis and the clinical stages. In vitro experiments showed that the proliferation, invasion, and migration of U2-OS osteosarcoma cells were inhibited after miR-106b inhibition. The transition from G1 to S phase of U2-OS osteosarcoma cells was inhibited after miR-106b inhibition. The Western blot analysis demonstrated that both the AKT phosphorylation in PI3K/AKT pathway and the PI3Kp100 expression were significantly reduced when the expression of miR-106b was down-regulated. CONCLUSIONS: miR-106b level was significantly up-regulated in osteosarcoma, which was positively correlated with the lung metastasis and clinical stages. The down-regulation of miR-16b inhibited the proliferation, invasion, and migration of osteosarcoma cells; thus, it may function as an oncogene to promote osteosarcoma proliferation and invasion through the PI3K/AKT signaling pathway.

Analysis of circulating long non-coding RNA UCA1 as potential biomarkers for diagnosis and prognosis of osteosarcoma.

OBJECTIVE: The aim of this investigation was to examine the potential usefulness of long non-coding RNA UCA1 (UCA1) as a biomarker for diagnosis and prognosis in osteosarcoma. PATIENTS AND METHODS: The expression level of UCA1 was determined using TaqMan real-time PCR in human osteosarcoma tissues and patients' sera. Next, we investigated to clarify the relationship of UCA1 with clinicopathological features. The receiver operating characteristic (ROC) curve was performed to estimate the diagnostic value of UCA1. Finally, the prognosis of patients with osteosarcoma was assessed by Kaplan-Meier method and Cox proportional hazards model. RESULTS: We observed that UCA1 was significantly increased in osteosarcoma tissue compared with normal bone tissue (p<0.001) and the serum expression of UCA1 was significantly higher in patients with osteosarcoma than that in healthy controls (p<0.01). Up-regulation of UCA1 was correlated with clinical stage (p=0.001) and metastasis (p=0.007). Furthermore, serum UCA1 levels were observed to be robust in differentiating osteosarcoma patients from control subjects [area under the curve (AUC) = 0.831; 95% confidence interval (CI)= 0.746 to 0.916]. Kaplan-Meier analysis showed that that high UCA1 expression level was associated with poorer overall survival (p<0.001) and disease-free survival (p<0.001). Finally, Cox regression analyses showed that UCA1 expression might be an independent prognostic parameter to predict poor prognosis. CONCLUSIONS: Our study firstly showed that UCA1 could be a specific and noninvasive candidate biomarker for the diagnosis and prognosis of UCA1.

Up-regulation of long non-coding RNA BCAR4 predicts a poor prognosis in patients with osteosarcoma, and promotes cell invasion and metastasis.

OBJECTIVE: The long non-coding RNA BCAR4 (BCAR4) has been reported to be associated with cancer development. The aim of our study was to investigate the expression of BCAR4 in osteosarcoma patients and its association with clinicopathologic parameters and the prognosis. PATIENTS AND METHODS: Quantitative RT-PCR (qRT-PCR) assay was used to detect the expression of BCAR4 and its correlations with clinicopathological factors were statistically analyzed. The clinical and prognostic significance of BCAR4 expression was analyzed statistically by Kaplan-Meier estimate and Cox regression model. Furthermore, Cell proliferation, migration, and invasion were evaluated using counting assay Kit-8 (CCK-8) and transwell assay, respectively. RESULTS: We found that BCAR4 expression was higher in osteosarcoma tissues and cell lines than that in normal controls. The BCAR4 levels were significantly correlated with clinical stage and distant metastasis. Kaplan-Meier analysis with the log-rank test indicated that high expression of BCAR4 had a decreased overall survival (OS). Univariate and multivariate analyses showed that BCAR4 expression was an independent predictor of overall survival. Furthermore, decreased expression of BCAR4 markedly inhibited osteosarcoma cell proliferation, migration, and invasion. CONCLUSIONS: The results of the present study identified a crucial tumor promotive role of BCAR4 in the progression of osteosarcoma, and suggested that BCAR4 may be a potential therapeutic agent for the treatment of osteosarcoma.

Evaluating the dynamical characteristics of particle matter emissions in an open ore yard with industrial operation activities.

A study to investigate the dynamical characteristics of particle matter emissions in a working open yard is conducted in Caofeidian Port of Hebei Province, China. The average diurnal concentrations of the total suspended particulate (TSP) matter and respirable particulate matter (PM10 and PM5) are monitored during the field measurement campaign. Sampling is performed at a regular interval at 8 monitoring stations in the yard with normal industrial activities. The average TSP, PM10 and PM5 concentrations range from 285 to 568, 198 to 423 and 189 to 330 µg.m-3 in the yard, respectively. The linear regression correlation coefficient of TSP/PM10 and TSP/PM5 is 0.95±0.01 and 0.88±0.02, respectively.By using the Spearman correlation method, the wind speed and relative humidity are both weakly correlated with the PM10 and PM5 concentrations according to the measurements. In addition, industrial operation activities, such as vehicular traffic in the yard and the loading time of stackers, are significantly positively correlated with the PM concentration. Using the multivariate regression method, the main parameters influencing the TSP concentration variations are integratedly analysed. The traffic volume is found to be a significant predictor of TSP concentration variation, with the smallest P value (P<0.05).To understand the dynamical characteristics of particle emissions in the yard, the emissions from the truck transports, that is, from unpaved haul roads and from the loading process, are established. Then, the dynamical emission factor (EFD) based on the industrial activities in the yard is proposed. The dynamical emissions average 5.25x105 kg.year-1 and EFD is evaluated to be 0.29 kg.(ton.day)-1 during the measurement period. These outcomes have meaningful implications not only for understanding the dynamical characteristics of particle emissions in the working stockyard but also for implementing effective control measures at appropriate sites in the harbour area.

Influence of schedule on regulated sensitivity of AML blasts to cytosine arabinoside.

Regulatory molecules that affect the growth culture of blast cells from acute myeloblastic leukemia (AML) may also alter drug sensitivity, a phenomenon that may be called regulated drug sensitivity. Previous studies have shown: (i) blast cells exposed to retinoic acid before cytosine arabinoside (Ara-C) usually show increased sensitivity, but after some retinoic acid exposure times, sensitivity may be decreased; (ii) factor-sensitive or responsive blasts cultured with granulocyte colony-stimulating factor (G-CSF) are regularly more Ara-C-sensitive than when cultured with granulocyte-macrophage CSF (GM-CSF). This paper is concerned with the effects of schedule on drug sensitivity as regulated by either retinoic acid or the myelopoietic growth factors, G-CSF and GM-CSF. We measured the effects of retinoic acid on the sensitivity of blasts cells from the two continuous AML lines to Ara-C or arabinofuranosyl 5-azacytosine (Ara-AC). Cells from seven patients with AML were tested for Ara-C sensitivity in conjunction with retinoic acid. The cells were treated with retinoic acid before or after administration of the drug. Both increases and decreases in Ara-C sensitivity were seen for both schedules. Consistent increases in Ara-C sensitivity were obtained when retinoic acid was included in the methylcellulose cultures used to determine clonogenic cell recovery at each drug dose. In studies of growth factors, a single factor-dependent cell line (OCI/AML-5) was used to compare the effects of G-CSF and GM-CSF on Ara-C sensitivity. An experimental design was used that permitted factors to present in culture for 24 h before Ara-C, during the next 24 h period with the drug, for a subsequent day in suspension without drug, and during the 5-7 days required for colony formation in methylcellulose cultures. G-CSF and GM-CSF were most effective in increasing or decreasing Ara-C, respectively, when the factor under test was included in the methylcellulose cultures. Thus, like retinoic acid, growth factors influenced drug sensitivity when they were present after the drug had been removed. These data, therefore, are compatible with the hypothesis that repair mechanism may contribute to regulated drug sensitivity.

Fluorescence-labeling of nicks in DNA from leukemic blast cells as a measure of damage following cytosine arabinoside. Application to the study of regulated drug sensitivity.

Damage to DNA can be assessed using a technique for labeling nicks in DNA by incubating paraformaldehyde-fixed cells in a mixture of biotin-labeled dUTP, dATP with dNTP and DNA polymerase I. The addition of labeled nucleotides can then be identified by fluorescence by their reaction with streptavidin. We have used this method to examine damage to the DNA of OCI/AML-2 cells caused by cytosine arabinoside (ara-C) and the effects of hydrocortisone and retinoic acid on this damage (regulated drug sensitivity). Concurrent measurements of clonogenic cells were used to allow a comparison of damage as shown by labeled nicks in DNA with loss of colony-forming capacity. Both methods gave comparable ara-C dose-response curves, for cells incubated with the drug for 24 h. Both methods showed that exposure of OCI/AML-2 cells to hydrocortisone before ara-C greatly reduced the toxicity of the drug; and that retinoic acid given after ara-C increased both its lethal effects on colony formation and the extent of DNA damage as assessed by labeled nicks. Clonogenic assays required for colony formation are not readily adapted to the study of development and repair of damage. The labeled nick assay is suitable for such kinetic studies. OCI/AML-2 cells were exposed in suspension to either hydrocortisone before ara-C or retinoic acid after ara-C. At 24 h intervals thereafter, cells were harvested, assayed by both methods, and recultured after dilution to the original cell concentration. In cultures exposed only to ara-C (controls), the number of cells with labeled nicks increased during the first 24 h and cells with damaged DNA could be detected for 48-72 h, depending on the ara-C dose in spite of the dilution at each passage. OCI/AML-2 cells exposed to hydrocortisone before drug showed fewer nick-labeled cells than controls at the first observation and damaged cells rapidly disappeared from the population with increasing time. For cells treated with retinoic acid after ara-C, the nick-labeled cell population was greater than controls and remained greater throughout subsequent observations. We propose that in the control cultures, sublethal damage either became lethal with time and was seen as increased numbers of cells with damaged DNA, or alternatively, sublethal damage was repaired. From this point of view we consider that hydrocortisone promotes repair of sublethal damage while retinoic acid inhibits repair.