Anthropogenic, biogenic, and photochemical influences on surface formaldehyde and its significant decadal (2006-2017) decrease in the Lewiston-Clarkston valley of the northwestern United States.

Formaldehyde (HCHO) is a key carcinogen and plays an important role in atmospheric chemistry. Both field measurements and Positive Matrix Factorization (PMF) modeling have been employed to investigate the concentrations and sources of HCHO in the Lewiston-Clarkston (LC) valley of the mountainous northwestern U.S. Different instruments were deployed to measure surface formaldehyde and other related compounds in July of 2016 and 2017. The measurements reveal that the average HCHO concentrations have significantly decreased to 2-5 ppb in the LC valley in comparison to its levels (10-20 ppb) observed in July 2006. This discovery with surface measurements deserves attention given that satellite retrievals showed an increasing long-term trend from 2005 to 2014 in total vertical column density of HCHO in the region, suggesting that satellite instruments may not adequately resolve small valleys in the mountainous region. Our PMF modeling identified four major sources of HCHO in the valley: (1) emissions from a local paper mill, (2) secondary formation and background, (3) biogenic sources, and (4) traffic. This study reveals that the emissions from the paper mill cause high HCHO spikes (6-19 ppb) in the early morning. It is found that biogenic volatile organic compounds (VOCs) in the area are influenced by national forests surrounding the region (e.g., Nez Perce-Clearwater, Umatilla, Wallowa-Whitman, and Idaho Panhandle National Forests). The results provide useful information for developing strategies to control HCHO levels and have implications for future HCHO studies in atmospheric chemistry, which affects secondary aerosols and ozone formation.

C/EBPα gene as a genetic marker for beef quality improvement.

Intramuscular fat (IMF) or intramuscular triglycerides are interspersed throughout the skeletal muscles. The IMF, also called marbling, imparts meat with flavor and juiciness and is one of the core criteria for judging carcass value. The quantity of IMF is influenced entirely by genetics. Recently, understanding the underlying genetic bases of IMF has been a focus particularly in the beef industry. In this study, with the deep insights of ameliorating the beef quality by genetic means, the role of the CCAAT/enhancer binding protein alpha (C/EBPα) gene was investigated by over-expressing C/EBPα in bovine muscle stem cells (MSCs) to initiate the adipogenic program. Prior to this, bovine MSCs were isolated and induced to differentiate into adipocytes from cells that were exposed to dexamethasone isobutylmethylxanthine and indomethacin; the presence of insulin and fetal bovine serum was examined. Either ectopic expression of C/EBPα or treatment with dexamethasone and insulin induced the accumulation of fat droplets and the expression of adipogenic induction genes (LPL, PPARγ, C/EBPß, and C/EBPδ). The expression levels of myoblast-related genes (MyoD, Myf5, and Pax7) were also measured to assess the accuracy of the differentiation process. This study provides evidence that the C/EBPα gene is essential for cattle adipose tissue growth and development. Hence, this finding can contribute to improving beef carcass quality.

Polymorphisms of the Osteocrin gene and its association with meat quality traits in Qinchuan cattle.

Here, we detected 2 SNPs, A85C and T335C, that were located on the 3rd exon and the 3 untranslated regions of the bovine Osteocrin gene, respectively, using 413 Qinchuan cattle DNA samples. PCR-SSCP and DNA sequencing methods were specifically used. Three genotypes (AA, AC, and CC) were found at A85C; yet, only 2 genotypes (TC and CC) were found at T335C. Association analysis showed that both loci were associated with certain meat quality traits, including back fat thickness and loin muscle area. At the A85C locus, individuals with the CC genotype had greater back fat thickness. In comparison, at the T335C locus, individuals with the TC genotype had greater back fat thickness and a larger loin muscle area. Therefore, these 2 SNPs could be used as genetic markers to enhance Qinchuan cattle breeding programs.

Comparative study on the immunogenicity between Hsp70 DNA vaccine and Hsp65 DNA vaccine in human Mycobacterium tuberculosis.

The BALB/c mice were immunized with Hsp70 DNA and Hsp65 DNA vaccines in human Mycobacterium tuberculosis. Eight weeks after immunization, the eyeballs were removed, blood and spleen taken, and intraperitoneal macrophages were harvested. The lymphocytic stimulating index (SI) was used to measure the cellular proliferating ability and NO release to measure the phagocytic activity of the macrophages. With ELISA kit, the levels of interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) in serum and the splenic lymphocytic cultured supernatant were detected. The results showed that after the mice were immunized with 100 micrograms/mouse of Hsp70 DNA vaccine intramuscularly, the splenic lymphocytic proliferating ability in the mice was significantly increased as compared with that in the control group, vector group and Hsp65 DNA vaccine group (P < 0.01); The contents of NO in the intraperitoneal macrophages of the mice were significantly lower than in the control group and Hsp65 DNA vaccine group (P < 0.01); The levels of serum IL-2 in the mice were significantly higher than in the control group, but there was no statistical difference between Hsp65 DNA group and vector group (P > 0.05); The contents of serum IFN-gamma in the mice were significantly higher than in the control group, but significantly lower than in the Hsp65 DNA vaccine group (P < 0.05). It was indicated that immunization with Hsp70 DNA vaccine could obviously enhance the immune response, but its intensity seemed inferior to Hsp65 DNA vaccine. The anti-infection mechanisms and clinical use in the future of the vaccines of Hsp70 DNA and Hsp65 DNA are worth further studying.

A multicenter, randomized, controlled trial of interferon alfacon-1 compared with alpha-2a-interferon in Chinese patients with chronic hepatitis C virus infection.

BACKGROUND: Alpha-interferons are the accepted therapy for patients infected with chronic hepatitis C virus (HCV) in China. However, consensus interferon (CIFN) for HCV treatment is effective in patients with chronic hepatitis C from Western countries. METHODS: This randomized, controlled trial was conducted to determine the safety and efficacy of CIFN at two doses, and to compare it with alpha-2a-interferon (IFN-alpha-2a) in Chinese patients with chronic HCV. Interferon-naive patients with chronic HCV infection (n = 187) were randomly chosen to receive 15 microg CIFN or 9 microg or 3 MU IFN-alpha-2a subcutaneously, three times a week for 24 weeks, followed by a 24 week observation period. Efficacy was evaluated by the normalization of serum alanine aminotransferase (ALT) and the non-detectability disappearance of serum HCV-RNA by using reverse-transcription-polymerase chain reaction. The safety of CIFN was evaluated by recording the type and severity of adverse effects. RESULTS: The combined ALT and HCV-RNA end-of-treatment and sustained responses were observed to be greater for treatment with 15 microg CIFN (59.0% and 55.7%, respectively) compared to IFN alpha-2a (36.1% and 39.3%, respectively; P = 0.01 for the end-of-treatment, P = 0.07 for the sustained response). The combined ALT and HCV-RNA end-of-treatment and sustained responses for treatment with 9 microg CIFN (both 49.2%) were higher than those for IFN-alpha-2a (not statistically significant). Data were analyzed by using a logistic-multiple-variate regression model, which indicated that the higher IFN dose (15 microg or 9 microg CIFN vs 3 MU IFN-alpha-2a; P < 0.01) appeared to be associated with a better sustained response. The type, frequency and severity of adverse effects were comparable across treatment groups. CONCLUSIONS: Consensus interferon appears to be safe and effective at concentrations of 9 and 15 microg, but 15 microg CIFN may be more effective than 3 MU IFN-alpha-2a, without increased toxicity.

[Construction of recombinant BCG bearing Schistosoma japonicum 26Ku antigen gene and study on its immunogenicity on mice].

OBJECTIVE: To construct recombinant BCG vaccine bearing Schistosoma japonicum 26Ku glutathione S-transferase (Sj26GST) gene and determine its immunogenicity on BALB/c mice. METHODS: Using techniques of molecular biology, human mycobacterium tuberculosis HSP70 promoter and Sj26GST gene were linked to produce a fused gene. The fused gene was cloned into an E. coli-Mycobacterium shuttle plasmid pBCG-2000 to construct an E. coli-Mycobacterium expression shuttle plasmid pBCG-Sj26 that could express Sj26GST gene. Then, the pBCG-Sj26 was introduced by electroporation into mycobacterium bovis BCG to construct a recombinant BCG vaccine bearing Sj26GST gene (rBCG- Sj26GST). The expression of Sj26GST gene in BCG was induced by heating. The lymphocyte stimulating index (SI), macrophage activity and IL-2, IFN-gamma levels of the serum and culture supernatant of spleen lymphocytes were tested after immunization of BALB/c mice with rBCG-Sj26GST vaccine. RESULTS: The fused gene of HSP70 promoter and Sj26GST cDNA was inserted into an E. coli-Mycobacterium shuttle expression plasmid by analysing electrophoresis results on PCR products using plasmid pBCG-Sj26 as a templet. The content of rSj26GST contained 15% of total bacterial protein of BCG. The SI of the experimental group was 2.26 +/- 0.43, which was significantly higher than those in the control group (1.61 +/- 0.28, P < 0.05), vector group (1.48 +/- 0.30, P < 0.05) and BCG group (1.42 +/- 0.26, P < 0.05). The macrophage NO level of the experimental group was (357.42 +/- 84.11) nmol/ml which was significantly higher than those in the control group (183 nmol/ml +/- 33 nmol/ml, P < 0.01) and vector group (203 nmol/ml +/- 56 nmol/ml, P < 0.01). The serum IL-2 level of the experimental group was (267 pg/ml +/- 130 pg/ml), which was significantly higher than those in the control group (45 pg/ml +/- 15 pg/ml, P < 0.01) and vector group (52 pg/ml +/- 29 pg/ml, P < 0.05. Compared with the control group, the serum IFN-gamma level increased by 20%, the IL-2 level of the culture supernatant of spleen lymphocytes increased by 44%. CONCLUSIONS: The foreign gene encoding Sj26 GST can be expressed in BCG. rBCG Sj26GST vaccine may induce stronger immune response in BALB/c mice than in control, vector and BCG groups.

The construction of Schistosoma japonicum vaccine BCG-Sj26GST and its identification.

The expression of foreign gene, Schistosoma Japonicum 26 ku antigen (Sj26GST), in Bacillus Calmette-Guerin (BCG), Mycobacterium (M. smegmatis) and Escherichia coli (E. coli) were studied. The cDNA fragment encoding Sj26GST was amplified by PCR using plasmid pGEX, which could express Sj26GST in E. coli as template. The Sj26GST cDNA was cloned into the down-stream of human M. tuberculosis heat shock protein (hsp) 70 promoter with correct reading frame, and then the DNA fragment containing hsp70 promoter and Sj26GST gene were subcloned together into E. coli-Mycobacteria shuttle plasmid pBCG-2000 to construct the expression shuttle plasmid pBCG-Sj26. The recombinant BCG and M. smegmatis mc(2)155, which were electroplated with pBCG-Sj26, could express Sj26GST and the recombinant Schistosoma Japonicum vaccine BCG-Sj26GST was made. The recombinant Sj26GST (rSj26GST) were soluble and could be observed on SDS-PAGE at molecular weight of 26 ku. The content of rSj26GST accounted for 15% and 10% of total bacterial protein in BCG and M. smegmatis respectively. The results of Western blot showed the combination of rSj26GST with antibody of GST.

[Study of HSP70 gene upstream regulation element on expression efficiency of GST gene in M. smegmatis].

Four different expression vectors were constructed by cloning foreign gene which encode Schistosoma japonicum 26 kD antigen (Sj26GST) into Escherichia coli-Mycobacteria shuttle plasmid pBCG-2000 and investigated their expression efficiency in mycobacteria smegmatis. The plasmid which contains promoter of human mycobacterial tuberculosis heat shock protein 70 was firstly digested with Nco I and modified with two different ways to lead to two kinds of SD sequences, and ligated with Sj26GST encoding gene. Then, the DNA fragment contained HSP70 promoter and Sj26GST gene was obtained and cloned into E. coil-mycobacteria shuttle plasmid pBCG-2000, and finally four recombinant mycobacterial expression vectors that differenciated in SD sequence, orientation and copy number were selected. The expressed native recombinant Sj26GST(rSj26GST) was soluble and could be observed on SDS-PAGE about at the molecular weight of 26 kD obviously. Analysis with protein density scanning indicated: the expression efficiency that containing double-copy promoter-foreign gene vector was the highest and the expressed protein which was about 1.6 times than others was 28% of total protein of Mycobacteria smegmatis. The cloned direction and SD sequence had no significant effect on expression efficiency.

Expression of foreign gene in mycobacterium regulated by human Mycobacterium tuberculosis heat shock protein 70 promoter.

The DNA fragments of 150bp length promoter of human Mycobacterium (M.) tuberculosis heat shock protein (hsp) 70 and 650bp length foreign gene, the Schistosoma japonicum glutathione S-transferase (Sj26GST) gene, were obtained by amplification with polymerase chain reaction. And the 150bp DNA sequence upstream initiation codon ATG of the human M. tuberculosis hsp 70 promoter that contains the sequence TTGAG and ATCATA which consensus with E. coli promoter's -35 and -10 region respectively, as well as ribosome binding site GGAGG at position -12--8 upstream the ATG were determined by SangerDideoxyribonucleotide-mediated chain-termination method. Then, the human M. tuberculosis hsp70 promoter and Sj26GST cDNA were cloned into E. coli-mycobacteria shuttle plasmid pBCG- 2000 to construct E. coli-Mycobacterium expression shuttle plasmid pBCG- Sj26 that can express Sj26GST gene. The M. smegmatis were electroporated and the positivecolonies were selected by kanamycin. The M. smegmatis containing the vector pBCG-Sj26 can be induced by heating and hydrogen peroxide (H2O2) to express GST. The molecular weight of the recombinant GST (rGST) was 26,000. The rGST contents that were about 10 percent of the total bacterial protein were analyzed by density scanning after running SDS-PAGE. This study would provide scientific evidences for application of hsp70 promoter in expressing foreign gene in mycobacterium and development of mycobacterium as multiple-valent vectoral vaccine.

Construction of shuttle expression plasmid and stable expression of foreign gene in mycobacteria and E. coli.

By employing the pUC19 as a backbone, the regulatory and signal sequences which encode kanamycin resistance, and mycobacterial plasmid origin of replication (oriM) were cloned into the pUC19. The recombinant E. Coli-mycobacteria shuttle expression plasmid pBCG-8000 was constructed. The pBCG-8000 was able to replicate in both E. Coli and mycobacteria (including BCG) systems, and to confer stable kanamycin resistance upon transformants. The study should facilitate the development of BCG and other mycobacteria into multivalent vaccine vectors.

The production of strains highly expressing human interleukin-2 cDNA.

The plasmid pTLIL-2, which only directed a low-level expression of human interleukin 2(IL-2) cDNA in E. coli, was reconstructed: a series of deletions were made in 3' non-coding region of human IL-2 cDNA, and 7 recombinant plasmids with different deletions were selected, on the other hand, a Tac promoter sequence from pDR540 was inserted to upstream of IL-2 cDNA in pTLIL-2 so that pTLIL-2DT, which contains double Tac promoters, was constructed. And then, E. coli JM103 was transformed with the above 8 recombinant plasmids respectively. The expression efficiency of IL-2 cDNA in E. coli transformed with different plasmids was evaluated by means of SDS-PAGE and measuring 3H-TdR incorporation of IL-2-dependent activated T lymphocytes in the presence of bacterial extracts. Three engineering strains with high efficiency of IL-2 expression were selected, and all of these strains could produce recombinant IL-2(rIL-2) 4 times more than E. coil JM103/pTLIL-2.

[Experimental study of the protective action of re-du-qing on H(+)-APT synthase activity of mitochondria in the rabbit].

To observe the effect of Re-Du-Qing on the toxic effect of endotoxin, the rabbit model of general Shwartzman reaction was made by injection of endotoxin into ear vein and the H(+)-ATP synthase activities of rabbit liver mitochondria were measured. The results showed that the H(+)-ATP synthase activity of endotoxin group was significantly lower than that of the normal group and group which had been simultaneously injected Re-Du-Qing (P less than 0.05). The level of H(+)-ATP synthase activity was almost raised to normal value, when Re-Du-Qing or dexamethasone was injected simultaneously with endotoxin respectively. It suggested that Re-Du-Qing was able to alleviate the nocuous effect of endotoxin on H(+)-ATP synthase activity in the mitochondria of rabbit liver cells.

Normal and leukemic lymphocyte membrane fluidity and response to stimulation with ConA and PHA.

In our experiment, lymphocyte membrane was labeled by DPH fluorescence probe. The rate of rotation of the probe can be measured from the value of fluorescence polarization (PDPH). With this method useful information could be provided about membrane fluidity of lymphocytes. It was found that the F value (unit of lipid fluidity of membrane) of leukemic lymphocytes was obviously higher than that of normal ones. Furthermore, the F value of cultured leukemic Ts lymphocytes was the highest. In contrast with normal spleen T-lymphocytes or mixed lymphocytes, the response of malignant lymphocytes to the stimulation with ConA or PHA was reflected in the decrease of PDPH value or the increase of F value. Unexpectedly, the F value of T-lymphocytes from "615" mouse not injected with tumour cells was also higher than that of the mixed. The possibility of using the membrane fluidity as a diagnostic criterion was also discussed.