Intrafollicular expression and potential regulatory role of cocaine- and amphetamine-regulated transcript in the ovine ovary.

Follicular growth is regulated by a complex interaction of pituitary gonadotropins with local regulatory molecules. Previous studies demonstrated an important role for cocaine- and amphetamine-regulated transcript (CART) in regulation of granulosa cell estradiol production associated with dominant follicle selection in cattle. However, intraovarian expression and actions of CART in other species, including sheep, are not known. The objective of this study was to investigate the expression of CART in sheep follicles and determine the effects of CART on indices of ovine granulosa cell function linked to follicular development. Results demonstrated the expression of CART messenger RNA and prominent intraovarian localization of CART peptide in granulosa cells of sheep follicles. Granulosa cell CART messenger RNA was lower, but follicular fluid estradiol concentrations were higher in large (>5 mm) follicles vs smaller 3- to 5-mm follicles harvested from sheep ovaries of abattoir origin. CART treatment inhibited follicle stimulating hormone-induced estradiol production by cultured ovine granulosal cells and also blocked the follicle stimulating hormone-induced increase in granulosa cell numbers. Results demonstrate expression of CART in sheep follicular tissues and suggest potential biological actions of CART, which are inhibitory to ovine follicular growth and development.

DNA methylation regulates expression of VEGF-C, and S-adenosylmethionine is effective for VEGF-C methylation and for inhibiting cancer growth

DNA hypomethylation may activate oncogene transcription, thus promoting carcinogenesis and tumor development. S-adenosylmethionine (SAM) is a methyl donor in numerous methylation reactions and acts as an inhibitor of intracellular demethylase activity, which results in hypermethylation of DNA. The main objectives of this study were to determine whether DNA hypomethylation correlated with vascular endothelial growth factor-C (VEGF-C) expression, and the effect of SAM on VEGF-C methylation and gastric cancer growth inhibition. VEGF-C expression was assayed by Western blotting and RT-qPCR in gastric cancer cells, and by immunohistochemistry in tumor xenografts. VEGF-C methylation was assayed by bisulfite DNA sequencing. The effect of SAM on cell apoptosis was assayed by flow cytometry analyses and its effect on cancer growth was assessed in nude mice. The VEGF-C promoters of MGC-803, BGC-823, and SGC-7901 gastric cancer cells, which normally express VEGF-C, were nearly unmethylated. After SAM treatment, the VEGF-C promoters in these cells were highly methylated and VEGF-C expression was downregulated. SAM also significantly inhibited tumor growth in vitro and in vivo. DNA methylation regulates expression of VEGF-C. SAM can effectively induce VEGF-C methylation, reduce the expression of VEGF-C, and inhibit tumor growth. SAM has potential as a drug therapy to silence oncogenes and block the progression of gastric cancer.

DNA methylation regulates expression of VEGF-C, and S-adenosylmethionine is effective for VEGF-C methylation and for inhibiting cancer growth.

DNA hypomethylation may activate oncogene transcription, thus promoting carcinogenesis and tumor development. S-adenosylmethionine (SAM) is a methyl donor in numerous methylation reactions and acts as an inhibitor of intracellular demethylase activity, which results in hypermethylation of DNA. The main objectives of this study were to determine whether DNA hypomethylation correlated with vascular endothelial growth factor-C (VEGF-C) expression, and the effect of SAM on VEGF-C methylation and gastric cancer growth inhibition. VEGF-C expression was assayed by Western blotting and RT-qPCR in gastric cancer cells, and by immunohistochemistry in tumor xenografts. VEGF-C methylation was assayed by bisulfite DNA sequencing. The effect of SAM on cell apoptosis was assayed by flow cytometry analyses and its effect on cancer growth was assessed in nude mice. The VEGF-C promoters of MGC-803, BGC-823, and SGC-7901 gastric cancer cells, which normally express VEGF-C, were nearly unmethylated. After SAM treatment, the VEGF-C promoters in these cells were highly methylated and VEGF-C expression was downregulated. SAM also significantly inhibited tumor growth in vitro and in vivo. DNA methylation regulates expression of VEGF-C. SAM can effectively induce VEGF-C methylation, reduce the expression of VEGF-C, and inhibit tumor growth. SAM has potential as a drug therapy to silence oncogenes and block the progression of gastric cancer.

Plant height affects Fusarium crown rot severity in wheat.

Effects of plant height on Fusarium crown rot (FCR) disease severity were investigated using 12 pairs of near-isogenic lines (NILs) for six different reduced height (Rht) genes in wheat. The dwarf isolines all gave better FCR resistance when compared with their respective tall counterparts, although the Rht genes involved in these NILs are located on several different chromosomes. Treating plants with exogenous gibberellin increased FCR severity as well as seedling lengths in all of the isolines tested. Analysis of the expression of several defense genes with known correlation with resistance to FCR pathogens between the Rht isolines following FCR inoculation indicated that the better resistance of the dwarf isolines was not due to enhanced defense gene induction. These results suggested that the difference in FCR severity between the tall and dwarf isolines is likely due to their height difference per se or to some physiological and structural consequences of reduced height. Thus, caution should be taken when considering to exploit any FCR locus located near a height gene.

Effect of individual Sumai 3 chromosomes on resistance to scab spread within spikes and deoxynivalenol accumulation within kernels in wheat.

Two sets of substitution lines were developed by crossing individual monosomic lines of Chinese Spring (recipient) with scab (Fusarium graminearum) resistant cultivar Sumai 3 (donor) and then using the monosomics as the recurrent male parent for four backcrosses (without selfing after each backcross). The disomic substitution lines were separated from selfed BC4F2 plants. Chromosome specific SSR markers were analyzed for polymorphism between Sumai 3 and Chinese Spring. Polymorphic markers were used to identify substitution lines for specific chromosomes. Based on the specific SSR markers, chromosome substitutions occurred in thirty-six lines, and six lines segregated alleles from the two parents or were homozygous for the allele from Chinese Spring. These substitution lines were used to evaluate Type II (spread within the head) and Type V (deoxynivalenol accumulation within kernels) scab resistance. The objective was to use the substitution lines to evaluate the effect of individual chromosomes of Sumai 3 on Type 11 and Type V scab resistance in the greenhouse. Significant differences in Type II scab resistance and deoxynivalenol (DON) levels among different Chinese Spring (Sumai 3) substitution lines were detected. Positive chromosome substitution effects on Type II scab resistance were found on chromosomes 2B, 3B. 6B, and 7A from Sumai 3. Chromosomes 3B and 7A also reduced DON accumulation within the kernels, while chromosomes IB, 2D, and 4D from Sumai 3 increased DON concentration. Chromosome 7A from Sumai 3 had the largest effect on resistance to scab spread and DON accumulation. Additional research is in progress on the scab resistance conferred by chromosome 7A.

Enhanced production of superoxide anion by microglia from trisomy 16 mice.

Disruption of normal oxygen radical metabolism in the CNS may contribute to the neuropathological changes associated with Down syndrome (trisomy 21) and its mouse counterpart, the trisomy 16 (Ts16) mouse. One potent source of oxyradicals is the CNS-specific macrophage, the microglial cell. We prepared primary glial cultures from the cerebral cortices of Ts16 and normal littermate mice taken at day 15 of gestation. Microglia were isolated from confluent cultures after 14 days in vitro and assayed for superoxide anion production using a cytochrome C reduction assay. Stimulation by either opsonized zymosan (OPZ) or phorbol myristate acetate (PMA), produced significantly higher levels (2.8-20 fold) of superoxide per mg protein in Ts16 microglial cultures. Resting, i.e. unstimulated secretion, was not significantly different from littermate controls. Astrocyte enriched cultures, stimulated by OPZ, exhibited low levels of superoxide production which was higher in Ts16 mice than normal littermates. Microglial enriched cultures from rat neonatal cerebral cortices were exposed for 24 h to medium from the Ts16 glial cultures. Superoxide production in the Ts16 media treated rat microglia was significantly higher than in those treated with littermate conditioned media.