Preliminary Investigation on Regulating Effects of Different TCM Treatments on Transcription of the Correlated Genes of Liver Cancer in Rats

The regulating effects of TCM treatments including clearing away heat and toxic materials,promoting blood circulation and removing blood stasis,and strengthening the spleen and regulating qi on the oncogene transcription were observed in the liver cancer model rats.The preliminary results indicated that the mRNA levels of H-ras N-ras and K-ras,and signal molecules correlated with the ras/MAPK signal transduction pathway were down-regulated by the different TCM treatments in varying degrees.Also,the regulating effects of the treatments on differently-displayed genes were discrepant.It is suggested that the molecular mechanisms of the TCM treatments for liver cancer was complex with different target genes.

GABA transporter 1 transcriptional starting site exhibiting tissue specific difference

GABA transporter 1(GAT1) takes important roles in multiple physiological processes through the uptake and release of GABA, but the regulation of GAT1 gene expression in different tissues is rarely known. To address the question, first, 5’ Rapid amplification of cDNA end (RACE) was used to determine GAT1 transcriptional starting sites in neonatal mouse cerebral cortex and intestine, adult mouse brain and adult rat testis. The products of 5’RACE were confirmed by DNA sequencing. We found that the transcript of GAT1 in neonatal mouse cerebral cortex and adult mouse brain starts at the same site (inside of exon 1), while in mouse intestine, GAT1 starts transcription in intron 1, and in rat testis, the transcript of GAT1 has an additional untranslation exon to the 5’ direction.

An Integrated Genetic,Physical and Transcript Map of Homoeologous Chromosomes 12 and 26 in Upland Cotton(Gossypium hirsutum L.)

While Upland cotton(Gossypium hirsutum L.) represents 95% of the world production,its genetic improvement is hindered by the shortage of effective genomic tools and resources.The

Prepartion and identification of activity of anti-HPV-6b/11E1 universal ribozyme——Rz1198 in vitro

Aim: To study the preparation and cleavage activity of Rz1196 directed against HPV-6bE1 and HPV-11E1 (HPV-6b/11E1) transcripts in vitro. Methods: HPV-6b/11E1 gene fragments were cloned into T-vector under the control ofT7 promoter, ～(32)P-labeted HPV-6b/11E1 transcripts as target-RNAs were transcribed in vitro and purified by PAGE.Rz1198 gene designed as a universal ribozyme for both HPV-6b/11E1 transcripts was cloned into vector p1.5 between5'-cis-Rz and 3'-cis-Rz. a2P-labeled Rzl198 transcript was gel-purified, incubated with target-RNAs at different condi-tions and autoradiographed after denaturing gel-electrophoresis. Results: Rz1198 was active at 37℃. The optimaltemperature was 50℃. For HPV-6bE1, k_m = 12.2 nmol/L, k_cat = 0.18 min～(-1); For HPV-11E1, k_m= 14.7 nmol/L,k_cat =0.14 min～(-1). All these revealed that the design of Rz1198 was correct. It could be a universal ribozyme for thetwo substrates--HPV-6bE1 and HPV-11E1 transcripts. Conclusion: Rz1198 prepared in vitro possesses the perfectspecific catalytic cleavage activity. It leads to the expectation that, in the future, it will be possible to develop a newnucleic acid drug from Rz1198 which can efficiently inhibit the replication of HPV-6b/11 DNA in vivo. (Asian J An-drol 1999 Dec; 1: 195-201)

Expression of estrogen receptor (ER) -α and -βtranscripts in the neonatal and adult rat cerebral cortex, cerebellum, and olfactory bulb

In the present study expression of estrogen receptor subtype -alpha (ERalpha) and -beta (ERbeta) in the cerebral cortex, cerebellum, and olfactory bulb was investigated and compared between neonatal (1 to approximately 3-days-old) and adult (250 to approximately 350 g) rats, using reverse transcription-polymerase chain reaction (RT-PCR). No ERalpha transcripts were detectable in the adult cerebellum and olfactory bulb, whereas very weak expression of ERalpha was present in the adult cerebral cortex. No significant difference in ERbeta transcripts was detectable between the neonatal and adult rats. While transcripts for both ER subtypes were co-expressed in these brain areas of neonatal rats, although ERalpha expression was significantly weaker than ERbeta. Even in the cerebral cortex known to contain both ER subtypes in adult rats, ERalpha transcripts in neonatal rats were much higher than in adult. These observations provide evidence for the existence of different expression patterns of ERalpha/ERbeta transcripts in these three brain areas between the neonatal and adult rats, suggesting that each ER subtype may play a distinct role in the regulation of differentiation, development, and functions of the brain by estrogen.

Dynamic changes in the collagen metabolism of liver fibrosis at the transcription level in rabbits with Schistosomiasis japonica

OBJECTIVE: To study the role of the synthesis and degradation of collagen at the transcription level during liver fibrogenesis due to schistosomiasis japonica in rabbits. METHODS: New Zealand rabbits challenged by cercariae of Schistosoma japonicum (S. japonicum) were served as animal models for liver fibrosis. Liver specimens were collected through operations at 4, 6, 8, 10, 12, 16, 20, 24 and 28 wks after challenge. Type I collagen, type III collagen, type IV collagen, MMP-1 and MMP-9 mRNA levels of liver tissue were detected by RT-PCR + Dot blot. The size of egg granulomas and the degree of liver fibrosis were measured by histopathological examinations. RESULTS: Type I collagen, type III collagen, type IV collagen, MMP-1 and MMP-9 mRNA levels increased simultaneously in the early stage after challenge. Most of them reached their peak at 10 weeks, and compared with normal controls, type I collagen, type III collagen, type IV collagen, MMP-1 and MMP-9 mRNA levels increased by 12.0-, 11.0-, 6.6-, 10.0- and 11.0-fold, respectively, coinciding with the change of egg granulomas, i.e., the change in the inflammatory process. Then both collagen and collagenase mRNA levels decreased. Type I, III and IV collagen mRNA levels declined to 2-fold to 3-fold as compared with normal controls (P 0.05) at 28 wks. This study shows that the synthesis and degradation of collagen keep a dynamic balance at the early stage of schistosomiasis japonica challenge, while at the later stages the quantity of collagen synthesis was higher than that of collagen degradation. CONCLUSIONS: It was confirmed at transcription level that when the quantity of collagen synthesis was higher than that of collagen degradation liver fibrogenesis may be resulted in.

Effects of hepatocyte growth factor-mediated activation of Dll4-Notch-Hey2 signaling pathway

Background Hepatocyte growth factor （HGF） treats ischemic disease by promoting arteriogenesis, however, its mechanism of action is not known. The notch signaling pathway plays an important role in neovascularization. The relationship between the proliferation and migration ability of artery endothelial cells and the Dll4-Notch-Hey2 signaling pathway in the process of arteriogenesis was investigated as a mechanism of action of HGE Methods Based on the prophase study cells and supernatant were harvested at the indicated time after human femoral artery endothelial cells （HFAECs） were infected with adenovirus-HGF （Ad-HGF） at 200 pfu/cell. Cells were analyzed for HGF expression and Notch1, DII4 and Hey2 expression by ELISA and reverse transcription-PCR （RT-PCR）. The changes in the proliferation and migration ability of HFAECs were observed by M-I-I- and Transwell migration experiments Ad-GFP-infected HFAECs were used as control. Results Compared with the control group the Ad-HGF group＇s HGF expression was not increased with time, and the induction by HGF of Notch1, DII4 and Hey2 gene transcription was not enhanced with an increase of HGF. The proliferation ability of Ad-HGF-transduced HFAECs was enhanced and their migration ability was also enhanced in the presence of HGF. Conclusions Through activating the DII4-Notch-Hey2 signaling pathway, HGF indirectly promotes the proliferation and migration ability of cells, so that offspring artery branches are formed.

Polymorphisms and functions of the aldose reductase gene 5' regulatory region in Chinese patients with type 2 diabetes mellitus

OBJECTIVE: To screen the 5' regulatory region of the aldose reductase (AR) gene for genetic variabilities causing changes in protein expression and affecting the promoter function. METHODS: The screenings were carried out by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). All SSCP variants were submitted for DNA sequencing and inserted into the plasmid chloromycetin acetyl transferase (CAT) enhancer vector. The constructs were used to transfect Hela cells, and CAT assays were performed to assess promoter activity. Gel mobility shift and footprinting assays were also performed to determine the interaction between the DNA and nuclear proteins. RESULTS: Two polymorphisms, C (-106) T and C (-12) G, were identified in the regulatory region in 123 Chinese control subjects and 145 patients with type 2 diabetes mellitus. The frequencies of genotypes WT/WT, WT/C (-12) G and WT/C (-106) T were not significantly different between the subjects and patients. In the patients with and without retinopathy, frequencies of WT/C (-106) T were 31.5% and 17.5% (P 0.05) respectively. The total frequency of WT/C (-12) G and WT/C (-106) T in patients with retinopathy was 41.8%, significantly higher than that (20.0%) in patients without retinopathy (P