Biomedical subjects

Li Chen

Publications and source records attributed to Li Chen.

At least 55 records · Page 3Linked to original sources

PubMed2006

Selective small-molecule inhibitor reveals critical mitotic functions of human CDK1.

CDK1 is a nonredundant cyclin-dependent kinase (CDK) with an essential role in mitosis, but its multiple functions still are poorly understood at a molecular level. Here we identify a selective small-molecule inhibitor of CDK1 that reversibly arrests human cells at the G(2)/M border of the cell cycle and allows for effective cell synchronization in early mitosis. Inhibition of CDK1 during cell division revealed that its activity is necessary and sufficient for maintaining the mitotic state of the cells, preventing replication origin licensing and premature cytokinesis. Although CDK1 inhibition for up to 24 h is well tolerated, longer exposure to the inhibitor induces apoptosis in tumor cells, suggesting that selective CDK1 inhibitors may have utility in cancer therapy.

Apoptosis↗

PubMed2006

The tetanus patterns for the induction of long-term depression in the adult rat hippocampus.

Previous reports suggested that a low-frequency stimulus (LFS) of 1~2 Hz (600~900 pulses) induced a homosynaptic long-term depression (LTD) of synaptic efficacy in the hippocampal CA1 area of young rats (< 4-week old). However, these stimulation protocols often failed to induce LTD in the adult CA1 hippocampus. In the present study, we examined the effects of two novel tetanus patterns on LTD induction in adult rat hippocampal slices. We determined that these novel stimulation protocols induced LTD in the adult hippocampus, and that the characteristics of induced LTD were parameter-specific, including latency (period from the end of tetanus to a beginning of LTD) and the amplitude of LTD. These results suggest that LFS with certain patterns can induce LTD in the CA1 area of adult rat hippocampal slices, and that the multi-trains of 2-Hz protocol provided more effective response than the 5-Hz protocol.

Animals↗

PubMed2006

Endoplasmic reticulum stress response is involved in the pathogenesis of stress induced gastric lesions in rats.

Stress gastric ulcer is a serious complication, but the mechanism involved is not fully clarified. It is well known that mucosal cell apoptosis plays a crucial role in the pathogenesis of gastric ulceration. Recent studies have shown that endoplasmic reticulum (ER) stress is an important pathway leading to cellular apoptosis. To investigate the role of ER stress in the pathogenesis of stress gastric ulcer, we studied the alteration in the expression of ER stress markers GRP78 (glucose-regulated protein 78) and caspase-12 (an ER stress-specific proapoptotic molecule) and their relations with gastric mucosal apoptosis during development of stress gastric lesions in the water-immersion and restraint stress (WRS) model in rats. Rats developed severe gastric lesions after 6 h of WRS. Typical apoptosis was observed at the edge cells of WRS induced gastric lesions. Western blot analysis showed that GRP78 and activated caspase-12 were over-expressed in the gastric tissues of WRS rats. Immunohistochemical analysis demonstrated that increased GRP78 and caspase-12 were distributed only under the lesions. In addition, dithiothreitol and tunicamycin (ER stress inducers), which increased the expression of GRP78 and activated caspase-12, caused gastric mucosal injury and mucosal cell apoptosis in vitro. These findings suggest that ER stress might be involved in the development of stress gastric ulcer through an apoptotic mechanism.

2-Pyridinylmethylsulfinylbenzimidazoles↗

PubMed2006

STI571 combined with vincristine greatly suppressed the tumor formation of multidrug-resistant K562 cells in a human-nude mice xenograft model.

BACKGROUND: The development of the targeted signal transduction inhibitor STI571 has prompted us to treat chronic myeloid leukemia in different ways. Since STI571 may reverse multidrug-resistance of K562/MDR cells in vitro, we studied the effect of STI571 on multidrug-resistant K562 cells in vivo. METHODS: Multidrug-resistant human leukemia cell line K562-n/VCR expresses both bcr/abl fusion gene and multi-drug resistance (mdr1) gene. It is a vincristine resistant cell line subcloned from the vincristine (VCR) sensitive cell line K562-n induced by vincristine in vitro. K562-n and K562-n/VCR cells were inoculated subcutaneously into both sides of nude mice breast (5 x 106 cells/each) to establish a human leukemia xenograft model. The incidence and volume of tumor were observed. In the tumor-bearing nude mice, anti-tumor drugs vincristine, daunorubicin (DNR), STI571, and STI571 plus VCR for the treatment of mdr1 and bcr/abl double positive leukemia were studied respectively. RESULTS: The tumor incidence was 100% in the nude mice inoculated with either K562-n or K562-n/VCR. The transcription of the mdr1 gene and expression of P-gp were negative in K562-n cells but positive in K562-n/VCR cells. The intracellular accumulation of DNR in K562-n cells was higher than that in K562-n/VCR cells (P < 0.05). The tumor incidence of K562-n/VCR cells in nude mice was much higher than that of K562-n cells in chemotherapy groups, and the mean volume of the tumors was also larger (P < 0.05). STI571 combined with VCR significantly suppressed the proliferation of K562-n/VCR cells. CONCLUSIONS: The MDR characteristics of K562-n/VCR in vivo were the same as in vitro. STI571 had a significant tumor-suppressing effect on VCR-sensitive leukemia cells and a moderate effect on MDR leukemia cells. VCR combined with STI571 had an excellent tumor-suppressing effect on both K562-n/VCR and K562-n in the human-nude mice xenograft model.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2006

Proteasome dysfunction in aged human alpha-synuclein transgenic mice.

A deficit in proteasome function in Parkinson's disease has been speculated. We characterized the ubiquitin-proteasome system in three regions of brain from transgenic and nontransgenic littermates. Mice expressing a doubly mutated form of human alpha-synuclein had significant impairments whereas mice expressing the wild-type gene had lesser changes compared to nontransgenic littermates. Significant abnormalities in line hm2 alpha-SYN-39 included declines in 20S-mediated proteolytic activity, the level of the 19S proteasome subunits Rpt1 and Rpn2, and the level of soluble total high MW ubiquitin cross-reacting proteins. Line hw alpha-SYN-5 had significant, but restricted proteasome abnormalities. The severity of impairment was proportional to the substantia nigra dopaminergic neuronal loss previously identified. There were significant correlations between the level of Rpn2 with the level of Rpt1, the activity of the 20S proteasome, and the level of soluble high MW ubiquitin cross-reacting proteins. These abnormalities in symptomatic line hm2 alpha-SYN-39 mice are consistent with abnormalities identified in tissue from patients with Parkinson's disease.

Aging↗

PubMed2006

Evidence for distinct functions for human DNA repair factors hHR23A and hHR23B.

Rad23 proteins bind ubiquitinated substrates and the proteasome, consistent with an important role in protein degradation. Although human Rad23 proteins (hHR23A and hHR23B) have redundant roles in DNA repair, we determined they formed distinct interactions with proteasomes and multiubiquitinated proteins, but similar binding to Ataxin-3. Threonine-79 contributed to the weak proteasome-binding property of hHR23A, and its conversion to proline (T79P), which is the residue present in hHR23B, increased proteasome interaction. We also determined that hHR23A and hHR23B could be co-purified with unique proteolytic and stress-responsive factors from human breast cancer tissues, indicating that they have unique functions in vivo.

Amino Acid Sequence↗

PubMed2006

Simvastatin promotes Th2-type responses through the induction of the chitinase family member Ym1 in dendritic cells.

Statins, best known for their lipid-lowering actions, also possess immunomodulatory properties. Recent studies have shown a Th2-biasing effect of statins, although the underlying mechanism has not been identified. In this study, we investigated whether simvastatin can exercise a Th2-promoting effect through modulation of function of dendritic cells (DCs) without direct interaction with CD4+ T cells. Exposure of DCs to simvastatin induced the differentiation of a distinct subset of DCs characterized by a high expression of B220. These simvastatin-conditioned DCs up-regulated GATA-3 expression and down-regulated T-bet expression in cocultured CD4+ T cells in the absence of additional simvastatin added to the coculture. The Th2-biased transcription factor profile induced by simvastatin-treated DCs also was accompanied by increased Th2 (IL-4, IL-5, and IL-13) and decreased Th1 (IFN-gamma) cytokine secretion from the T cells. The Th2-promoting effect of simvastatin was found to depend on the chitinase family member Ym1, known to be a lectin. Anti-Ym1 antibody abolished the Th2-promoting effect of simvastatin-treated DCs. Also, simvastatin was unable to augment Ym1 expression in DCs developed from STAT6-/- or IL-4R alpha-/- mice. Thus, modulation of Ym1 production by DCs identifies a previously undescribed mechanism of Th2 polarization by statin.

Animals↗

PubMed2006

Chronic oral nicotine normalizes dopaminergic function and synaptic plasticity in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-lesioned primates.

Our recent studies show that chronic oral nicotine partially protects against striatal damage in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated nonhuman primates. To identify the cellular changes associated with this protective action, we investigated the effects of nicotine treatment on stimulus-evoked dopamine release, dopamine turnover, and synaptic plasticity in striatum from lesioned and unlesioned animals. Monkeys were chronically (6 months) treated with nicotine in the drinking water and subsequently lesioned with the dopaminergic neurotoxin MPTP (6 months) while nicotine was continued. Nigrostriatal damage increased nicotinic acetylcholine receptor (nAChR)-mediated fractional dopamine release from residual terminals, primarily through changes in alpha3*/alpha6* nAChRs. In contrast, fractional receptor-evoked dopamine release was similar to control in unlesioned and lesioned animals with chronic oral nicotine. Long-term nicotine administration also attenuated the enhanced K(+)-evoked fractional dopamine release from synaptosomes of MPTP-lesioned animals, suggesting that nicotine treatment had a generalized effect on dopaminergic function. This premise was further supported by experiments showing that nicotine dosing decreased the elevated dopamine turnover that occurs after nigrostriatal damage. We next investigated changes in synaptic plasticity with lesioning and nicotine treatment. Nicotine treatment alone enhanced synaptic plasticity by lowering the threshold for long-term depression (LTD) in the corticostriatal pathway. MPTP lesioning led to a loss of LTD, a measure of short-term synaptic plasticity. In contrast, LTD was preserved in nicotine-treated lesioned animals. Thus, the present data show that the disruptions in striatal dopaminergic function after nigrostriatal damage were attenuated with chronic nicotine administration. These cellular alterations may underlie the ability of nicotine to maintain/restore normal function with nigrostriatal damage.

Administration, Oral↗

PubMed2006

Hyperhomocysteinemia associated with decreased renal transsulfuration activity in Dahl S rats.

Elevated plasma homocysteine (Hcys) has been reported to participate in the development of arterial and glomerular sclerosis in Dahl salt-sensitive hypertensive (SS) rats. The mechanism resulting in hyperhomocysteinemia in these animals remains unknown. Disposal of Hcys in the kidneys plays an important role in regulating the plasma Hcys level. We, therefore, examined the activities and expressions of the enzymes involved in the metabolism of Hcys in the kidneys of SS rats, compared with that of Brown Norway rats and SSBN13 rats, a consomic subcolony of SS rats that carries a substituted chromosome 13 from Brown Norway rats. High-performance liquid chromatography analysis demonstrated that plasma Hcys levels were significantly higher in SS rats. The conversion of S-adenosylhomocysteine into Hcys via S-adenosylhomocysteine hydrolase by renal tissue was not different among these 3 rat strains. However, the metabolic rate of Hcys into cysteine was markedly reduced in the SS rat kidneys. The mRNA and protein levels of cystathionine beta-synthase (CBS), one of the key enzymes in the transsulfuration pathway in the kidneys, were significantly lower in SS rats. In microdissected nephron segments, CBS mRNA was shown to be mainly present in renal proximal tubules (PTs). The mRNA levels of CBS in the PTs were also significantly decreased in SS rats, accompanied by a reduced CBS activity in PTs. We conclude that hyperhomocysteinemia is associated with a decreased activity and expression of CBS in renal PTs because of the defect of chromosome 13 in SS rats.

Adenosylhomocysteinase↗

PubMed2006

[Effect of hyperhomocysteinemia on cardiac remodeling in rats].

OBJECTIVE: To study the effects of hyperhomocysteinemia on cardiac remodeling and systolic function. METHODS: Twenty-four Wistar rats were divided randomly into experiment and control group. 12 rats of experiment group were bred with 1 g/(kg.d) L-methionine. After 4 weeks, 6 of them were killed (HHcy), others were continuously bred with normal diet for 4 weeks (Quit hcy, QHcy). Twelve rats of control group with normal diet for 4 and 8 weeks. The plasma Hcy concentrations were measured by immuno-fluorescence technique. Left ventricular structure and systolic function were assessed by echocardiography. The cardiac morphology and collagen were studied by optical microscopy and image analysis system after hematoxylin eosin-staining and picrosirius red-staining. Immuno-histochemistry for actin-smooth muscle and factor VIII-related antigen was performed to identify changes in cardiac smaller arteriolar and microvascular numbers. RESULTS: The plasma Hcy concentration was (106.19+/-19.75) micromol/L in HHcy, (6.52+/-0.94) micromol/L in QHcy and (4.90+/-0.10) micromol/L in the control. Left ventricular systolic function was decreased 10.7% in HHcy. Left ventricular wall strain and cardiac smaller arteriolar wall thickness were increased by 62.1% and 2.9 fold in HHcy, 27.1% and 2.4 fold in QHcy as compared with the control. Cardiomyocyte diameter was not changed in HHcy and increased by 15.4% in QHcy as compared with the control. Microvascular number was decreased by 45.9% in HHcy as compared with the control, but smaller arteriolar number did not differ among the groups. Left ventricular systolic function and microvascular number resumed in QHcy. Interstitial and perivascular collagen deposition was significantly increased in HHcy and QHcy. CONCLUSION: Hyperhomocysteinemia may increase cardiomyocyte diameter, cardiac smaller arteriolar wall thickness and left ventricular wall strain; decrease microvascular number; induce interstitial and perivascular collagen deposition, directly and indirectly affect on cardiac remodeling and systolic function. The cardiac remodeling couldn't resume completely with plasma Hcy concentration being decreased.

Animals↗

PubMed2006

Exercise training promotes expression of apelin and APJ of cardiovascular tissues in spontaneously hypertensive rats.

Because apelin may play an important regulatory role in human cardiac dysfunction, we investigated alterations in cardiovascular content of apelin and its receptor, APJ, during hypertension and the effect of exercise training on the cardiovascular apelin/APJ system in hypertensive animals. Spontaneously hypertensive rats (SHRs) underwent swimming training consisting of 54 swimming sessions of 60 min each (6 days/week for 9 weeks). Systolic blood pressure (SBP) was verified weekly by tail-cuff plethysmography. Apelin levels in plasma and cardiovascular tissues were determined by radioimmunoassay. The level of apelin/APJ mRNA was determined by RT-PCR. SHRs showed severe hypertension and pathological cardiomegaly. The level of apelin immunoreactivity (apelin-ir) in plasma and ventricular and aortic tissues was lower, by 40%, 40% and 42% (all P<0.01), respectively, in SHRs than in control Wistar-Kyoto rats, and the mRNA level of apelin and APJ in myocardium and aorta was markedly decreased. Compared with sedentary SHRs, swimming-trained SHRs showed decreased SBP and elevated mRNA expression of apelin and APJ in cardiovascular tissues and elevated apelin-ir level in plasma, myocardium and aorta (all P<0.01). SBP and level of apelin-ir in plasma and cardiovascular tissues were negatively correlated. Long-term swimming training relieved the pathogenesis of hypertension and reversed the downregulation of the cardiovascular apelin/APJ system induced by hypertension, which suggests that the improving effect of exercise training on hypertension could be mediated by upregulating the cardiovascular apelin/APJ system.

Animals↗

PubMed2006

Intrauterine ethanol exposure results in hypothalamic oxidative stress and neuroendocrine alterations in adult rat offspring.

Prenatal ethanol (EtOH) exposure is associated with low birth weight, followed by increased appetite, catch-up growth, insulin resistance, and impaired glucose tolerance in the rat offspring. Because EtOH can induce oxidative stress, which is a putative mechanism of insulin resistance, and because of the central role of the hypothalamus in the regulation of energy homeostasis and insulin action, we investigated whether prenatal EtOH exposure causes oxidative damage to the hypothalamus, which may alter its function. Female rats were given EtOH by gavage throughout pregnancy. At birth, their offspring were smaller than those of non-EtOH rats. Markers of oxidative stress and expression of neuropeptide Y and proopiomelanocortin (POMC) were determined in hypothalami of postnatal day 7 (PD7) and 3-mo-old (adult) rat offspring. In both PD7 and adult rats, prenatal EtOH exposure was associated with decreased levels of glutathione and increased expression of MnSOD. The concentrations of lipid peroxides and protein carbonyls were normal in PD7 EtOH-exposed offspring, but were increased in adult EtOH-exposed offspring. Both PD7 and adult EtOH-exposed offspring had normal neuropeptide Y and POMC mRNA levels, but the adult offspring had reduced POMC protein concentration. Thus only adult offspring preexposed to EtOH had increased hypothalamic tissue damage and decreased levels of POMC, which could impair melanocortin signaling. We conclude that prenatal EtOH exposure causes hypothalamic oxidative stress, which persists into adult life and alters melanocortin action during adulthood. These neuroendocrine alterations may explain weight gain and insulin resistance in rats exposed to EtOH early in life.

Animals↗

PubMed2006

Temporal transcriptome of mouse ATDC5 chondroprogenitors differentiating under hypoxic conditions.

The formation of cartilage takes place in vivo in an environment of reduced oxygen tension. To study the effect of hypoxia on the process of chondrogenesis, ATDC5 mouse chondroprogenitor cells were induced to differentiate by the addition of insulin and cultured under ambient and hypoxic conditions corresponding to 21% and 1% O2 in the gas phase, respectively. The production of extracellular proteoglycans as well as the transcriptional profile of 104 selected genes was determined by real-time RT-PCR. Hypoxia alone induced early chondrogenesis as evidenced by the synthesis of glycosaminoglycans and expression of aggrecan and collagen type II genes. Surprisingly, however, hypoxic incubation of insulin-treated cells delayed and suppressed the insulin-mediated early chondrogenesis and almost completely blocked hypertrophic differentiation. Analysis of the gene expression yielded several clues as to the mechanisms involved. In addition, a group of genes was identified that have not previously been associated with hypoxia, including Ak4, Akt3, Col X, Fmod, Ier3, IGFbp4, MafF, Mxi1, Rcor2, Rras, Sox6, Tnni2, Wnt5a, and Zfp313.

Aggrecans↗

PubMed2006

Determination of trace lead by solid substrate room temperature phosphorescence enhancing method based on heavy atom effect and dissoluble manganese supramolecule containing rhodamine 6G luminescent particles.

Dissoluble manganese supramolecule containing rhodamine 6G luminescent particles (M2) are synthesized, based on dissoluble manganese supramolecule (M1) doping rhodamine 6G (R.6G), by crystalline method. The particle diameters of M1 and M2 determined by ETM are both of micron degree. M1 and M2 can emit solid substrate room temperature phosphorescence (SS-RTP) on filter paper. The transition probability from the singlet state (S1) to triplet state (T1) of the luminescent molecules was greatly enhanced, based on the increment of luminescent molecules for each spot and the heavy atom effect of certain amount of Pb2+. As a result, the phosphorescence intensity (Ip) of M2 was increased sharply, and the enhancing value of phosphorescence intensity (DeltaIp) is directly proportional to the concentration of Pb2+. Thus, a new method of SS-RTP enhancing for the determination of trace lead is established based on manganese supramolecule containing rhodamine 6G luminescent particles. The linear range of this method is 0.0040-0.400 pg spot-1 of Pb2+ (corresponding concentration, 0.01-1.0 ng mL-1; sample volume, 0.4 microL spot-1), with a detection limit (LD) of 0.0011 pg spot-1 (corresponding concentration, 2.8x10(-12) g mL-1 of Pb2+, n=11). For the working solutions containing 0.0040 and 0.40 ng mL-1 of Pb2+, they were determined repeatedly for seven times, respectively. The R.S.D.s were 3.2 and 3.8%, respectively. This method has good repeatability, sensitivity and high precision. It has been applied to the determination of trace lead in human hair and tea samples with satisfactory results.

Cations, Divalent↗

PubMed2006

Cloning and developmental expression of the DEC1 ortholog gene in zebrafish.

DEC1/STRA13/SHARP2 is a transcription factor of the bHLH family that has been suggested to play key roles in mammalian cell differentiation, the cell cycle and circadian regulation. However, the function of the DEC1 gene during embryogenesis is not well understood. In the present study, we cloned a zebrafish ortholog of the human DEC1 gene and analyzed its expression during development of zebrafish. The predicted protein encoded by zebrafish DEC1 consists of 403 amino acids, and shares 59%, 60% and 59% identity in overall amino acid sequence with human DEC1, mouse STRA13 and rat SHARP2, respectively. Zebrafish DEC1 contains a bHLH domain exhibiting 97% identity with that of the mammalian ortholog. During zebrafish embryogenesis, DEC1 is expressed in a strong ubiquitous manner before early segmentation. At 15-72 hpf, DEC1 shows a specific and dynamic expression in the developing eyes, somites, pineal gland (epiphysis), heart, brain, spinal cord, notochord, pronephric duct, common cardinal vein and blood cells. In older zebrafish, DEC1 also is expressed in multiple tissues including the brain, eye, gut, liver and pancreas. Our data provide evidence that expression of DEC1 is evolutionally conserved in zebrafish.

Amino Acid Sequence↗

PubMed2006

[Effects of tissue inhibitor of matrix metalloproteinases-4 on the activities of matrix metalloproteinases and collagen of artery: experiment with rats].

OBJECTIVE: To investigate the effects of tissue inhibitor of matrix metalloproteinases-4 (TIMP-4) on the activities of matrix metalloproteinases (MMPs) and the collagen deposition. METHODS: Vascular smooth muscle cells (VSMCs) of rat thoracic aorta were cultured and divided into 3 groups: 2 groups to be transfected with adenovirus vector containing green fluorescence protein (AdGFP) or adenovirus vector containing TIMP-4 (AdTIMP-4), and one group un-transfected. Zymography and reverse zymography were used to detect the activities of MMP-2, MMP-9 and TIMP-4 in the supernatants. Male Wistar rats underwent balloon injury of common carotid artery and then divided into 3 groups: pure injury group, AdGFP transfection group, and AdTIMP-4. transfection group. Four and 28 days later 3 and 6 rats were killed in each group respectively to undergo microscopy and examination of the activities of MMP-2, MMP-9, and TIMP-4, and collagen quantity. RESULTS: The MMP-2 activity in the VSMC culture fluid supernatant of the AdTIMP-4 group was decreased dose-dependently, however, the activity of MMP-9 did not changed significantly among the 3 groups. Bands of activated MMP-2 and MMP-9 could not be examined in the normal vessel tissues. Four hours after the injury, the activity of MMP-2 was significantly increased in the pure injury group and AdGFP group, however, was significantly decreased in the AdTIMP-4 group. Four days later no MMP activity could be detected in either group. The neoformation of tunica intima was inhibited by 66% in the AdTIMP-4 group, The collagen quantity per vessel cell was 12.1 +/- 1.0 in the AdGFP group, not significantly different from that of in the AdTIMP-4 group (11.9 +/- 1, P > 0.05), and the collagen quantity per tunica adventitia cell in the AdTIMP-4 group was 118 +/- 13, not significantly different from that of the pure injury group (135 +/- 11, P > 0.05). CONCLUSION: The regulation of MMP/TIMP balance by TIMP-4 may control the metabolism of collagen and play an important role in the vascular repair process.

Adenoviridae↗

PubMed2006

Human prolyl-4-hydroxylase alpha(I) transcription is mediated by upstream stimulatory factors.

Prolyl-4-hydroxylase alpha(I) (P4Halpha(I)) is the rate-limiting subunit for P4H enzyme activity, which is essential for procollagen hydroxylation and secretion. In the current study, we have characterized the human P4Halpha(I) promoter for transcription factors and DNA elements regulating P4Halpha(I) expression. Using a progressive deletion cloning approach, we have constructed pGL3-P4Halpha(I) recombinant plasmids. We have identified a positive regulatory region at the positions of bp -184 to -97 responsible for approximately 80% of the P4Halpha(I) promoter efficiency. Three E-boxes were located within this region, and the E-box at position bp -135 explains most of the regulatory capacity. Upstream stimulatory factors (USF1/USF2) were shown to bind on the E-box using chromatin immunoprecipitation assay. Suppression of USF1 and/or USF2 using specific short interference RNA resulted in a significant reduction in P4Halpha(I) promoter activity, and overexpressed USF1 or USF2 increased P4Halpha(I) promoter activity significantly. Although transforming growth factor beta1 increased the USF1/USF2-E-box binding and P4Halpha(I) promoter activity, this up-regulatory effect can be largely prevented by USF1/USF2-specific short interference RNA. On the other hand, cigarette smoking extracts, which have been shown to suppress P4Halpha(I) expression, inhibited the binding between the USF1/USF2 and E-box, resulting in a reduced P4Halpha(I) promoter activity. Furthermore, the E-box on the P4Halpha(I) promoter appeared to indiscriminately bind with either USF1 or USF2, with a similar outcome on the promoter efficiency. In conclusion, our study shows that USF1/USF2 plays a critical role in basal P4Halpha(I) expression, and both positive (transforming growth factor beta1) and negative (cigarette smoking extract) regulators appear to influence the USF-E-box interaction and affect P4Halpha(I) expression.

Base Sequence↗

PubMed2006

Expression of HBcAg mutant with long internal deletion in Saccharomyces cerevisiae and observation of its self-assembly particles by atomic force microscopy (AFM).

An internally truncated C gene of adr hepatitis B virus core antigen with long internal deletion (aa81-aa116) (DeltaHBcAg with 36aa truncation) was expressed in Saccharomyces cerevisiae and the products (DeltarHBcAg) were purified from a crude lysate of the yeast by three steps: Sephrose CL-4B chromatography, sucrose step-gradient ultracentrifugation and CsCl-isopycnic ultracentrifugation. Results of ELISA test and density analysis of CsCl-isopycnic ultracentrifugation indicated that the purified products (DeltarHBcAg protein) with HBeAg antigenicity mainly located at the densities of 1.23 g ml-1. Observation and analysis of the purified DeltarHBcAg products by AFM indicated that the DeltarHBcAg (core) protein produced in S. cerevisiae could self-assemble into three or more size classes of core particles which exhibited a polymorphous distribution of DeltarHBcAg (core) particles. These different size classes of core particles mainly centred on the range whose mean diameter was from 10 nm to 48 nm, especially on the position of 11 nm, 15.6 nm and the range from 27 nm to 41 nm, respectively. Furthermore, the most number of core particles mainly centred on the range whose mean diameter was from 27 nm to 41 nm. These results above indicated that the truncated internal long fragment (aa81-aa116) probably had no effect on self-assembly of the HBcAg core particles which implied the internal length fragment (aa81-aa116) was not the sole domain for self-assembly of HBcAg dimer or the truncated HBcAg protein subunit formed the fresh interactive domain with each other. These initial results above by AFM analysis were very important for further research on the self-assembly, ultrastructure, subunit interaction and core internal deletion mutant (CIDM) function of HBcAg core particles.

Amino Acid Sequence↗