Biomedical subjects

Tong Cheng

Publications and source records attributed to Tong Cheng.

At least 19 recordsLinked to original sources

PubMed2006

Activated protein C inhibits tissue plasminogen activator-induced brain hemorrhage.

Brain hemorrhage is a serious complication of tissue plasminogen activator (tPA) therapy for ischemic stroke. Here we report that activated protein C (APC), a plasma serine protease with systemic anticoagulant, anti-inflammatory and antiapoptotic activities, and direct vasculoprotective and neuroprotective activities, blocks tPA-mediated brain hemorrhage after transient brain ischemia and embolic stroke in rodents. We show that APC inhibits a pro-hemorrhagic tPA-induced, NF-kappaB-dependent matrix metalloproteinase-9 pathway in ischemic brain endothelium in vivo and in vitro by acting through protease-activated receptor 1. The present findings suggest that APC may improve thrombolytic therapy for stroke, in part, by reducing tPA-mediated hemorrhage.

Animals↗

PubMed2006

Variants of green fluorescent protein GFPxm.

As research progresses, fluorescent proteins useful for optical marking will evolve toward brighter, monomeric forms that are more diverse in color. We previously reported a new fluorescent protein from Aequorea macrodactyla, GFPxm, that exhibited many characteristics similar to wild-type green fluorescent protein (GFP). However, the application of GFPxm was limited because GFPxm expressed and produced fluorescence only at low temperatures. To improve the fluorescent properties of GFPxm, 12 variants were produced by site-directed mutagenesis and DNA shuffling. Seven of these mutants could produce strong fluorescence when expressed at 37 degrees C. The relative fluorescence intensities of mutants GFPxm16, GFPxm18, and GFPxm19 were higher than that of EGFP (enhanced GFP) when the expression temperature was between 25 and 37 degrees C, and mutants GFPxm16 and GFPxm163 could maintain a high fluorescence intensity even when expressed at 42 degrees C. Meanwhile, at least 4 mutants could be successfully expressed in mammalian cell lines. The fluorescence spectra of 6 of the 12 mutants had a progressive red shift. The longest excitation-emission maximum was at 514/525 nm. In addition, 3 of the 12 mutants had two excitation peaks including an UV-excitation peak, while another mutant had only one UV-excitation peak.

Animals↗

PubMed2006

Hypervolemia of pregnancy is not maintained in mice chronically overexpressing angiotensinogen.

OBJECTIVE: Women who develop pre-eclampsia show significantly less hypervolemia of pregnancy, compared with controls. We have shown that chronically elevated angiotensinogen expression increases a woman's risk of developing pre-eclampsia. Our objective was to determine whether increased angiotensinogen expression is sufficient to cause failed hypervolemia. STUDY DESIGN: To isolate the effects of elevated angiotensinogen expression, we studied transgenic mice with either 2 or 3 copies of the murine angiotensinogen gene. Plasma volume was measured by Evans blue dye dilution, and kidney sections were immunostained for angiotensinogen and renin. RESULTS: Three-copy mice failed to maintain hypervolemia after midgestation (P < .01) and failed to up-regulate renin expression in the distal nephron, compared with 2-copy controls. Intrarenal angiotensinogen was up-regulated during pregnancy in both genotypes. CONCLUSION: Chronically elevated angiotensinogen expression is sufficient to cause failed hypervolemia of pregnancy. Whether this observation is related to failed up-regulation of distal tubule renin expression requires further study.

Angiotensinogen↗

PubMed2006

The promise of protein C.

Protein C, a vitamin K-dependent serine protease zymogen that circulates in plasma, is converted by limited proteolysis to activated protein C (APC) by the thrombin-thrombomodulin complex. APC exerts anticoagulant, antiinflammatory, cytoprotective, and antiapoptotic activities. Recombinant APC therapy reduces mortality in severe sepsis patients. This review summarizes data from clinical observations, from in vitro studies, and from animal models of focal ischemic injury that provide a compelling rationale for clinical trials of APC for ischemic stroke.

Animals↗

PubMed2006

RNA interference inhibits hepatitis B virus gene expression and replication in HepG2-N10 cells.

OBJECTIVE: RNA interference (RNAi) refers to the phenomenon of sequence-specific degradation of homologous mRNA induced by double-stranded RNA. It has been successfully utilized to down-regulate endogenous gene expression or suppress the replication of various pathogens in mammalian cells. In this study, the effect of vector-based small interfering RNA (siRNA) promoted by pSilencer2.0-U6 inhibit hepatitis B virus (HBV) replication in cell culture was evaluated. METHODS: Three fragments of short nucleic acids, respectively, targeting on S, X and C region of HBV genome were inserted into pSilencer vectors after they were annealed with their partly antisense strands. The recombination plasmids were pS, pX and pC. These expression plasmids were transfected into HepG2-N10 cells, a cell line which stably expresses hepatitis B virus surface antigen (HBsAg), hepatitis B virus e antigen (HBeAg) and adw2 subtype Dane particles. The effect of RNAi was evaluated from the changes of DNA, RNA and protein levels. Viral antigens were measured by ELISA. Viral mRNA was analyzed by RT-PCR. The covalent closed circular DNA and genome DNA of HBV secreted into the culture media were measured by quantitative real-time PCR. Analysis of variance was performed for the results. RESULTS: Vector-based RNA interference could potently reduce HBsAg (pS vs pN: 47%, pX vs pN: 30%, and pC vs pN: 25%, P < 0.001) and HBeAg (pX vs pN: 57% and pC vs pN: 66%, P < 0.001) expression in cell culture. Furthermore, RT-PCR analysis showed that viral mRNAs were effectively degraded, thus eliminating the messengers for protein expression as well as templates for reverse transcription (pS and pC vs pN, P < 0.001; pX vs pN, P = 0.003). Quantitative real-time PCR analysis of HBV DNA revealed that vector-based RNA interference can inhibit HBV replication efficiently (pS, pX and pC vs pN, P < 0.001). CONCLUSIONS: Our results indicate that RNAi can inhibit HBV gene expression and replication, and it might have the potential to revolutionize the treatment of HBV.

Cell Line↗

PubMed2006

[Establishment and application of human papillomavirus type 16 pseudovirions neutralization assay].

Human papillomaviruses (HPV) are causally associated with cervical cancer and genital warts. Lack of permissive and productive cell cultures for HPV has hindered the study of HPV and evaluation of virus-neutralizing antibodies. So generation of infectious virions in vitro is highly desirable. In this report, we got high titer infectious HPV16 pseudovirions by calcium phosphate co-transfection of codon optimized HPV16 capsid genes L1 and L2 and reporter plasmids into 293FT cell line. Electron micrograph indicated that the pseudovirions were morphologically similar with the intact HPV16 virions. To evaluate the feasibility of using the pseudovirions to identify neutralizing monoclonal antibodies (mAbs), pseudovirions were incubated with 2-fold gradient dilution of the well identified mAbs V5, E70, U4 and D9 and then used to infect 293FT cells preplated in 96-well tissue culture plate. The infection of pseudovirions could be inhibited by neutralizing mAbs V5, E70 and U4 that recognize surface conformational epitopes on L1 VLP, but not by mAb D9 that is reactive to a linear epitope buried in VLP, which indicated that the pseudovirions could be used to evaluate the neutralization efficiency of mono- and polyclonal antibodies. The pseudovirions were then employed to identify neutralizing mAbs from 18 mAbs generated previously in our lab, 8 of which were conformational and 10 were linear. PD1 and 3D10, both of which recognized conformational epitopes on L1 VLP, had obviously strong neutralizing efficiency, with the neutralizing titer reached 81,920 and 20,480 respectively, while none of the linear mAbs were neutralizing, which reflected that rare linear mAbs have neutralization activity. The mechanism of PD1 and 3D10 block the infection of HPV16 pseudovirions need to be further studied. The technologies about generation of HPV16 pseudovirions and screening neutralizing mAb in our report are economical and efficient, can be easily used in large scale. They pave the way for rapid and precise evaluation of the protection efficiency of our prophylactic HPV vaccine being developed now.

Animals↗

PubMed2005

Hydrophobicity of reactive site loop of SCCA1 affects its binding to hepatitis B virus.

AIM: To investigate the role of SCCA2 and other SCCA1 molecules in the process of hepatitis B virus (HBV) binding to mammalian cells. METHODS: SCCA1 and SCCA2 were isolated from HepG2. Binding protein (BP) genes were obtained through PCR. Recombinant baculoviruses expressing SCCA1, SCCA2, BP, and different mutants were constructed and utilized to infect mammalian cells to investigate the binding ability of infected cells to HBV. RESULTS: A SCCA1 gene (A1) was isolated from HepG2, but it appeared to lack the binding ability of infected cells to HBV. Two mutants, A1-BP and BP-A1, were constructed by interchanging the carboxyl terminal of A1 and BP. Cells expressing A1-BP showed an increased virus binding capacity, but not BP-A1. Comparison of A1 sequence with the sequence of BP indicated the presence of only three amino acid changes in the carboxyl terminal, two of them were found in the reactive site loop (RSL) of SCCA1. Primary structure assay revealed that the hydrophobicity of BP and AJ515706 in this domain was strong, but A1 was relatively weak. Changing the aa349 of A1 from low hydrophobic glutamic acid to high hydrophobic valine enhanced HBV binding. In contrast, HBV binding was reduced by changing the aa349 of BP from valine to glutamic acid. CONCLUSION: The results suggest that the hydrophobicity of RSL of SCCA1 may play an important role in HBV binding to cells.

Antigens, Neoplasm↗

PubMed2005

Expression and immunoactivity of chimeric particulate antigens of receptor binding site-core antigen of hepatitis B virus.

AIM: To improve the immunogenicity of receptor binding site of hepatitis B virus (HBV) on preS1 antigen using HBV core antigen as an immuno-carrier. METHODS: One to 6 tandem copies of HBV preS1 (21-47) fragment were inserted into HBcAg at the sites of aa 78 and 82, and expressed in E.coli. ELISA, Western blot and animal immunization were used to analyze the antigenicity and immmunogenicity of purified particulate antigens. The ability to capture HBV by antibodies elicited by chimeric particles was detected with immuno-capture PCR. RESULTS: Recombinant antigens CI, CII, CIII carrying 1-3 copies of HBV preS1 (21-47) individually could form virus-like particles (VLPs), similar to HBcAg in morphology. But recombinant antigens carrying 4-6 copies of HBV preS1 (21-47) were poorly expressed in E.coli. Chimeric antigens were lacking of immunoreactivity with anti-HBc monoclonal antibodies (McAbs), but still reserved good immunoreactivity with anti-HBe McAbs. CI, CII, CIII could strongly react with anti-preS1 McAb, suggesting that preS1 (21-47) fragment was well exposed on the surface of chimeric VLPs. Three chimeric VLP antigens (CI, CII and CIII) could stimulate mice to produce high-level antibody responses, and their immunogenicity was stronger than non-particulate antigen 21-47*6, containing 6 copies of preS1 (21-47). Mouse antibodies to CI, CII and CIII were able to capture HBV virions in immuno-capture PCR assay in vitro. CONCLUSION: Chimeric particulate antigens of receptor binding site-core antigen of HBV can elicit strong antibody responses to preS1. They have a potential to be developed into prophylactic or therapeutic vaccines against HBV infection.

Animals↗

PubMed2005

Significance of urinary angiotensinogen in essential hypertension as a function of plasma renin and aldosterone status.

OBJECTIVE: This study was performed to test the significance of urinary angiotensinogen (UAGT) in essential hypertensive patients stratified as a function of plasma renin and aldosterone. METHODS AND RESULTS: A sample of 248 essential hypertensives, investigated under their usual sodium diet and either off-medication or under a standardized treatment, was separated into two groups on the basis of upright plasma active renin and aldosterone medians. Patients with plasma active renin and aldosterone below medians are referred to as the low renin-aldosterone essential hypertensive group (LRA-EH). Others subjects are defined as other essential hypertensives (O-EH). Blood pressure (BP) was recorded by 24-h ambulatory monitoring. UAGT was measured by a specific enzyme-linked immunosorbent assay for total angiotensinogen. Because UAGT was markedly increased in the presence of overt proteinuria (>/= 300 mg/24 h), proteinuric patients (n = 29) were excluded from subsequent analyses. UAGT was a significant predictor of systolic and diastolic BP in LRA-EH females (P < 0.01 and P = 0.05, respectively) but not in males. By contrast, urinary sodium excretion (P < 0.001) and maintenance of treatment (P = 0.002) were significant predictors of systolic BP in males. These correlations were not observed in O-EH, whether males or females. CONCLUSIONS: In the present study, UAGT stands as a strong predictor of BP in women with low plasma renin/aldosterone, suggesting an involvement of the tubular renin-angiotensin system in these subjects. Higher sodium intake or the need to maintain treatment may account in part for the lack of a similar relationship in males.

Aged↗

PubMed2005

[Hydrophobicity of reactive site loop of SCCA1 affects its binding to HBV].

Squamous cell carcinoma antigen 1 (SCCA1), a member of the ovalbumin family of serine protease inhibitors, includes several variants. It was reported that expression of two SCCA1 (BP and AJ515706) in cells results in increased binding of HBV to these cells by the interaction of the expressed BP and AJ515706 with HBV pre-S1 domain. In this study, a SCCA1 (A1) was isolated from HepG2, but it appears to lack this ability. A possible role of two mutants, A1-BP and BP-A1, constructed by interchanging the carboxyl terminal of A1 and BP, was investigated. Cells expressing A1-BP rather than BP-A1 showed an increased virus binding capacity. Comparison of A1 sequence with the sequence of BP indicated the presence of only three amino acid changes in the carboxyl terminal, two of them in the reactive site loop (RSL) of SCCA1. Primary structure analysis revealed that the hydrophobicity of BP and AJ515706 in this domain is higher than that of A1. Changing the aa349 of A1 from low hydrophobic glutamic acid to high hydrophobic valine enhanced HBV binding. In contrast, changing the aa349 of BP from valine to glutamic acid reduced HBV binding. Our finding suggests that the hydrophobicity of RSL of SCCA1 may play an important role in HBV binding to cells.

Antigens, Neoplasm↗

PubMed2004

APC and GSK-3beta are involved in mPar3 targeting to the nascent axon and establishment of neuronal polarity.

In developing hippocampal neurons in culture, the evolutionarily conserved polarity complex mPar3/mPar6/aPKC selectively accumulates at the tip of one, and only one, of the immature neurites of a neuron and thus specifies the axon and generates neuronal polarity. How mPar3/mPar6 is enriched at the tip of the nascent axon, but not the dendrites, is not fully understood. Here, we report that mPar3 forms a complex with adenomatous polyposis coli (APC) and kinesin superfamily (KIF) 3A, proteins that move along microtubules. In polarizing hippocampal neurons, APC selectively accumulates at the nascent axon tip and colocalizes with mPar3. Expression of dominant-negative C terminus deletion mutants of APC or ectopic expression of APC leads to dislocalization of mPar3 and defects in axon specification and neuronal polarity. In addition to spatial polarization of APC, the selective inactivation of the GSK-3beta activity at the nascent axon tip is required for mPar3 targeting and polarization and establishing neuronal polarity. These results suggest that mPar3 is polarized in developing neurons through APC- and kinesin-mediated transport to the plus ends of rapidly growing microtubules at the nascent axon tip, a process that involves a spatially regulated GSK-3beta activity.

Adenomatous Polyposis Coli Protein↗

PubMed2004

Tissue plasminogen activator neurovascular toxicity is controlled by activated protein C.

Although thrombolytic effects of tissue plasminogen activator (tPA) are beneficial, its neurotoxicity is problematic. Here, we report that tPA potentiates apoptosis in ischemic human brain endothelium and in mouse cortical neurons treated with N-methyl-D-aspartate (NMDA) by shifting the apoptotic pathways from caspase-9 to caspase-8, which directly activates caspase-3 without amplification through the Bid-mediated mitochondrial pathway. In vivo, tPA-induced cerebral ischemic injury in mice was reduced by intracerebroventricular administration of caspase-8 inhibitor, but not by caspase-9 inhibitor, in contrast to controls in which caspase-9 inhibitor, but not caspase-8 inhibitor, was protective. Activated protein C (APC), a serine protease with anticoagulant, anti-inflammatory and antiapoptotic activities, which is neuroprotective during transient ischemia and promotes activation of antiapoptotic mechanisms in brain cells by acting directly on endothelium and neurons, blocked tPA vascular and neuronal toxicities in vitro and in vivo. APC inhibited tPA-induced caspase-8 activation of caspase-3 in endothelium and caspase-3-dependent nuclear translocation of apoptosis-inducing factor in NMDA-treated neurons and reduced tPA-mediated cerebral ischemic injury in mice. Data suggest that tPA shifts the apoptotic signal in stressed brain cells from the intrinsic to the extrinsic pathway which requires caspase-8. APC blocks tPA's neurovascular toxicity and may add substantially to the effectiveness of tPA therapy for stroke.

Animals↗

PubMed2004

The immunoglobulin family member dendrite arborization and synapse maturation 1 (Dasm1) controls excitatory synapse maturation.

In the developing mammalian brain, a large fraction of excitatory synapses initially contain only N-methyl-D-aspartate receptor and thus are "silent" at the resting membrane potential. As development progresses, synapses acquire alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPA-Rs). Although this maturation of excitatory synapses has been well characterized, the molecular basis for this developmental change is not known. Here, we report that dendrite arborization and synapse maturation 1 (Dasm1), an Ig superfamily member, controls excitatory synapse maturation. Dasm1 is localized at the excitatory synapses. Suppression of Dasm1 expression by using RNA interference or expression of dominant negative deletion mutants of Dasm1 in hippocampal neurons at late developmental stage specifically impairs AMPA-R-mediated, but not N-methyl-D-aspartate receptor-mediated, synaptic transmission. The ability of Dasm1 to regulate synaptic AMPA-Rs requires its intracellular C-terminal PDZ domain-binding motif, which interacts with two synaptic PDZ domain-containing proteins involved in spine/synapse maturation, Shank and S-SCAM. Moreover, expression of dominant negative deletion mutants of Dasm1 leads to more immature silent synapses. These results suggest that Dasm1, as a transmembrane molecule, likely provides a link to bridge extracellular signals and intracellular signaling complexes in controlling excitatory synapse maturation.

Animals↗

PubMed2004

A rapid and efficient method to express target genes in mammalian cells by baculovirus.

AIM: To investigate the modification of baculovirus vector and the feasibility of delivering exogenous genes into mammalian cells with the culture supernatant of Spodoptera frugiperta (Sf9) cells infected by recombinant baculoviruses. METHODS: Two recombinant baculoviruses (BacV-CMV-EGFPA, BacV-CMV-EGFPB) containing CMV-EGFP expression cassette were constructed. HepG2 cells were directly incubated with the culture supernatant of Sf9 cells infected by recombinant baculoviruses, and reporter gene transfer and expression efficiencies were analyzed by flow cytometry (FCM). The optimal transduction conditions were investigated by FCM assay in HepG2 cells. Gene-transfer and expression efficiencies in HepG2 or CV1 cells by baculovirus vectors were compared with lipofectAMINE, recombinant retrovirus and vaccinia virus expression systems. Twenty different mammalian cell lines were used to investigate the feasibility of delivering exogenous genes into different mammalian cells with the culture supernatant of infected Sf9 cells. RESULTS: CMV promoter could directly express reporter genes in Sf9 cells with a relatively low efficiency. Target cells incubated with the 1:1 diluted culture supernatant (moi=50) for 12 h at 37 degrees C could achieve the highest transduction and expression efficiencies with least impairment to cell viability. Under similar conditions the baculovirus vector could achieve the highest gene-transfer and expression efficiency than lipofectAMINE, recombinant retrovirus and vaccinia virus expression systems. Most mammalian cell lines could be transduced with recombinant baculovirus. In primate adherent culture cells the recombinant baculovirus could arrive the highest infection and expression efficiencies, but it was not very satisfactory in the cell lines from mice and suspended culture cells. CONCLUSION: Mammalian cells incubated with the culture supernatant of infected Sf9 cells could serve as a very convenient way for rapid and efficient expression of foreign genes in mammalian cells, but it might be more suitable for primate adherent culture cells.

Animals↗

PubMed2004

Activated protein C prevents neuronal apoptosis via protease activated receptors 1 and 3.

Activated protein C (APC), a serine protease with anticoagulant and anti-inflammatory activities, exerts direct cytoprotective effects on endothelium via endothelial protein C receptor-dependent activation of protease activated receptor 1 (PAR1). Here, we report that APC protects mouse cortical neurons from two divergent inducers of apoptosis, N-methyl-D-aspartate (NMDA) and staurosporine. APC blocked several steps in NMDA-induced apoptosis downstream to nitric oxide, i.e., caspase-3 activation, nuclear translocation of apoptosis-inducing factor (AIF), and induction of p53, and prevented staurosporine-induced apoptosis by blocking caspase-8 activation upstream of caspase-3 activation and AIF nuclear translocation. Intracerebral APC infusion dose dependently reduced NMDA excitotoxicity in mice. By using different anti-PARs antibodies and mice with single PAR1, PAR3, or PAR4 deletion, we demonstrated that direct neuronal protective effects of APC in vitro and in vivo require PAR1 and PAR3. Thus, PAR1 and PAR3 mediate anti-apoptotic signaling by APC in neurons, which may suggest novel treatments for neurodegenerative disorders.

Animals↗

PubMed2004

Activated protein C and ischemic stroke.

OBJECTIVE: To summarize clinical observations, animal model experimentation, and in vitro studies that advance knowledge of the protein C system, including activated protein C (APC), in the setting of ischemic stroke. DATA SOURCE: Narrative review of selected published primary basic and clinical literature from MEDLINE for 2000-2003. DATA SYNTHESIS: Low levels of plasma APC and a poor response to APC in clotting assays may be markers or risk factors for ischemic stroke. Ischemia during routine endarterectomy causes APC generation in the affected region of the human brain. The prospective epidemiologic Atherosclerosis Risk in Communities (ARIC) study reported that plasma protein C may be protective for ischemic stroke. In murine models of focal cerebral ischemia, APC provided remarkable anti-inflammatory and neuroprotective effects in vivo and increased survival at 24 hrs. Recent in vitro and in vivo studies provide remarkable insights into mechanisms of the neuroprotective activities of APC. Independent of its well-known anticoagulant activity, APC acts directly on cells and alters gene expression profiles, inhibits apoptosis, and down-regulates inflammation. These effects require protease-activated receptor-1 and the endothelial protein C receptor. In an in vitro model involving hypoxia-induced apoptosis of human brain endothelial cells, protease-activated receptor-1 and endothelial protein C receptor were required for APC to exert its anti-apoptotic effects. In these cells, APC blunts hypoxia-induced increases in p53 messenger RNA and protein, reduces pro-apoptotic Bax, and increases anti-apoptotic Bcl-2, thereby inhibiting mitochondrial-dependent apoptosis. Murine ischemic stroke model studies have provided in vivo evidence for the physiologic roles of protease-activated receptor-1 and endothelial protein C receptor in the neuroprotective activities of APC. Because the low doses required for recombinant murine APC to provide neuroprotection do not cause observable anticoagulant effects, the in vivo neuroprotective action of APC seems, at least in part, to be independent of its anticoagulant activity and is likely to involve its anti-apoptotic activity. CONCLUSIONS: There is compelling evidence that ischemic stroke is an attractive target for therapy with APC.

Animals↗

PubMed2004

Expression of angiotensinogen in proximal tubule as a function of glomerular filtration rate.

BACKGROUND: Proximal tubule (PT) angiotensinogen (AGT) is part of a tubular renin-angiotensin system (RAS) that participates in the regulation of sodium reabsorption along the entire nephron. Physiologic maneuvers affecting AGT expression in PT also affect systemic RAS. Here, we tested the hypothesis that PT AGT is regulated by increased glomerular filtration rate (GFR). METHODS: Complete unilateral nephrectomy (UNX) in mice was used to induce a sustained increase in GFR in the remaining kidney. AGT expression was monitored by quantitative reverse transcription-polymerase chain reaction (RT-PCR). AGT protein in PT was investigated by semiquantitative histology. We also measured AGT concentration in plasma and in 24-hour urine by a specific enzyme-linked immunosorbent assay (ELISA). RESULTS: Seven weeks after nephrectomy, UNX animals exhibited a 2-fold increase in tubular AGT mRNA (P <.001) compared with sham-operated control animals. The proportion of PT sections exhibiting AGT immunostaining was significantly increased at day 3 (P <.05), and remained elevated at seven weeks (UNX = 0.63 +/- 0.09, sham = 0.38 +/- 0.02, P <.01), revealing recruitment of AGT-producing cells along the PT. AGT excretion in final urine corrected for creatinine and kidney weight was also elevated by UNX at seven weeks (UNX = 209 +/- 42 pmol/mg/g, sham = 147 +/- 29 pmol/mg/g, P <.05), with no difference in plasma AGT between UNX and control animals. CONCLUSION: These observations suggest that AGT expression in PT adapts in the long-term to changes in GFR. In the UNX model, urinary AGT excretion is also elevated as a consequence of increase in net tubular flow.

Adaptation, Physiological↗

PubMed2004

[Mammalian gene-transfer and expression efficiencies of baculovirus bacV-CMV-EGFPA].

It has been reported that baculoviruses could serve as a new gene-transfer vehicle for mammalian cells. We have previously constructed recombinant baculovirus BacV-CMV-EGFPA and have proven that mammalian cells could be effectively infected by the recombinant baculovirus. In this report, we studied the efficiency of baculovirus to deliver exogenous gene into twenty mammalian cells, including twelve human cell lines (WI-38, Hela, HepG2, 293, PLC/PRF/5, 143B, MCF-7, BGC-223, DMS 114, CNE, Raji, LCL-cm), seven murine cell lines (BNL 1ME A.7R.1, CHO-K1, L-929, JC, PT67, NIH3T3, P815) and one monkey cell line (CV1). Results showed that most mammalian cell lines could be transduced by the recombinant baculovirus, the transduction efficiencies of the human and monkey cell lines were markedly higher than that of murine cell lines, and the transduction efficiencies in adherent culture cell lines higher than that of suspend culture cell lines, implying that the infection efficiency of the baculovirus may be correlative with the organism used and the growth properties of the cell lines. The plasmid pcDNA3. 1-EGFP, which contains the CMV promoter and EGFP reporter gene, was next transfected by LipofectAMINE into a number of mammalian cells, especially those cells that were low in the baculovirus transfection. Results showed that the CMV promoter could effectively direct the expression of the reporter gene in these mammalian cells. Therefore the gene-expression efficiencies in different mammalian cell lines by the recombinant baculovirus which contains the same CMV promoter were dictated by the ability of the baculovirus to enter the cell lines. This study suggested that the recombinant baculovirus vector is more suitable for gene expression in primate adherent culture cells than in murine cells and suspend culture cells.

Animals↗