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Biomedical subjects

F Yao

Publications and source records attributed to F Yao.

At least 37 records · Page 2Linked to original sources
PubMed1998

Tetracycline repressor, tetR, rather than the tetR-mammalian cell transcription factor fusion derivatives, regulates inducible gene expression in mammalian cells.

This article describes the first (to our knowledge) tetracycline-inducible regulatory system that demonstrates that the tetracycline repressor (tetR) alone, rather than tetR-mammalian cell transcription factor fusion derivatives, can function as a potent trans-modulator to regulate gene expression in mammalian cells. With proper positioning of tetracycline operators downstream of the TATA element and of human epidermal growth factor (hEGF) as a reporter, we show that gene expression from the tetracycline operator-bearing hCMV major immediate-early enhancer-promoter (pcmvtetO) can be regulated by tetR over three orders of magnitude in response to tetracycline when (1) the reporter was cotransfected with tetR-expressing plasmid in transient expression assays, and (2) the reporter unit was stably integrated into the chromosome of a tetR-expressing cell line. This level of tetR-mediated inducible gene regulation is significantly higher than that of other repression-based mammalian cell transcription switch systems. In an in vivo porcine wound model, close to 60-fold tetR-mediated regulatory effects were detected and it was reversed when tetracycline was administered. Collectively, this study provides a direct implementation of this tetracycline-inducible regulatory switch for controlling gene expression in vitro, in vivo, and in gene therapy.

Animals↗
PubMed1998

In vivo gene transfer to skin and wound by microseeding.

BACKGROUND: Gene transfer to skin has many potential applications but lacks a safe, practical delivery method. This report presents a new technique, microseeding, for in vivo gene transfer to skin and wounds and for DNA-mediated vaccination. The plasmid DNA solution was delivered directly to the target cells of the skin by a set of oscillating solid microneedles driven by a modified tattooing device. MATERIALS AND METHODS: Skin and partial-thickness excisional wounds in pigs were microseeded with either hEGF expression plasmid or beta-galactosidase expression plasmid. Human EGF was also delivered by single injection or particle bombardment. hEGF expression in wound fluid and in target tissue was determined by ELISA with anti-hEGF-specific antibodies. Additionally, weanling pigs were microseeded with a hemagglutinin of swine influenza virus expression plasmid and production of anti-HA-specific antibodies was determined by blocking ELISA. RESULTS: hEGF expression in microseeded partial thickness wounds (5664 pg/site) and skin sites (969 pg/site) peaked 2 days after transfection being four- to seven-fold higher than gene transfer by a single intradermal injection and two- to three-fold higher than particle-mediated gene transfer. The beta-galactosidase-expressing cells were detected in dermis and epidermis. Pigs microseeded with HA expression plasmid were protected from infection by the Swine influenza virus. CONCLUSIONS: These results demonstrate that microseeding is a simple and effective method for in vivo gene transfer to skin and wounds and is more efficient than single injection and particle-mediated gene transfer.

Animals↗
PubMed1998

Tissue engineering of skin.

The skin plays a crucial role in protecting the integrity of the body's internal milieu. The loss of this largest organ is incompatible with sustained life. In reconstructive surgery or burn management, substitution of the skin is often necessary. In addition to traditional approaches such as split- or full-thickness skin grafts, tissue flaps and free-tissue transfers, skin bioengineering in vitro or in vivo has been developing over the past decades. It applies the principles and methods of both engineering and life sciences toward the development of substitutes to restore and maintain skin structure and function. Currently, these methods are valuable alternatives or complements to other techniques in reconstructive surgery. This review article deals with the evolution and current approaches to the development of in vitro and in vivo epidermis and dermis.

Biomedical Engineering↗
PubMed1998

HLA-DQA1, -DQB1 polymorphism distribution in Chinese women with pregnancy induced hypertension in Shanghai area.

OBJECTIVE: To explore the association of human leukocyte antigen (HLA) with pregnancy induced hypertension (PIH). METHODS: We oligotyped HLA-DQA1, -DQB1 locus of 30 Chinese PIH families and 14 control families in Shanghai area by polymerase chain reaction-sequence specific oligonucleotide (PCR-SSO) hybridization method (probes labeled by nonradioactive technique). RESULTS: Compared with the control group, the allelic frequency of HLA-DQB1 * 0502 was significantly higher in PIH couples, and the sharing of HLA-DQA1 increased in PIH couples as well. No difference was found in HLA-DQA1 allelic frequencies or HLA-DQB1 sharing between the two groups. Analysis of neither HLA-DQA1 nor HLA-DQB1 allelic frequencies in PIH patients and PIH mother-and-fetuses showed positive result. CONCLUSION: HLA-DQB1 * 0502 may be a marker of susceptibility to PIH. DQB1 * 0502 itself or some gene(s) located in HLA class II region and in linkage disequilibrium with 0502 affect maternal T cell immunity during pregnancy. The increase of compatibility in HLA-D region causes the production of blocking antibody to decrease.

Adult↗
PubMed1998

[Synthesis and cytotoxicity of the new taxoids].

Sinenxan A is a new biosynthetic taxane. In order to search for new taxol analogues with cytotoxicity, sinenxan A was modified and studied. Ten new taxoids containing the tetrahydrofuran ring were esterified at C14 or C10 positions with taxol/taxotere side chain. Preliminary pharmacological tests showed that some of the compounds have certain cytotoxicity on KB, HCT-8 and A2780. Meanwhile, the relationship between their structures and activity was discussed.

Antineoplastic Agents, Phytogenic↗
PubMed1998

[Cloning and characterization of melanin biosynthesis genes from Streptomyces lincolnensis 78-11].

Using mel C1-C2 gene from pIJ702 as probe to hybridize wity chromosome DNA of S. lincolnensis 78-11, a series of positive bands were found, including a 3.2 kb BanHI fragment and a 2.6 kb SphI fragment. So a gene library of S. lincolnensis, containg 3.0-3.5 kb BamHI fragment, was constructed. Melanin biosynthesis genes, mel C1 and mel C2, were then cloned and the fragment, inserted in recombinant plasmid pRSB336 were sequenced. Among five ORFs of the 3152 bp BamHI fragment, mel C1 and mel C2 were highly homologous to the respective genes of the other three strains of Streptomyces, morever, the product of mel C2 gene from S. lincolnensis 78-11 was slightly homologous to tyrosinase from human-being by 17.3% and from rat by 24.5%. All of the above results indicated that mel C1, mel C2 and orf 3 constitute the melanin biosynthesis operon of S. lincolnensis. In the further study of the cloned melanin biosynthesis genes of S. lincolnensis 78-11, recombinant plasmids, pPZ518 and pPZ519, which contain promoter of neo gene and mel gene in different orientation respectively, were constructed, and then were used to transform S. lividans TK23. 12 transformants of pPZ518, selected at random, secreted light brown pigment in R2YE culture medium, but all transformants of pPZ519 and pES1 were white.

Amino Acid Sequence↗
PubMed1996

Fibrolamellar hepatocellular carcinoma. Case reports and a review of the literature.

Fibrolamellar hepatocellular carcinoma is an uncommon malignancy seen in young adults without underlying liver disease. Physical signs are minimal and laboratory values are noncontributory. Diagnosis is suggested by clinical history, supported by radiographic studies, and confirmed by histologic examination. Individuals with fibrolamellar carcinoma generally have a greater survival than those with hepatocellular carcinoma. Although most patients with fibrolamellar carcinoma undergo curative surgery, two of the three patients we report had inoperable tumors.

Adult↗
PubMed1995

Temporal profile of in situ DNA fragmentation after transient middle cerebral artery occlusion in the rat.

We measured the temporal profile and anatomic distribution of cells exhibiting DNA fragmentation at various durations of reperfusion after middle cerebral artery (MCA) occlusion in the rat. Focal cerebral ischemia was induced in male Wistar rats (n = 62) using an intraluminal monofilament blockade of the MCA. After 2 h of MCA occlusion, the animals were killed at different durations of reperfusion (0.5, 3, 6, 9, and 12 h and 1, 2, 4, 7, 14, 21, and 28 days, n = 4 per time point). Sham-operated rats (n = 4) and normal rats not subjected to any surgical procedure (n = 4) were used as controls. Coronal brain sections (5 microns) were analyzed, using an in situ ApopTag kit, hematoxylin and eosin, and immunohistochemical double-staining methods. Six rats subjected to 2 h of MCA occlusion were killed at 24 h for measurement of DNA fragmentation by gel electrophoresis. Our data indicate that within a coronal section, DNA fragmentation was present in zero to three cells in each hemisphere of normal and sham-operated rats as well as in the contralateral hemisphere of ischemic rats. The number of cells exhibiting DNA fragmentation increased as early as 0.5 h (8 +/- 6), peaked at 24-48 h (213 +/- 59), and persisted for 4 weeks (10 +/- 2) after onset of reperfusion (p < 0.01). Groups of cells exhibiting DNA fragmentation (> 95% neurons) were located primarily in the inner boundary zone of the infarct. With use of gel electrophoresis, purified DNA obtained from the ischemic tissue exhibited the characteristic nucleosome ladder pattern associated with apoptosis.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

An activity specified by the osteosarcoma line U2OS can substitute functionally for ICP0, a major regulatory protein of herpes simplex virus type 1.

Among the five immediate-early regulatory proteins of herpes simplex virus (HSV) type 1, only ICP0 is capable of activating all kinetic classes of viral genes. Consistent with its broad transactivating activity, ICP0 plays a major role in enhancing the reactivation of HSV from latency both in vivo and in vitro. Although not essential for viral replication, ICP0 confers a significant growth advantage on the virus, especially at low multiplicities of infection. In this report we describe the expression of a novel activity by the osteosarcoma cell line U2OS that can substitute functionally for ICP0. Compared with Vero cells, both U2OS cells and cells of the ICP0-expressing line 0-28 significantly enhanced the plating efficiency of an ICP0 null mutant, 7134. In contrast, the plating efficiencies of the wild-type virus in all three cell types were similar. Single-step growth experiments demonstrated that the yield of 7134 in U2OS cells was severalfold higher than that in 0-28 cells and about 100-fold higher than that in Vero cells. In order to identify the viral genes whose expression is enhanced by the activity in U2OS cells, levels of expression of selected viral proteins in extracts of Vero and U2OS cells were compared by Western blot (immunoblot) analysis following low-multiplicity infection. At a multiplicity of 0.1 PFU per cell, the levels of expression of the immediate-early protein ICP4 and the early protein gD in 7134-infected U2OS cells were significantly higher than those in 7134-infected Vero cells. When infections were carried out at a multiplicity of 1 PFU per cell, however, no major differences in the levels of expression of these proteins in U2OS and Vero cells were observed. Cycloheximide reversal experiments demonstrated that the cellular activity expressed in U2OS cells that promotes high-level expression of ICP4 is not synthesized de novo but appears to exist as a preformed protein(s). To confirm this observation and to determine whether, like immediate-early genes, early, delayed-early, and late viral genes are also responsive to the cellular activity, transient-expression assays were performed. The results of these tests demonstrated that basal levels of expression from immediate-early, early, and delayed-early promoters, but not that from a late promoter, were significantly higher in U2OS cells than in Vero cells and that this enhancement occurred in the absence of viral proteins.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1995

[A study on two gut hormones in breast milk].

OBJECTIVE: To determine if motilin and gastrin are present in breast milk and to measure their concentrations in human milk and cow milk. METHODS: The concentration of motilin was measured in 17 samples of human colostrum, 18 samples of human mature milk, 8 samples of cow colostrum and 20 samples of cow mature milk by radioimmunoassay. The concentration of gastrin in human milk was also determined by radioimmunoassay. RESULTS: Motilin concentration in human colostrum was the highest (416.34 +/- 183.95ng/L), being higher than that in human mature milk (272.91 +/- 148.73ng/L, that in cow colostrum (229.51 +/- 63.68ng/L) and that in cow mature milk (35.46 +/- 16.94ng/L). Evidently the difference in motilin concentration was very significant between human milk and cow milk. The gastrin concentration in human colostrum was 17.20 +/- 11.98ng/L, being higher than that in human mature milk (5.62 +/- 2.33ng/L). CONCLUSION: Human milk, especially human colostrum, contains high concentrations of motilin and gastrin. Breast feeding, especially early breast feeding, may promote the maturation of the developing gut in neonates and infants.

Adult↗
PubMed1994

Physical interaction between the herpes simplex virus type 1 immediate-early regulatory proteins ICP0 and ICP4.

The herpes simplex virus type 1 immediate-early protein ICP0 enhances expression of a spectrum of viral genes alone and synergistically with ICP4. To test whether ICP0 and ICP4 interact physically, we performed far-Western blotting analysis of proteins from mock-, wild-type-, and ICP4 mutant virus-infected cells with in vitro-synthesized [35S]Met-labeled ICP0 and ICP4 as probes. The ICP4 and ICP0 polypeptides synthesized in vitro exhibited molecular weights similar to those of their counterparts in herpes simplex virus type 1-infected cells, and the in vitro-synthesized ICP4 was able to bind to a probe containing the ICP4 consensus binding site. Far-Western blotting experiments demonstrated that ICP0 interacts directly and specifically with ICP4 and with itself. To further define the interaction between ICP0 and ICP4, we generated a set of glutathione S-transferase (GST)-ICP0 fusion proteins that contain GST and either ICP0 N-terminal amino acids 1 to 244 or 1 to 394 or C-terminal amino acids 395 to 616 or 395 to 775. Using GST-ICP0 fusion protein affinity chromatography and in vitro-synthesized [35S]Met-labeled ICP0 and ICP4, ICP4 was shown to interact preferentially with the fusion protein containing ICP0 C-terminal amino acids 395 to 775, whereas ICP0 interacted efficiently with both the N-terminal GST-ICP0 fusion proteins and the C-terminal GST-ICP0 fusion proteins containing amino acids 395 to 775. Fusion protein affinity chromatography also demonstrated that the C-terminal 235 amino acid residues of ICP4 are important for efficient interaction with ICP0. Collectively, these results reveal a direct and specific physical interaction between ICP0 and ICP4.

Amino Acid Sequence↗
PubMed1993

Capillary electrophoretic separation of human recombinant erythropoietin (r-HuEPO) glycoforms.

Free zone capillary electrophoresis was investigated for the separation of glycoforms in recombinant erythropoietin (r-HuEPO). Reduction of electroendoosmotic flow through the addition of 1,4-diaminobutane and in the presence of urea resulted in optimum separation of all of the major glycoforms. Incubation of r-HuEPO with neuraminidase caused the slowest migrating species to diminish first in response, indicating that these contained the highest numbers of sialic acids. Additional evidence was obtained from spiking r-HuEPO with individual glycoforms isolated by preparative isoelectric focusing. These results indicate that the separation occurs in a predictable manner in order of increasing number of sialic acids contained in the glycoforms. As a consequence, the electropherograms provide the same information as that obtained from conventional gel isoelectric focusing, although separating by a different mechanism.

Animals↗
PubMed1992

Association of ICP0 but not ICP27 with purified virions of herpes simplex virus type 1.

Recent studies have shown that ICP4, one of the major immediate-early proteins of herpes simplex virus type 1 is present within the tegument region of the virion (F. Yao and R. J. Courtney, J. Virol. 63:3338-3344, 1989). With monoclonal antibodies to two additional immediate-early proteins, ICP0 and ICP27, and Western blot (immunoblot) analysis, ICP0, but not ICP27, was also found to be associated with purified virus particles. In an effort to localize the ICP0 within the virion, purified virions were treated with trypsin in the presence and absence of detergent. The data suggest that ICP0 is located within the tegument region of the virion and is not localized in the envelope or within the nucleocapsid. The number of molecules of ICP0 per virion was estimated to be approximately 150.

Animals↗
PubMed1992

Limitations of liver surface US in the diagnosis of cirrhosis.

Ultrasound (US) of the liver surface with a high-frequency, small-parts, short-focused probe has been proposed as a method of diagnosing cirrhosis. US of the liver was performed in 50 consecutive patients undergoing diagnostic liver biopsy to assess the clinical usefulness of this noninvasive procedure in diagnosing hepatic cirrhosis. Eight patients had histologically proved cirrhosis, and 42 had no histologic evidence of cirrhosis. Seven of the eight patients with cirrhosis had a normal liver surface at US, and five of the 42 patients without cirrhosis had an abnormal liver surface. US of the liver surface with this probe was not reliable in this heterogeneous patient population.

Humans↗