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R H Xu

Publications and source records attributed to R H Xu.

At least 37 records · Page 2Linked to original sources
PubMed1996

Involvement of Ras/Raf/AP-1 in BMP-4 signaling during Xenopus embryonic development.

Previously, we elucidated the role of bone morphogenetic protein 4 (BMP-4) in the dorsal-ventral patterning of the Xenopus embryo by using a dominant negative mutant of the BMP-4 receptor (DN-BR). The present paper describes the involvement of Ras, Raf, and activator protein 1 (AP-1) in BMP-4 signaling during Xenopus embryonic development. The AP-1 activity was determined by injecting an AP-1-dependent luciferase reporter gene into two-cell-stage Xenopus embryos and measuring the luciferase activity at various developmental stages. We found that injection of BMP-4 mRNA increased AP-1 activity, whereas injection of DN-BR mRNA inhibited AP-1 activity. Similar inhibitory effects were seen with injection of mRNAs encoding dominant negative mutants of c-Ha-Ras, c-Raf, or c-Jun. These results suggest that the endogenous AP-1 activity is regulated by BMP-4/Ras/Raf/Jun signals. We next investigated the effects of Ras/Raf/AP-1 signals on the biological functions of BMP-4. DN-BR-induced dorsalization of the embryo, revealed by the formation of a secondary body axis or dorsalization of the ventral mesoderm explant analyzed by histological and molecular criteria, was significantly reversed by coinjection of [Val12]Ha-Ras, c-Raf, or c-Jun mRNA. Furthermore, the BMP-4-stimulated erythroid differentiation in the ventral mesoderm was substantially inhibited by coinjection with the dominant negative c-Ha-Ras, c-Raf, or c-Jun mutant. Our results suggest the involvement of Ras/Raf/AP-1 in the BMP-4 signaling pathway.

Animals↗
PubMed1996

The protective role of the immunomodulator AS101 against chemotherapy-induced alopecia studies on human and animal models.

The immunomodulator AS101 has been demonstrated to exhibit radioprotective and chemoprotective effects in mice. Following phase-I studies, preliminary results from phase-II clinical trials on non-small-cell-lung-cancer patients showed a reduction in the severity of alopecia in patients treated with AS101 in combination with chemotherapy. To further substantiate these findings, the present study was extended to include 58 patients treated either with the optimal dose of 3 mg/m2 AS101 combined with carboplatin and VP-16, or with chemotherapy alone. As compared with patients treated with chemotherapy alone, there was a significant decrease in the level of alopecia in patients receiving the combined therapy. The newly developed rat model was used to elucidate the protective mechanism involved in this effect. We show that significant prevention of chemotherapy-induced alopecia is obtained in rats treated with Ara-C combined with AS101, administered i.p. or s.c. or applied topically to the dorsal skin. We show that this protection by AS101 is mediated by macrophage-derived factors induced by AS101. Protection by AS101 can be ascribed, at least in part, to IL-1, since treatment of rats with IL-1 RA largely abrogated the protective effect of AS101. Moreover, we demonstrate that in humans there is an inverse correlation between the grade of alopecia and the increase in IL-1 alpha. In addition, protection by AS101 could be related to PGE2 secretion, since injection of indomethacin before treatment with AS101 and Ara-C partly abrogated the protective effect of AS101. To assess the ability of AS101 to protect against chemotherapy-induced alopecia, phase-II clinical trials have been initiated with cancer patients suffering from various malignancies.

Alopecia↗
PubMed1996

Sp1 binding plays a critical role in Erb-B2- and v-ras-mediated downregulation of alpha2-integrin expression in human mammary epithelial cells.

The human alpha2-integrin gene is transcriptionally downregulated in a nontumorigenic human mammary epithelial cell line, MTSV1-7, and its clonal variant HB2, overexpressing the Erb-B2 oncogene. In this study, we have used deletion mutations within the alpha2-integrin promoter inserted 5' of the chloramphenicol acetyltransferase or luciferase reporter genes to identify the element that is responsible for the Erb-B2-mediated downregulation. The results of the transient-transfection assay showed that the Sp1 binding element located in the core region (positions --64 to +1) of the alpha2-integrin promoter plays an essential role in the alpha2-integrin promoter activity and its downregulation by Erb-B2. By gel shift assay, we have demonstrated that this element binds with a high degree of affinity not only to Sp1, but also to Sp3. The downregulation of the alpha2-integrin promoter activity could also be achieved by overexpression of v-Hras (v-ras), suggesting that the signals generated by Erb-B2, which lead to downregulation of the alpha2-integrin gene expression, may proceed through the ras pathway. Both the Erb-B2- and the v-ras-overexpressing cells exhibited a Sp1 DNA binding activity lower than that of the parental line, while the relative levels of Sp1 protein in these cells were not altered. The Erb-B2- and v-ras-mediated downregulation could be reversed by the overexpression of Sp1 and by a dominant negative variant of ras (rasN17), confirming the importance of Sp1 and the ras pathway. The inhibitory effects of Erb-B2 on transcriptional activity of the alpha2-integrin promoter were observed in transient-cotransfection assays using alpha2-integrin reporter plasmids and plasmids expressing the Erb-B2 or v-ras oncogene. The same effects were seen when an alpha2-integrin reporter gene construct was transfected into MTSV1-7 or HB2 cells permanently overexpressing Erb-B2 or v-ras. The effects of Erb-B2 or v-ras on the transcriptional activity of the alpha2-integrin promoter were observed in nontumorigenic luminal epithelial cell lines (MTSV1-7 and HB2) as well as in the breast cancer cell line T47D. These data suggest that in luminal epithelial cells and the breast cancers which develop from them, the Erb-B2 proto-oncogene signaling leads to inhibition of (alpha)2(beta)1-integrin gene expression and could contribute to the disruption of tissue architecture seen in breast cancers.

Antigens, CD↗
PubMed1995

A dominant negative bone morphogenetic protein 4 receptor causes neuralization in Xenopus ectoderm.

Injection of DN-BR mRNA encoding a dominant negative type I receptor for bone morphogenetic protein 4 (BMP4) converted prospective ectoderm into neural tissue in Xenopus animal cap explants, in the absence of expression of mesodermal marker genes. The injected caps expressed a general neural marker NCAM and the forebrain marker opsin. Coinjection of wild-type BMP4 receptor mRNA completely reversed the neuralization by DN-BR. No expression of known neuralizing factors, i.e., noggin and follistatin, was detected in the DN-BR-injected animal caps. Furthermore, neuralization elicited by noggin or 3m, a LIM domain mutant of Xlim-1, was substantially inhibited by co-injection of BMP4 mRNA. Since BMP4 is expressed in the prospective ectoderm during gastrulation, our results suggest that the ventralizing factor BMP4 acts also as a physiological inhibitor of neuralization in the development of Xenopus ectoderm.

Animals↗
PubMed1995

A mutation in the pore of the sodium channel alters gating.

Ion permeation and channel gating are classically considered independent processes, but site-specific mutagenesis studies in K channels suggest that residues in or near the ion-selective pore of the channel can influence activation and inactivation. We describe a mutation in the pore of the skeletal muscle Na channel that alters gating. This mutation, I-W53C (residue 402 in the mu 1 sequence), decreases the sensitivity to block by tetrodotoxin and increases the sensitivity to block by externally applied Cd2+ relative to the wild-type channel, placing this residue within the pore near the external mouth. Based on contemporary models of the structure of the channel, this residue is remote from the regions of the channel known to be involved in gating, yet this mutation abbreviates the time to peak and accelerates the decay of the macroscopic Na current. At the single-channel level we observe a shortening of the latency to first opening and a reduction in the mean open time compared with the wild-type channel. The acceleration of macroscopic current kinetics in the mutant channels can be simulated by changing only the activation and deactivation rate constants while constraining the microscopic inactivation rate constants to the values used to fit the wild-type currents. We conclude that the tryptophan at position 53 in the domain IP-loop may act as a linchpin in the pore that limits the opening transition rate. This effect could reflect an interaction of I-W53 with the activation voltage sensors or a more global gating-induced change in pore structure.

Amino Acid Sequence↗
PubMed1995

Alternatively spliced forms of the alpha subunit of the epithelial sodium channel: distinct sites for amiloride binding and channel pore.

The amiloride-sensitive epithelial sodium channel (ENAC) consists of at least three subunits, alpha, beta, and gamma. Sodium conductance occurs when only the alpha subunit is expressed in Xenopus oocytes, but it is greatly enhanced by coexpression of all three subunits. All three subunits have two transmembrane domains. Whether the amiloride binding site exists in the extracellular portion or a transmembrane domain has not been established. Using reverse transcription-polymerase chain reaction in rat taste tissues, we have identified two alternatively spliced transcripts of ENAC (alpha ENACa and alpha ENACb) with deletions of nucleotides that introduce a premature stop codon and may result in proteins shortened by 199 and 216 amino acids, respectively, at the carboxyl terminus. Genomic Southern blots indicate that a single gene accounts for alpha ENAC and the alternatively spliced variants. Reverse transcription-polymerase chain reaction and RNase protection assays demonstrate that alpha ENACa is expressed to a lesser extent than alpha ENAC in kidney, lung, and taste tissues. alpha ENACa differs from alpha ENAC by a deletion in the second transmembrane domain. Despite this deletion, alpha ENACa expression in transfected human embryonic kidney 293 cells or CV-1 cells augments [3H]phenamil binding. The [3H]phenamil binding of alpha ENACa resembles that of alpha ENAC, being inhibited more potently by phenamil (Kd = 65 nM) than amiloride. Unlike alpha ENAC, expression of alpha ENACa in Xenopus oocytes fails to generate amiloride-sensitive Na+ or Li+ currents. These results suggest that the amiloride binding site resides on the extracellular loop of the alpha subunit of ENAC and not the putative second transmembrane domain, which forms a channel pore. Heterogeneity in alpha ENAC isoforms may contribute to the complexity of multimeric structures and functional variation of ENAC.

Alternative Splicing↗
PubMed1995

The cytoprotective effect of the immunomodulator AS101 against hydrochloride induced gastric lesions.

AS101 as a new immunomodulator has been shown to induce production of a variety of cytokines such as interleukin-1, interleukin-2, colony-stimulating factor, interferon-gamma and tumor necrosis factor, and demonstrate a potential chemo-protection from chemotherapy induced immunosuppression and hematopoietic toxicity in tumor bearing mice and cancer patients on phase I and II clinical trials. This study was designed to verify whether AS101 exerts a cytoprotective effect in rats and mice with gastric lesions induced by intragastric (i.g.) instilling of 0.6N hydrochloride (HCl). AS101 given intraperitoniously (i.p) 2h before a HCl administration markedly prevented HCl-induced gastric lesions both in rats and mice. It also accelerated the ulcer repair when given i.p. 1h after a HCl treatment. Indomethacin (IND), a cyclo-oxygenase inhibitor, given i.p. at a non-ulcerogenic dose of 5mg/kg 1h before AS101 administration, abolished its protective effect. Mechanistic analysis showed that the gastric cytoprotective property of AS101 appears to be mediated through the induction of prostaglandin E2 (PGE2) and epidermal growth factor (EGF), of which, both prevent the gastric mucosa from HCl ulceration while EGF also contributes to the promotion of ulcer repair. This study adds another cytoprotective property to the known immunomodulating role of AS101.

Adjuvants, Immunologic↗
PubMed1994

[Osteogenesis role played by dermal fibroblasts cultured in vitro].

Fibroblasts were harvested by in vitro culture of split-thickness skin of New-Zealand rabbit. Some opaque bone nodules were formed in the culture flasks when the fibroblasts underwent subculture for 8 days. Some of the nodules might enlarge, extend and form trabecula-like or flake-like structures, when the fibroblasts underwent subculture for 37 days. The above mentioned structures revealed golden-yellow fluorescence under the blue-violet light fluorescence microscope when the specimens were labeled with Tetracycline intravitally, denoting that they were newly formed bone tissues. In these structures, the calcium salt staining and collagen staining also showed positive reaction, fully reflecting the calcium and collagen composition. Osteogenesis caused by dermal fibroblast cultured in vitro was thus verified. There were evenly scattered oval spots in the osteogenic areas, after the specimens were treated with Alcina Blue-Sirius Red stain. Their diameter was about 10 microns, and they stained yellow color by Picric Acid. Therefore, these spots colored yellow with Picric Acid might be the dermal epithelia (and/or endothelia), which induced the fibroblasts to form the bone tissues.

Animals↗
PubMed1994

[Osteogenic potential of rabbit dermal fibroblasts cultured in vitro: a histochemical and scanning electron microscopic study].

Split-thickness rabbit skin was processed into small chips that were cultured in vitro and subjected to inverted phase contrast microscopic and scanning electron microscopic observation. The fibroblasts swam out of the skin chips and they increased in number rapidly and became confluenced. The cells exhibited squamous configuration, possessing arboreal bifurcations and forming multi-layer structure. The fibroblasts then excreted numerous minute granules, heaping up on and around the cells. Henceforth emerged on the cell surface fine needle-like crystals, that agglomerated with the granules to form nodules. The fibroblasts orientated themselves in a radiating pattern around the large nodules. Neighbouring nodules could be linked up into trabecular structure. Histochemical study of both the nodules and the trabecular structure with specific new bone labeller (Tetracycline and Alizarin red S) revealed positive reaction, denoting that the nodules and the trabecular structure produced by the fibroblasts were bone tissues. These results pointed out clearly to the osteogenic potential of the dermal fibroblasts.

Animals↗
PubMed1993

Synthesis and expression in Escherichia coli of a human neutrophil activating protein-1/interleukin-8 gene.

The complete gene coding for human neutrophil activating protein-1/interleukin-8 was synthesized using a semi-chemical semi-enzymatic method. The synthetic gene was then overexpressed in Escherichia coli under the temperature-regulated control of the PRPL tandem promoters. As determined by SDS-PAGE and densitometry, the overexpressed protein comprised up to 18.5% and 10.9% of the total soluble protein in E. coli cells grown in shake flasks and in batch fermentation, respectively. The recombinant NAP-1/IL-8 was then purified to > 95% homogeneity by gel filtration and cation exchange chromatography. The purified protein appeared as a single band on the SDS-PAGE gel and possessed potent chemotactic activity in the concentration of < 10 ng/ml, as assayed by the agarose plate method. An early skin reactivity was also observed when the pure NAP-1/IL-8 was injected subcutaneously into the rabbits. The N-terminal 36 amino acid sequence of the recombinant NAP-1/IL-8 was determined using the Edman method and was shown to be identical to that of the native protein.

Animals↗
PubMed1993

Effects of osteogenesis on dermal fibroblasts cultured in vitro.

Fibroblast were harvested from in vitro culture of split-thickness skin of New Zealand rabbit. Some opaque bone nodules were found in the culture flasks when the fibroblasts underwent subculture for 8 days. After a 37-day subculture of fibroblasts, some of the nodules might enlarge extend and form trabecular-like or flake-like structures. The above-mentioned structures revealed golden-yellow fluorescence under the blueviolet light fluorescence microscope when the specimens were labeled with tetracycline intravitally, denoting that they were newly formed bone tissues. Both calcium salt and collagen stainings for these structures showed positive reaction, fully reflecting the calcium and collagen composition. Osteogenesis caused by dermal fibroblast cultured in vitro was thus verified.

Animals↗
PubMed1992

Effects of cordyceps sinensis on natural killer activity and colony formation of B16 melanoma.

This paper reports the study on the effects of the ethanol extract of Cordyceps sinensis (CS-II), a potent herbal tonic, on murine and human in vitro natural killer cell (NK) activities and on murine in vivo NK activity (by 125I clearance assay), and on colony formation of B16 melanoma in mouse lungs. The results revealed that: 1. the in vivo and in vitro NK activities of mouse were both significantly augmented by intraperitoneal (ip) injection of CS-II. Besides, the inhibition of mouse NK activity by cyclophosphamide (Cy) was prevented following the administration of CS-II; 2. the in vitro NK activity of human peripheral blood mononuclear cells (PBMs) was elevated by preincubation of PBMs with CS-II; and 3. the colony formation of B16 melanoma in mouse lungs was reduced significantly by ip pretreatment of the mice with CS-II. This study indicates that CS-II may be used as an immunopotentiating agent in treating cancer and immunodeficient patients.

Adjuvants, Immunologic↗
PubMed1991

Transformation of chloroplasts with the psaB gene encoding a polypeptide of the photosystem I reaction center.

A chloroplast photosystem I reaction center mutation, ac-u-g-2.3, of Chlamydomonas reinhardtii has been complemented with a wild type psaB gene to restore photosynthetic competence. The mutation was mapped in the psaB coding sequence by chloroplast transformation using subcloned restriction fragments of psaB. The mutation was found to be a single base pair deletion resulting in a reading frame shift and premature termination of the polypeptide. Transformants were verified by insertion of a site-directed mutation which created a new restriction enzyme site. These transformations demonstrate the feasibility of insertion of site-directed mutations into the psaB gene in order to elucidate amino acid residues involved in photosystem I assembly and function.

Amino Acid Sequence↗
PubMed1991

Observation of the activity of factor VIII in the endometrium of women pre- and post-insertion of three types of IUDs.

The endometrial materials were obtained from 90 women who had been randomly inserted with three types of IUDs (Stainless steel ring, SS; copper T 220, TCu 220, and levonorgestrel-releasing device, LNG). An immunoperoxidase reaction, PAP method, with the antiserum of Factor VIII as the primary antibody, was carried out to detect the Factor VIII activity in the endometrial endothelium pre- (control) and post-insertion of the IUDs. The results revealed that: 1. There was a generalized lower Factor VIII activity in the endometrium of women post-insertion of IUDs (except LNG). 2. Comparison of the Factor VIII activity in the endometrium of women using different types of IUDs showed that the TCu type and the SS type decreased the activity significantly whereas the activity remained unchanged after 3-6 months' use of the LNG-IUD. The different types of IUDs seemed to influence the coagulation regulatory system in different ways; the lower Factor VIII activity, the more tendency to bleeding. 3. The Factor VIII activity in the endometrium of women using IUDs was lower in all phases of the menstrual cycle including the proliferative phase when the Factor VIII activity is normally high. It cannot be excluded that this could be a contributing factor to IUD-induced bleedings.

Adult↗
PubMed1991

[Histochemical study on effect of radix Salviae miltiorrhizae on growth of isolated cells from embryonic chicken frontal bone cultured in vitro].

12-day embryonic chicken frontal bone digested with trypsin to prepare the suspension of isolated bone cells. 3 x 10(6) cells were harvested altogether. The cells were divided equally into five parts. Then the Eagle medium and 0.1%, 0.2%, 0.4% and 0.8% Radix Salviae Miltiorrhizae in Eagle medium were added respectively and cultured in 5% CO2 incubator. It was observed under the inverted microscope every day. At the 26th day of culture, the cells were studied. The specimens were stained with H. E., Alcian Blue-Sirius Red, Alizarin Red S staining and alkaline phosphatase-acid phosphatase reaction for comparison. It was found that the maturation of the osteoblast-like cells could be accelerated by Radix Salviae Miltiorrhizae. Secretion of the collagenous substance, positive alkaline phosphatase reaction and deposition of mineral on the collagenous substance, forming bone nodules were found to be enhanced. But unduly high concentration of Radix Salviae Miltiorrhizae could lead to inhibition of osteoblast-like cell growth. The optimal concentration of Radix Salviae Miltiorrhizae was 0.2% in culture medium.

Animals↗
PubMed1990

Ultrastructural comparative study on endometrium of Chinese women using intrauterine devices of stainless steel ring, copper T220, and releasing levonorgestrel: morphometry of microvessels.

An electron microscopic morphometric study was performed for three kinds of IUDs, the stainless steel ring (SS), the copper T220 (TCu) and the WHO levonorgestrel IUD (LNG), for comparison of their effects on microvessels of the endometrium of women wearing the devices. The morphometry of plasmolemmal vesicles (PV) of the endothelial cell as well as the contracted and degenerated endothelial cell were undertaken. The results showed that all three IUDs led to significant changes in blood vessels, i.e., a higher percentage of contracted and degenerated cells than in the control group (p less than 0.01), and an increase in PV after insertion of the devices (p less than 0.01). The TCu caused more contracted endothelial cells than the SS (p less than 0.01) and the LNG (p less than 0.05). The levonorgestrel-releasing IUD resulted in a higher increase of PV than the SS and the TCu. For the differences between bleeding and non-bleeding groups, only the TCu showed a higher percentage of contracted cells (p less than 0.05).

Adult↗