Biomedical subjects

Y Sheng

Publications and source records attributed to Y Sheng.

At least 55 records · Page 3Linked to original sources

PubMed1997

Tolerance of human fetal retinal pigment epithelium xenografts in monkey retina.

BACKGROUND: RPE transplantation offers the possibility of treating certain forms of retinal degeneration. Understanding how to optimize the surgical technique for performing RPE transplantation, especially in primates, is therefore of considerable interest. METHODS: Fifteen patch RPE transplants were performed in six monkeys. The transplant sites were examined at follow-up by ophthalmoscopy, biomicroscopy, fluorescein angiography and histology. Foveal and peripheral retinal transplants were compared. RESULTS: Human fetal RPE xenografts can survive without rejection for at least 6 months after transplantation in monkey retina. Such grafts form a basal lamina and make intimate contacts with the outer segments of the host. Both rods and cones retain a normal appearance when in contact with unrejected transplants. Rejection occurred in only 30% (3/10) of the peripheral but in 60% (3/5) of the foveal transplants. CONCLUSIONS: Cultured human fetal RPE patch transplants can survive and maintain local photoreceptor integrity for relatively long periods of time in monkey subretinal space without immunosuppression. Rejection, when it occurs, is more frequent near the fovea.

Animals↗

PubMed1997

Neutral metoclopramide induces tumor cytotoxicity and sensitizes ionizing radiation of a human lung adenocarcinoma and virus induced sarcoma in mice.

Radiation induced cytotoxicity was potentiated by neutralized metoclopramide (nMCA; Neu-Sensamide, Oxigene Inc) when a human lung adenocarcinoma (H2981) transplanted into scid mice and an adeno-type 12 virus induced mouse sarcoma (A12B3) inoculated into CBA mice were exposed in vivo to low dose radiation at single doses of 1 and 2 Gy respectively. However, when the radiation dose was increased to 6, 10 or 18 Gy (single dose) and combined with a single dose nMCA (2 mg/kg), tumor cytotoxicity was not sensitized by the combination treatment. A fractionated dose of ionizing radiation (3 x 1 Gy) in combination with nMCA at a repeated dose of 3 x 10 mg/kg body weight (1 dose/day, i.m.) significantly increased cytotoxicity in H2981 compared with radiation given alone. nMCA alone also had a statistically significant dose dependent cytotoxic effect on H2981 growth when it was administered as repeated doses (8 doses) at 2 mg/kg or 10 mg/kg (1 dose every second day), and a similar result was achieved at 20 mg/kg but not at 2 and 10 mg/kg in the A12B3 tumor. In addition, the tumor volume at the start of treatment was important for the anti-tumor effect of nMCA (i.e. the larger initial tumor volume gave less effect on tumor growth). Taken together, our data propose that the mode of action of nMCA is different from radiation, and hence the two mechanisms are at least additive when in combination with lower radiation doses. The data further suggest that the cytotoxic mechanism is consistent with potentiating apoptosis because low and repeated doses of radiation (1-2 Gy), which are known to increase cytotoxicity by apoptosis, are sensitized by nMCA but not high doses and nMCA has more potent anti-tumor effects against H2981 tumors which have a higher constitutive apoptotic fraction of cells than A12B3.

Adenocarcinoma↗

PubMed1996

Transcriptional regulation of the N-acetylglucosaminyltransferase V gene in human bile duct carcinoma cells (HuCC-T1) is mediated by Ets-1.

N-Acetylglucosaminyltransferase V (GnT-V) catalyzes the transfer of N-acetylglucosamine from UDP-N-acetylglucosamine to alpha-6-D-mannoside to produce the beta1-6 linked branching of N-glycan oligosaccharides, which controls the polylactosamine content. The expression of N-acetylglucosaminyltransferase V, which contains 17 exons and spans 155 kilobase pairs, is expressed in a tissue- and cell type-specific manner and is regulated at the level of transcription by multiple promoters (Saito, H., Gu, J., Nishikawa, A., Ihara, Y., Fujii, J., Kohgo, Y., and Taniguchi, N. (1995) Eur. J. Biochem. 233, 18-26). To elucidate the mechanism by which the GnT-V gene is expressed in a cell- and tissue-specific manner, cell-restricted expression was analyzed using the 5'-upstream regions of the human GnT-V gene spanning base pairs -2760 to +23 in a human bile duct carcinoma cell line, HuCC-T1. We characterized two cis-acting elements that are potentially important in HuCC-T1 cell-specific expression. The two elements each contain an Ets-1 binding site, 5'-GGA-3'. Specific binding of Ets-1 to the respective elements was demonstrated by competition analysis as well as by antibody supershift experiments. Cotransfection of an Ets-1 expression plasmid along with a GnT-V promoter-luciferase reporter plasmid revealed the participation of Ets-1 in the regulation of the GnT-V gene transcription. These data indicated that the transcriptional regulation of the GnT-V gene was mediated by transcription factor Ets-1.

Bile Duct Neoplasms↗

PubMed1996

Site-directed mutagenesis of recombinant human beta 2-glycoprotein I identifies a cluster of lysine residues that are critical for phospholipid binding and anti-cardiolipin antibody activity.

beta2-Glycoprotein I (beta2GPI) is a phospholipid-binding serum protein with anticoagulant properties. It plays a vital role in the binding of anti-cardiolipin Abs purified from patients with autoimmune disease when assayed in a cardiolipin (CL) ELISA. Based on a three-dimensional model of beta2GPI, electrostatic calculations, and earlier peptide studies, a highly positively charged amino acid sequence, Lys282-Asn-Lys-Glu-Lys-Lys287, located in the fifth domain of beta2GPI, has been predicted to be the phospholipid binding site. We tested this hypothesis by site-directed mutagenesis of residues in the predicted phospholipid binding site and by assessing the mutants for phospholipid binding and anti-beta2GPI activity. A single amino acid change from Lys286 to Glu significantly decreased the binding of beta2GPI to CL. Double and triple mutants 2k (from Lys286, 287 to Glu286, 287), 2ka (from Lys284, 287 to Glu284, 287), and 3k (from Lys284, 286, 287 to Glu284, 286, 287) possessed no binding of Ab to beta2GPI in a CL ELISA, as well as no inhibitory activity on the binding of iodinated native beta2GPI to CL. These results indicate that the residues Lys284, Lys286, and Lys287 in the fifth domain of beta2GPI are critical for its binding to anionic phospholipids and its subsequent capture for binding of anti-beta2GPI Abs.

Amino Acid Sequence↗

PubMed1996

DNA damage and repair in tumour and non-tumour tissues of mice induced by nicotinamide.

In vivo DNA damage and repair was induced by nicotinamide (NAM) in adenotype 12 virus-induced mouse sarcoma A12B3 and sarcoma F inoculated into CBA mice. DNA damage, NAM and NAD concentrations were measured after in vivo exposure to NAM, in tumours and spleens by alkaline elution and by HPLC analysis. Our results indicate that NAM between 100-1000 mg kg-1 causes a high level of in vivo DNA strand breaks in tumours and normal tissues in mice bearing the immunogenic sarcoma A12B3 but not in the non-immunogenic sarcoma F. The repair process was also delayed by the NAM treatment probably owing to inhibition of the DNA repair enzyme, poly(ADP-ribose)polymerase, as evidenced by accumulation of NAM and NAD. These data are consistent with NAM having a mechanism of action as a radiosensitiser at least in part by DNA repair inhibition. In addition, it should also be considered that high doses of NAM might cause considerable complications to normal tissue in tumour-bearing individuals.

Animals↗

PubMed1996

Hypochlorous acid/N-chloramines are naturally produced DNA repair inhibitors.

Human mononuclear leukocytes (HML) respond to oxidative DNA damage by activation of ADP ribosylation and initiation of DNA repair synthesis (i.e. unscheduled DNA synthesis, UDS), whereas neutrophils do not. When neutrophils are added to HML cultures in ratios up to 4:1 ADP ribosylation becomes inhibited to approximately 50-60%. The ability of neutrophils to inhibit HML ADP ribosylation was shown to be dependent on H2O2, chloride ions and myeloperoxidase, which in turn are factors known to govern HOCl and N-chloramine production by phagocytic cells. HOCl and a model N-chloramine, chloramine T, were shown to give a dose-dependent inhibition of DNA repair using four independent estimates, namely ADP ribosylation, UDS and the repair of DNA strand breaks estimated by nucleoid sedimentation and alkaline elution profiles. All the DNA repair measurements used on HML were inhibited approximately 70-80% by 100 microM doses of HOCl or chloramine T, which was considered a biologically relevant dose because: (i) viable neutrophils equal in concentration to those found in blood could easily produce 100 microM levels in short-term culture; (ii) 100 microM doses of these agents were not acutely cytotoxic judged by trypan blue stained cells after 30-60 min exposure and under the conditions used for assay, but yet they abolished 86-95% of the growth response of HML to phytohemagglutinin.

Adenosine Diphosphate Ribose↗

PubMed1996

[Sixty years of study on history of chinese medical education].

This paper deals with the general study on ancient and modern Chinese medical education, namely the history of institution, history of medical thinking, regional medical education, minority medical education. The summary showed that there are certain achievements in this field of research. However, generally speaking, the basis is rather weak and the progress is sluggish. Many fields, including summary and systematization on heritage, developmental law of Chinese medical education, need to be further explored. The study on the contemporary part of the history of Chinese medical education should also be strengthened. Some suggestions are given in the summary of this paper.

China↗

PubMed1995

Transformation of Escherichia coli with large DNA molecules by electroporation.

We have examined bacterial electroporation with a specific interest in the transformation of large DNA, i.e. molecules > 100 kb. We have used DNA from bacterial artificial chromosomes (BACs) ranging from 7 to 240 kb, as well as BAC ligation mixes containing a range o different sized molecules. The efficiency of electroporation with large DNA is strongly dependent on the strain of Escherichia coli used; strains which offer comparable efficiencies for 7 kb molecules differ in their uptake of 240 kb DNA by as much as 30-fold. Even with a host strain that transforms relatively well with large DNA, transformation efficiency drops dramatically with increasing size of the DNA. Molecules of 240 kb transform approximately 30-fold less well, on a molar basis, than molecules of 80 kb. Maximum transformation of large DNA occurs with different voltage gradients and with different time constants than are optimal for smaller DNA. This provides the opportunity to increase the yield of transformants which have taken up large DNA relative to the number incorporating smaller molecules. We have demonstrated that conditions may be selected which increase the average size of BAC clones generated by electroporation and compare the overall efficiency of each of the conditions tested.

Chromosomes, Bacterial↗

PubMed1995

Expression of membrane-bound and secreted forms of equine herpesvirus 1 glycoprotein D by recombinant baculovirus.

Analyses of the synthesis and processing of recombinant full-length glycoprotein D of equine herpesvirus type 1 (EHV-1; gD392) or recombinant truncated gD (gD352) expressed in baculovirus-infected Sf9 cells revealed the following: (1) gD polypeptides encoded by both recombinant baculoviruses react with gD-specific antibodies including peptide-specific antiserum that neutralizes EHV-1 in a plaque reduction assay, (2) both the full-length recombinant gD392 and the truncated gD352 are expressed predominantly as gD species that contain high mannose-type oligosaccharides (55 kDa and 52 kDa, respectively), (3) both the full-length recombinant gD392 and the truncated gD352 are also expressed in lesser amounts as gD species that contain complex-type oligosaccharides (58 kDa and 55 kDa, respectively) as well as the unglycosylated forms of gD (43 kDa and 37 kDa, respectively), (4) flow cytometric analyses of cells expressing gD392 revealed that gD first appears on the cell surface at 24 h post infection; by 60 h, 95% of the cells express high levels of cell surface gD, (5) cells expressing gD352, in contrast to cells expressing gD392, secrete gD into the extracellular medium. This initial demonstration that immunoreactive EHV-1 glycoprotein D can be produced as a secreted polypeptide in the baculovirus system should provide reagents to assess the potential use of gD as a subunit vaccine in an animal model.

Animals↗

PubMed1995

Progress in identifying clinical relevance of inhibition, stimulation and measurements of poly ADP-ribosylation.

Our laboratory, in collaboration with Oxigene Inc, has been involved in identifying commercially feasible clinical applications of measurement or modulation of ADP-ribosylation as a core technology. For this purpose a pivotal regulatory role for ADP-ribosylation in the repair of DNA lesions leading to cytotoxic as well as mutagenic events has been hypothesized. A new class of DNA repair inhibitors, the N-substituted benzamides, has been identified which can react with radiation to produce reactive intermediates that oxidize thiol amino acids. Their proposed mechanisms of action are two-fold: ie they can interact with radiation: i) to directly enhance DNA damage; and ii) to react with thiols in the zinc finger DNA binding domain of poly ADP-ribosyl transferase to inhibit DNA repair and thereby increase DNA damage. Sensamide, a clinically relevant formulation of metoclopramide which is an N-substituted benzamide, has indicated enhancement of tumor response and survival in patients with inoperable squamous cell carcinoma of the lung when it was administered as a radiosensitizer in a phase I/II trial and compared to historical controls. A mechanism of endogenous regulation of human mononuclear leucocyte ADP-ribosylation has been identified to be HOCl/N-chloramine production via the oxidative burst of phagocytes. HOCl/N-chloramines are potent oxidants of thiol-containing proteins. Quantitative estimation of N-chloramine sensitive plasma thiols has been identified as an effective surrogate measure of leucocyte poly ADPRT.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

In vivo tumor measurement of DNA damage, DNA repair and NAD pools as indicators of radiosensitization by metoclopramide.

Metoclopramide (MCA), a N-substituted benzamide, causes DNA strand breaks and inhibits DNA repair in vitro and sensitizes radiation and chemotherapeutic drugs in human squamous cell carcinomas when xenographed into nude mice or in a rat glioma model. Here we report on the evaluation of the mechanism behind the radiosensitizing effects of MCA. DNA damage was measured in vivo in a CBA-mouse tumor line (A12B3, sarcoma tumor) by using both alkaline elution and nucleoid sedimentation analysis of cell suspensions prepared from either resected tumor, spleen tissues or whole blood samples. The amount of DNA damage caused by radiation alone, measured 30 min after the irradiation was started, was dose dependent up to 18 Gy in all tissues. The radiation-induced DNA damage in tumor tissue was elevated compared to radiation alone in the presence of MCA, but the level was not higher at 18 Gy compared to 6 Gy in the presence of MCA, and it was still not fully repaired 12 h after irradiation. HPLC analysis of the NAD pools in tumor tissue after DNA damage induction showed a delay in the recovery of the NAD pools (presumably due to the presence of still unrepaired DNA) after exposure to MCA (2 mg/kg) + radiation (6 Gy) compared to tumors exposed to radiation (6 Gy) only, which were fully restored after 48 h. These data confirm earlier published in vitro data on MCA as an inducer of DNA damage and an effector of DNA repair. In addition, the in vivo measurement of radiation-induced DNA damage and DNA repair using the nucleoid sedimentation and alkaline elution assays together with NAD pool determinations may prove to be effective intermediate endpoints in the evaluation of drugs as potential radiosensitizers.

Animals↗

PubMed1995

Acidic and neutralized metoclopramide formulations sensitize ionizing radiation induced cytotoxicity in a human lung adenocarcinoma xenografted to scid mice.

A neutralized formulation (Neu-Sensamide) of metoclopramide (MCA) has been shown to possess reduced sedative side effects compared with the conventional acidic formulations (Primperan). The acidic formulation of MCA has also been shown to sensitize the effect of ionizing radiation (6-8 Gy) using human squamous cell carcinomas from the head and neck xenografted to nude mice. In the present study, 2 mg MCA/kg body weight 1-3 h before treatment with 1 Gy radiation (single dose) was evaluated in scid mice xenografted with a human lung adenocarcinoma. MCA given alone in acidic or neutralized formulations did not show any effect on tumor growth retardation. However, when combined with radiation, both acidic and neutralized formulations of MCA sensitized the cytotoxic effect of radiation directed against the tumors by increasing tumor doubling time, tumor quadrupling time and specific growth delay, and by decreasing area under growth curve measurements. In addition, there was no statistically significant difference between the two formulations of MCA in the efficacy of sensitizing the cytotoxicity of a single low dose (1 Gy) of radiation.

Adenocarcinoma↗

PubMed1995

Release of platelet activating factor (PAF) and eicosanoids in UVC-irradiated corneal stromal cells.

Ultraviolet (UV) irradiation provokes acute inflammation of the eye, and can be used to model processes that occur in response to damage to the anterior segment. This study characterized ultraviolet-C (UVC, 254 nm) irradiation-induced PAF synthesis, and arachidonic acid (20:4) and eicosanoid release in rabbit corneal stromal cells maintained in vitro. PAF was measured by radioimmunoassay (RIA) after exposing cultured corneal stromal cells to UVC irradiation (20 min, 2, 5, 10 mW/cm2). 14C-20:4-labeled stromal cells were also stimulated with UVC and radiolabeled phospholipids, neutral lipids and eicosanoids were measured. Synthesis of cell-associated and secreted PAF from corneal stromal cells was increased by UV irradiation. UV irradiation (254 nm, 5mW/cm2) enhanced 20:4 release from triacylglycerols, phosphatidylinositol, phosphatidylserine and phosphatidylethanolamine, and increased levels of 20:4-diacylglycerol and unesterified 20:4. The released 20:4 entered both the cyclooxygenase and lipoxygenase pathways after UVC irradiation. The PAF antagonist, BN52021 (10 microM) reduced UVC irradiation-induced stimulation of prostaglandin production, but failed to inhibit UVC-induced 20:4 release and synthesis of lipoxygenase products. Furthermore, exogenous PAF (1 microM) stimulated prostaglandin production, but did not increase the synthesis of lipoxygenase products from radiolabeled 20:4. The effects of PAF on prostaglandin synthesis were inhibited by BN52021. These findings indicate that responses to injury in cultured corneal stromal cells include PAF synthesis, release of 20:4 from glycerolipids, accumulation of diacylglycerol and synthesis of eicosanoids. The data further suggest that during UVC irradiation in vitro, PAF is not a primary or initial mediator of 20:4 release and synthesis of lipoxygenase products, but may mediate UVC-induced prostaglandin synthesis.

Animals↗

PubMed1995

[Dynamics of IL-2 and IFN-gamma levels induced by sea or Con A in spleen cells of Schistosoma japonicum-infected mice].

The capacity of mitogen Con A and SEA-stimulated spleen cells to produce cytokines IFN-gamma and IL-2 was studied in S. japonicum-infected mice every two weeks from 0 to 14 wk after infection. The results showed that the levels of these two cytokines began to rise at the 4th wk after infection and reached a peak level at 6-8 wk, then declined to the levels similar to those pre-infection at 12-14 wk after infection. The IFN-gamma level reached the peak earlier than the IL-2 level. The dynamics of cytokine level in both mitogen and antigen-stimulated group was similar. The results suggest that IL-2 and IFN-gamma might be the essential cytokines involved in egg granuloma formation in schistosomiasis japonica.

Animals↗

PubMed1995

Patch transplants of human fetal retinal pigment epithelium in rabbit and monkey retina.

PURPOSE: To transplant human fetal retinal pigment epithelium (RPE) into the subretinal space of rabbits and monkeys as an organized monolayer without artificial support. METHODS: The method involves dissecting small patches of cultured RPE monolayers in sheets (1 to 5 mm2), sucking them into a glass pipette and injecting them into the subretinal space after producing a bleb detachment of the neural retina. RESULTS: These patches unfold and survive as a quasi-monolayer under the reattached neural retina intimately associated with the host photoreceptors and phagocytizing host outer segment material. Graft rejection is observed in most rabbits at 1 month but not in monkeys at 3 months after transplantation. CONCLUSIONS: Monolayer patches of cultured human fetal RPE can be transplanted to the subretinal space, where they survive in contiguity with healthy host outer segments. In primates, but not in rabbits, host-graft rejection does not occur for at least 2 to 3 months.

Animals↗

PubMed1994

Transplantation of fetal retinal pigment epithelium in age-related macular degeneration with subfoveal neovascularization.

BACKGROUND: Age-related macular degeneration (ARMD) is caused by abnormal retinal pigment epithelium (RPE) and may be complicated by choroidal neovascularization. The object of treatment would be to replace the diseased RPE with normal human RPE. METHOD: Five patients with ARMD (preoperative visual acuity 0.08-0.2) underwent removal of subretinal fibrovascular membranes using pars plana vitrectomy techniques. Human fetal RPE (15-17 weeks gestational age) was cultured and transplanted as a monolayer patch into the subretinal space. Transplants were followed by funduscopy and fluorescein angiography. Macular function was assessed using scanning laser ophthalmoscopic (SLO) microperimetry. RESULTS: Three RPE transplants were placed in the fovea; two were placed parafoveally. All transplants have survived for 3 months. They have grown and increased in size covering part of the epithelial defect caused by removal of the fibrovascular membrane. SLO microperimetry indicated that visual function was present in four of the transplants at 1 month but in only two at 3 months after surgery. Function over the transplants, especially those in the fovea, was compromised by cystoidlike macular edema. CONCLUSIONS: Human fetal RPE transplants survive well in the macula for as long as 3 months. They are capable of growing to cover epithelial defects caused by removal of subretinal neovascular membranes. The causes for development of macular edema in transplants directly in the fovea warrant further evaluation.

Aged↗

PubMed1994

Patch culturing and transfer of human fetal retinal epithelium.

Human retinal pigmented epithelium (RPE) can be cultured by removing small patches of this layer from the choroid of the fetal eye. Such RPE patches give rise to healthy, epithelioid monolayers in vitro within 1-2 weeks without contamination from retinal or choroidal cells. The viability and proliferative capacity of these cultures is independent of the initial polarity of the patch. These RPE monolayers develop apical/basal polarity and a basal lamina and rest on a field of collagen fibers; they are capable of phagocytizing outer segments. A patch can be lifted off a confluent monolayer and transferred to another culture dish without risking the viability of either the old or the new culture. This provides a means of transplanting an organized, polarized patch of human RPE from one place to another.

Adult↗

PubMed1994

[Pathological studies on alveolar hydatid in gerbils treated with albendazole and praziquantel].

Pathological changes in alveolar hydatids from experimentally infected gerbils were studied after administration of albendazole or praziquantel. The animals were divided into several groups including the control one, and drugs of different dosages were given separately to different groups with diverse durations of infection. According to the microscopical observation, the hydatids could be categorized into 3 grades: grade I, degenerated cysts; grade II, stable cysts (showing no proliferation); grade III, proliferated cysts. Larger proportion of grade I cysts were found in the group treated with albendazole at high dosage (33.3%) and the two groups treated with praziquantel (25.0% and 16.3%). The proportion of grade I cysts in gerbils with shorter duration (45 d) of infection was larger than those with longer duration (60 d). Lymphocyte infiltration to various extent was present surrounding the alveolar hydatid cysts in all the treated groups. More marked infiltration of lymphocytes was observed in albendazole treated gerbils (106.9 +/- 34.3-300.1 +/- 59.5) (P < 0.001). However, the combined medication of albendazole and praziquantel was less efficacious.

Albendazole↗