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Biomedical subjects

P W Chen

Publications and source records attributed to P W Chen.

At least 19 recordsLinked to original sources
PubMed2006

The role of target elevation in prey selection by tiger beetles (Carabidae: Cicindela spp.).

The elevation of objects in the visual field has long been recognized as a potential distance cue, but it has been demonstrated to a reasonable extent in only four species: humans, frogs, fiddler crabs and backswimmers. Many tiger beetles hunt in flat, sandy areas, and their eyes show "flat-world" adaptations, such as an extended visual streak of higher acuity that corresponds to the horizon. They are therefore possible candidates for the use of elevation as a cue for distance. We tested this empirically and with simulation. In a behavioral prey selection paradigm, in which starved beetles were presented moving prey-targets having different size, speed and elevation, the beetles showed a strong preference for large targets when these were low in the visual field and a weaker preference for small targets when these were near the horizon. Striking of targets above the horizon was reduced compared to sub-horizontal targets, and lacked the size-elevation interaction. We simulated these empirical results with a model that converted elevation to distance, and used distance to estimate the absolute size of the targets. Simulated strike probability was then determined by the similarity between this absolute size and an independently confirmed preferred prey size. The results of the simulation model matched the empirical data as well as the best statistical model of the behavioral results. While some aspects of the model, and the beetles' behavior, differ from the strict geometry of the "elevation hypothesis", our results nevertheless indicate that tiger beetles use elevation to estimate distance to prey, and that it is therefore one of the determinants of prey selection.

Altitude↗
PubMed2001

A disposable poly(methylmethacrylate)-based microfluidic module for protein identification by nanoelectrospray ionization-tandem mass spectrometry.

The design, fabrication, and analytical use of a poly(methylmethacrylate) (PMMA)-based microfluidic module for nanoelectrospray ionization-tandem mass spectrometry (nano-ESI-MS/MS) were described. The microfluidic module can be mass-produced at low costs and used as a disposable device to generate nano-ESI-MS/MS signals for protein identification from low amounts of protein samples. Compared with commercially available nanospray capillary tips, the module gave comparable signal quality and also offered advantages in convenience and easiness of operation, permitting repeated usage, and disposability.

Animals↗
PubMed1999

Immune privilege is extended, then withdrawn, from allogeneic tumor cell grafts placed in the subretinal space.

PURPOSE: To determine whether the subretinal space can extend immune privilege to allogeneic tumor cell grafts that do not possess their own inherent immune privilege. METHODS: P815 tumor cells were injected into the anterior chamber (AC), the subretinal (SR) space, or subconjunctivally in eyes of BALB/c (allogeneic), SCID (immune incompetent), normal DBA/2 (syngeneic), or DBA/2 mice presensitized with P815 cells transfected with interleukin-12 and B7.1. Tumor growth was observed clinically and histologically for up to 50 days. BALB/c recipients were tested for suppression of DBA/2-specific delayed hypersensitivity and concomitant immunity. The SR space of tumor-containing eyes was assessed for its capacity to support ovalbumin (OVA)-specific anterior chamber associated immune deviation (ACAID). RESULTS: P815 cells injected into the SR space of presensitized and normal DBA/2 and SCID mice grew progressively, resulting eventually in recipient death. Tumor cells injected into the SR space of eyes of BALB/c mice grew progressively until day 14, followed by tumor regression resulting in phthisis bulbi (14/35) or tumor elimination (19/35) with preserved ocular anatomy by day 35. Despite elimination of tumors from the SR space, BALB/c recipients exhibited DBA/2-specific ACAID and concomitant immunity. In addition, OVA injected into the SR space of eyes from which tumor has been eliminated induced ACAID. CONCLUSIONS: Various parameters of immune privilege, originally described for the AC, are characteristic of immune privilege within the SR space. However, because P815 cells placed in the AC prove lethal for BALB/c recipients, but P815 cells placed in the SR space resolve without jeopardizing the host's life, immune privilege in the SR space can be distinguished from immune privilege in the AC, and this may have implications for grafts of retinal tissue placed within the SR space.

Animals↗
PubMed1998

Uveal melanomas contain antigenically specific and non-specific infiltrating lymphocytes.

PURPOSE: Tumor-infiltrating lymphocytes (TIL) were recovered from a series of human choroidal melanomas and expanded in cultures containing interleukin-2 (IL-2) to determine whether TIL contained cytotoxic cells that could be activated in vitro. METHODS: TIL were recovered from six ocular melanoma patients and expanded in vitro with IL-2. Cytotoxic activity was tested in a standard 4-hr 51Cr release assay. The HLA class I phenotype of patients was determined, using peripheral blood lymphocytes and the Amos modified-cytotoxicity test. HLA class I expression on tumor cells was determined by flow cytometry. RESULTS: TIL from four patients lysed autologous ocular melanoma cells. Two of these patients possessed TIL that displayed specific cytotoxic activity and failed to lyse tumor cells from other patients (HLA-mismatched, or -matched). TIL from the remaining two patients possessed non-specific cytotoxic cells that lysed ocular melanoma cells from a variety of other patients (HLA-mismatched). TIL from patients that failed to lyse autologous tumor cells possessed cytotoxic activity for ocular melanoma cells from other HLA-mismatched patients. CONCLUSIONS: Ocular melanomas accumulate lymphocytes with the potential to kill tumor cells. Our results imply that elimination of tumor cells may be possible by activation of cytotoxic cells present within progressively growing ocular tumors.

Cells, Cultured↗
PubMed1998

Genomic cloning of 18 kDa oleosin and detection of triacylglycerols and oleosin isoforms in maturing rice and postgerminative seedlings.

Oleosins are hydrophobic proteins localized abundantly in the oil bodies of plant seeds. Two distinct oleosin isoforms of molecular masses 18 and 16 kDa are present in rice oil bodies. These isoforms were found in similar ratio in rice embryos and aleurone layers. To survey potential DNA sequences involved in the activation of oleosin genes, a genomic clone of rice 18 kDa oleosin was sequenced, and its 5'-flanking region was compared with that of the known rice 16 kDa oleosin gene. Corresponding mRNAs of the two rice oleosin isoforms appeared seven days after pollination and vanished in mature seeds. Triacylglycerols and oleosins were accumulated concomitantly in maturing rice reeds in accord with the active assembly of oil bodies, and partly mobilized in postgerminative seedlings. Approximately 60% of the stored triacylglycerols in rice were not utilized: while the majority of oil bodies in embryos were mobilized in five days after imbibition, those in aleurone layers remained intact in postgerminative seedlings.

Base Sequence↗
PubMed1997

Expression of MAGE genes in ocular melanoma during progression from primary to metastatic disease.

Primary melanomas that form within the eye have a unique pattern of disease progression as compared with melanomas that form within the skin. A high percentage of patients (approximately 50%) develop metastatic tumors that occur predominately in the liver. An unusual characteristic of ocular melanomas is the prolonged disease-free interval that extends for many years between the development of primary and metastatic tumors. It is estimated that the shortest interval between dissemination of tumor cells from the eye and the appearance of clinically detectable metastases is 6 years. A recent report indicated that fresh uveal melanoma tissue and metastatic tumor biopsies failed to express melanoma antigen gene (MAGE)-1, MAGE-2, or MAGE-3. In the present study, we examined the expression of MAGE genes on fresh and cultured tumor cells obtained from an ocular melanoma patient during different stages of progressive disease. MAGE gene expression was determined by reverse transcription-polymerase chain reaction using MAGE-1, MAGE-2 and MAGE-3 specific primers. Our results demonstrate that primary ocular tumor tissue and cultured tumor cells both express significant levels of MAGE-1, 2, and 3 at the time of enucleation. A high percentage of tumor cells within the primary tumor appear to express MAGE as demonstrated by consistent MAGE expression in 16 tumor cell clones. Metastatic liver tumors that developed 3 years after enucleation and 18 years after the initial formation of the primary tumor also expressed high levels of MAGE-1, -2, and -3. MAGE was expressed on fresh tumor tissue from a single biopsy and cultured tumor cells obtained from three of four different metastatic tumor nodules. When the MAGE-negative metastatic tumor cells were treated with the demethylating agent 5-Aza-2-Deoxycytidine (5-Aza-dC), transcription of MAGE-1 was restored, indicating the MAGE genes were not deleted. Our results demonstrate that in some patients, MAGE genes are expressed on primary and metastatic ocular melanomas.

Antigens, Neoplasm↗
PubMed1997

Expression of MAGE genes in ocular melanoma cell lines.

The pathobiology of melanomas that develop within the eye is distinct from melanomas that develop within the subcutaneous tissues of the skin. This may be related to the unique structural and functional differences between normal melanocytes present within the uveal tract of the eye and the epidermal layers of the skin. The purpose of the present study was to determine whether normal pigmented cells within the eye (melanocytes and retinal pigment epithelial cells) and cultured cells derived from malignant ocular melanomas express the MAGE genes that encode tumor antigens that are recognized by specific CD8+ cytotoxic T lymphocytes. In the present series of experiments, we examined MAGE expression in cultured ocular melanoma cells obtained from a group of 17 ocular melanoma patients. Normal ocular melanocytes and retinal pigment epithelial cells were recovered and cultured from eyes enucleated for trauma. MAGE gene expression was determined using reverse transcription-polymerase chain reaction specific for either MAGE-1, -2, or -3. Our results demonstrate that MAGE-1, MAGE-2, and MAGE-3 genes are transcribed in primary ocular melanoma cell lines and are detected in cells recovered from 41, 53, and 53% of the patients examined, respectively. Normal choroidal melanocytes and retinal pigment epithelial cells did not express MAGE genes. We conclude that cultured ocular melanoma cell lines express MAGE-1, MAGE-2, and MAGE-3 genes.

Antigens, Neoplasm↗
PubMed1997

Gene transfer of the CD80 costimulatory molecule into ocular melanoma cells using a novel episomal vector.

PURPOSE: The CD80 (B7.1) molecule, which is a necessary costimulatory signal for T-cell activation and proliferation, is a powerful inducer of antitumor immunity. In this study, primary human ocular melanoma cells were transfected with a novel vector (B45-Neo episomal vector) containing the complementary DNA (cDNA) for human CD80 to determine if this vector system is useful for stimulating CD8+ T cells. METHODS: Ocular melanoma cells were transfected with the B45-Neo episomal vector containing the cDNA for human CD80 and were positively selected in medium containing geneticin. The transcription of plasmid cDNA, plasmid copy number, and cell surface expression were determined on transfected tumor cell lines, and cloned tumor cells were obtained by limiting dilution techniques. The stability of CD80 expressed on tumor cells was determined after prolonged culture without geneticin and on irradiated cells. Autologous lymphocytes were restimulated with CD80+ tumor cells in the presence of recombinant interleukin-2 to determine whether CD8+ T cells were stimulated. RESULTS: CD80 was expressed on tumor cells transfected with the B45-Neo vector containing the cDNA for CD80. The level of CD80 expressed on different transfected tumor cell lines was heterogeneous and dependent on the plasmid copy number. High CD80 expression was observed on cloned tumor cells that possessed more than 520 plasmids per cell; intermediate levels were observed on tumor cells with approximately 240 to 520 plasmids. CD80+ ocular melanoma cells maintained a stable CD80 expression even after prolonged culture without geneticin, and on irradiated tumor cells. CD80 expressed on tumor cells was biologically functional and stimulated autologous CD8+ cells. CONCLUSIONS: The B45-Neo episomal vector induces stable expression of the CD80 costimulatory molecule on ocular melanoma cells. Our results indicate that this vector is suitable for experiments designed to genetically engineer ocular melanoma cells to stimulate CD8+ T cells.

B7-1 Antigen↗
PubMed1996

Therapeutic antitumor response after immunization with a recombinant adenovirus encoding a model tumor-associated antigen.

Recombinant adenovirus (rAd), deleted of critical genes that enable viral replication and replaced with genes encoding heterologous proteins, has been shown to be a safe and effective vector in gene therapy studies. To evaluate a potential role for rAd as an immunogen, we used two different replication-defective type 2 rAds encoding the model Ag, beta-galactosidase (beta-gal). To determine whether rAd elicited the kind of immune responses therapeutic in an anti-tumor setting, the beta-gal-expressing adenocarcinoma, CT26.CL25, was used. Splenocytes from BALB/c mice immunized with 1 x 10(7) infectious units (iu) of rAd demonstrated anti-beta-gal activity after in vitro culture with the relevant L(d) beta-gal peptide. Adoptive transfer of these same splenocytes produced dramatic regression of established pulmonary metastases. However, when tumor-bearing mice were treated with 1 x 10(7) iu of rAd, no reduction in established disease was observed even when rAd was given with exogenous IL-2. To increase the viral dose delivered to each animal, we used an E1-E4-deleted rAd that could be grown to much higher titers. Significant reduction occurred with 10-fold more rAd (1 x10(8) iu) was administered. Exogenous IL-2 administration with 1 x 10(8) iu of rAd resulted in augmentation of this anti-tumor effect. These findings demonstrate that when using a nonreplicating virus, the viral dose is directly related to the immune response generated. These data constitute the first reported use of rAd in the treatment of an established experimental cancer and may have implication for the treatment of human cancer.

Adenocarcinoma↗
PubMed1995

IL-2 enhances the function of recombinant poxvirus-based vaccines in the treatment of established pulmonary metastases.

Neoplastic cells are generally poor immunogens. Transfection of the murine tumor CT-26 with beta-galactosidase (beta-gal), a protein from Escherichia coli, did not alter its growth rate in vivo, or its lethality, and did not elicit a measurable anti-beta-gal immune response. Immunization with beta-gal-expressing recombinant vaccinia viruses (rVV) elicited specific anti-beta-gal cytolytic T lymphocytes, but rVV-beta-gal was only marginally therapeutic when given to tumor-bearing mice. With the aim of expanding the immune response against beta-gal, used here as a model tumor Ag, we gave mice exogenous IL-2 starting 12 h after the poxvirus. The therapeutic effectiveness of the combination of poxvirus and IL-2 was far greater than either of these treatments alone. When the cDNA for IL-2 was inserted into the viral genome of the rVV construct to make a double recombinant (drVV), antitumor activity was further augmented. One mechanism of action may be the enhanced activation or expansion of cytotoxic T cells, because a marked increase in primary cytotoxic responses against vaccinia determinants was observed. Interestingly, other cytokines (mGM-CSF, mTNF-alpha, and mIFN-gamma) inserted into the rVV genome did not modify the efficacy of the rVV constructs. The increase in specific CTL responses against beta-gal by drVV expressing the tumor-associated Ags (TAA) and IL-2 was more pronounced in mice bearing the lacZ-transduced tumor than in those bearing the parental cell line, suggesting that the TAA presented by growing tumor cells can either pre-activate or otherwise amplify the immune response induced by the rVV. Unfortunately, in several long-term surviving mice, tumor recurred that no longer expressed beta-gal. These results indicate that treatment of disseminated tumors by using recombinant viruses expressing TAA can be enhanced by IL-2 provided exogenously, or encoded within the recombinant virus.

Amino Acid Sequence↗
PubMed1995

Active immunotherapy of cancer with a nonreplicating recombinant fowlpox virus encoding a model tumor-associated antigen.

Some tumor cells express Ags that are potentially recognizable by T lymphocytes and yet do not elicit significant immune responses. To explore new immunotherapeutic strategies aimed at enhancing the recognition of these tumor-associated Ags (TAA), we developed an experimental mouse model consisting of a lethal clone of the BALB/c tumor line CT26 designated CT26.WT, which was transduced with the lacZ gene encoding beta-galactosidase, to create CT26.CL25. The growth rate and lethality of CT26.CL25 and CT26.WT were virtually identical despite the expression by CT26.CL25 of the model tumor Ag in vivo. A recombinant fowlpox virus (rFPV), which is replication incompetent in mammalian cells, was constructed that expressed the model TAA, beta-galactosidase, under the influence of the 40-kDa vaccinia virus early/late promoter. This recombinant, FPV.bg40k, functioned effectively in vivo as an immunogen, eliciting CD8+ T cells that could effectively lyse CT26.CL25 in vitro. FPV.bg40k protected mice from both subcutaneous and intravenous tumor challenge by CT26.CL25, and most surprisingly, mice bearing established 3-day pulmonary metastasis were found to have significant, Ag-specific decreases in tumor burden and prolonged survival after treatment with the rFPV. These observations constitute the first reported use of rFPV in the prevention and treatment of an experimental cancer and suggest that changing the context in which the immune system encounters a TAA can significantly and therapeutically alter the host immune response against cancer.

Adenocarcinoma↗
PubMed1995

Anti-tumor activity of cytotoxic T lymphocytes elicited with recombinant and synthetic forms of a model tumor-associated antigen.

The recent cloning of tumor-associated antigens (TAAs) recognized by CD8+ T lymphocytes (TCD8+) has made it possible to use recombinant and synthetic forms of TAAs to generate TCD8+ with anti-tumor activity. To explore new therapeutic strategies in a mouse model, we retrovirally transduced the experimental murine tumor CT26(H-2d), with the lacZ gene encoding our model TAA, beta-galactosidase (beta-gal). The transduced cell line, CT26.CL25, grew as rapidly and as lethally as the parental cell line in normal, immunocompetent animals. In an attempt to elicit TCD8+ directed against our model TAA by using purely recombinant and synthetic forms of our model TAA, we synthesized a nine-amino-acid long immunodominant peptide of beta-gal (TPH-PARIGL), corresponding to amino acid residues 876-884, which was known to be presented by the Ld major histocompatibility complex (MHC) class I molecule, and a recombinant vaccinia virus encoding the full-length beta-gal protein (VJS6). Splenocytes obtained from naïve mice and co-cultured with beta-gal peptide could not be expanded in primary ex vivo cultures. However, mice immunized with VJS6, but not with a control recombinant vaccinia virus, yielded splenocytes that were capable of specifically lysing CT26.CL25 in vitro after co-culture with beta-gal peptide. Most significantly, adoptive transfer of these cells could effectively treat mice bearing 3-day-old established pulmonary metastases. These observations show that therapeutic TCD8+ directed against a model TAA could be generated by using purely recombinant and synthetic forms of this antigen. These findings point the way to a potentially useful immunotherapeutic strategy, which has been made possible by the recent cloning of immunogenic TAAs that are expressed by human malignancies.

Amino Acid Sequence↗
PubMed1994

Presentation of endogenous tumor antigens to CD4+ T lymphocytes by murine melanoma cells transfected with major histocompatibility complex class II genes.

In a previous study, we demonstrated that K1735 transfectants expressing either Kk or Ak antigens alone produced tumors in syngeneic mice, whereas transfectants that expressed both antigens were rejected. In this study, we investigated whether K1735 transfectants expressing Ak molecules can present endogenous tumor antigens to CD4+ T lymphocytes in the absence of normal accessory cells. Our results indicate that K1735 transfectants expressing Kk and Ak molecules presented antigen to both CD4+ and CD8+ T lymphocytes, whereas K1735 transfectants expressing only the Ak or the Kk antigen preferentially stimulated either CD4+ or CD8+ T cells. Analogous to endogenous antigens, K1735 transfectants expressing Ak molecules also presented exogenous hen egg lysozyme (HEL) to HEL-specific 3A9 hybridomas in the absence of normal accessory cells. These results demonstrate that K1735 murine melanoma cells expressing Ak molecules can function as antigen-presenting cells and that the generation of an effective antitumor immune response by K1735 melanoma cells expressing Kk and Ak antigens is due to their ability to present endogenous tumor antigens to both helper and cytotoxic T cells.

Animals↗