G-protein gamma 7 is down-regulated in cancers and associated with p 27kip1-induced growth arrest.
(17/7715)
We previously identified and cloned human G protein gamma 7 (G-gamma 7) gene, which is down-regulated in pancreatic cancer. We examined G-gamma 7 expression in other gastrointestinal tract cancers. In 24 of 30 patients with gastrointestinal tract cancer, Northern blot assay and immunohistochemical staining revealed significantly lower G-gamma 7 expression in tumors than in normal tissues from the same patients. Semiquantitative reverse transcription PCRs also showed lower G-gamma 7 expression in tumors than in corresponding normal tissues in 69 of 90 patients. To examine the biological role of G-gamma 7 in cancer, the G-gamma 7 cDNA was transfected into a human esophageal carcinoma cell line, KYSE150, that lacks G-gamma 7 expression. G-gamma 7 expression suppressed cell growth and tritiated-thymidine uptake when cells were confluent G-gamma 7 expression also suppressed tumorigenicity in BALB/c nude mice until 3 weeks after transplantation. G-gamma 7 expression increased the Go/G1 population and decreased the S phase population when cells were at high density. We confirmed that this change was associated with p27K1P1 expression. These findings suggest that human G-gamma 7 is associated with p27kip1-induced growth arrest and may be a therapeutic target in cancers. (+info)
Overproduction of hyaluronan by expression of the hyaluronan synthase Has2 enhances anchorage-independent growth and tumorigenicity.
(18/7715)
Hyaluronan (HA) has long been implicated in malignant transformation and tumor progression. However, due to the lack of molecular tools to directly manipulate production of HA, which does not require a core protein for its synthesis, our understanding of the role of HA in tumor cells has been largely circumstantial. In this study, we genetically manipulated the production of HA by transfection of a mammalian HA synthase Has2 into human HT1080 cells and examined the malignant phenotype of transfected cells. We found that increased production of HA promotes anchorage-independent growth and tumorigenicity of the cells. Has2-transfected cells formed greater numbers of colonies in semisolid medium. Tumors in nude mice derived from Has2-transfected cells grew more rapidly and were 2-4 times larger than those derived from control cells at termination of experiments. Histological and biochemical analyses of tumors revealed no significant differences in cell density and tissue structures between them, indicating that the larger size of the tumors was due to enhanced cell proliferation, not to increased accumulation of tumor stroma or increased angiogenesis. These results demonstrate that HA production by tumor cells per se promotes proliferation of these cells in tissues and provides direct evidence for the role of HA in tumorigenicity. (+info)
Xenotransplantation.
(19/7715)
As transplantation waiting lists lengthen because of the shortage of donor organs, the death rates of patients continue to rise. Xenotransplantation offers the potential to solve the problem of organ shortage br providing an unlimited supply of healthy donor organs. However, there are several barriers to xenotransplantation, including graft rejection, potential xenozoonosis, physiologic incompatibilities and ethical concerns. Experimental xenotransplantation studies continue in several areas, ranging from tissue to whole- organ grafting. Clinical studies continue in the area of tissue xenotransplantation. Trials with extracorporeal xenografts in an acute setting to support fulminant organ failure are likely to begin in the near future. The reintroduction of whole-organ xenotransplantation must be based on sound scientific analysis with broad societal input so as to offer the maximal benefit to transplant recipients and their families. (+info)
Chimerism and xenotransplantation. New concepts.
(20/7715)
In both transplant and infectious circumstances, the immune response is governed by migration and localization of the antigen. If the antigenic epitopes of transgenic xenografts are sufficiently altered to avoid evoking the destructive force of innate immunity, the mechanisms of engraftment should be the same as those that permit the chimerism-dependent immunologic confrontation and resolution that is the basis of allograft acceptance. In addition to "humanizing" the epitopes, one of the unanswered questions is whether the species restriction of complement described in 1994 by Valdivia and colleagues also necessitates the introduction of human complement regulatory genes in animal donors. Because the liver is the principal or sole source of most complement components, the complement quickly is transformed to that of the donor after hepatic transplantation. Thus, the need for complementary regulatory transgenes may vary according to the kind of xenograft used. Much evidence shows that physiologically important peptides produced by xenografts (e.g., insulin, clotting factors, and enzymes) are incorporated into the metabolic machinery of the recipient body. To the extent that this is not true, xenotransplantation could result in the production of diseases that are analogous to inborn errors of metabolism. In the climate of pessimism that followed the failures of baboon to human liver xenotransplantation in 1992-1993, it seemed inconceivable that the use of even more discordant donors, such as the pig, could ever be seriously entertained; however, this preceded insight into the xenogeneic and allogeneic barriers that has brought transplantation infectious immunity to common ground. With this new insight and the increasing ease of producing transgenic donors, the goal of clinical xenotransplantation may not be so distant. (+info)
Establishment and characterization of human neuroblastoma cell lines.
(21/7715)
Three new tissue culture cell lines, CHP-100, CHP-126, and CHP-134, have been established from explant cultures of human neuroblastoma. The cell lines have been characterized with respect to morphology, chromosomes constitution, growth, neural enzyme content, and their ability to grow in nude mice. The cells grow as dense masses comprised of fibroblast-or neuroblast-like cells with small processes. The cell lines differ in their neural enzyme acitivity. The chromosomal content of the 3 cell lines is near diploid, and all are capable of forming tumors in nude mice. The morphological findings indicate that the cells in culture resemble those found in the tumor, and the enzyme activities are consistent with those of nervous tissue. This the morphological, biochemical, and tumorigenic properties confirm that the 3 cell lines are neoplastic cells of neural origin. (+info)
Cytokine treatment or accessory cells are required to initiate engraftment of purified primitive human hematopoietic cells transplanted at limiting doses into NOD/SCID mice.
(22/7715)
Little is known about the cell types or mechanisms that underlie the engraftment process. Here, we have examined parameters affecting the engraftment of purified human Lin-CD34+CD38- normal and AML cells transplanted at limiting doses into NOD/SCID recipients. Mice transplanted with 500 to 1000 Lin-CD34+CD38- cord blood (CB) or AML cells required the co-transplantation of accessory cells (ACs) or short-term in vivo cytokine treatment for engraftment, whereas transplantation of higher doses (>5000 Lin-CD34+CD38- cells) did not show these requirements suggesting that ACs are effective for both normal and leukemic stem cell engraftment in this model. Mature Lin+CD34- and primitive Lin-CD34+CD38+ cells were capable of acting as ACs even though no repopulating cells are present. Cytokine treatment of NOD/SCID mice could partially replace the requirement for co-transplantation of AC. Furthermore, no difference was seen between the percentage of engrafted mice treated with cytokines for only the first 10 days after transplant compared to those receiving cytokines for the entire time of repopulation. Surprisingly, no engraftment was detected in mice when cytokine treatment was delayed until 10 days posttransplant. Together, these studies suggest that the engraftment process requires pluripotent stem cells plus accessory cells or cytokine treatment which act early after transplantation. The NOD/SCID xenotransplant system provides the means to further clarify the processes underlying human stem cell engraftment. (+info)
Human embryonic gastric xenografts in nude mice: a new model of Helicobacter pylori infection.
(23/7715)
In vitro or animal models have been used to investigate the pathogenesis of Helicobacter pylori infection. However, extrapolation to humans of results obtained with these heterologous models remains difficult. We have developed a new model for the study of H. pylori infection that uses human entire embryonic stomachs engrafted in nude mice. At 80 days after implantation, 22 of these xenografts, which exhibited a mature gastric epithelium, were inoculated with 10(7) to 10(8) CFU of either H. pylori LB1, a freshly isolated H. pylori strain (n = 12), or H. pylori ATCC 49503 (n = 10). After 12-week examination, H. pylori LB1 persistently colonized the antrum of all inoculated grafts, as assessed by culture (mucus and mucosa), immunohistochemistry (mucosa), and a rapid urease test (mucus). H. pylori ATCC 49503, either before or after in vivo passage, permitted only a transient 2-week colonization in one of the five inoculated grafts in both groups. Colonization was always associated with an increase of gastric juice pH. A mild neutrophil infiltration of the gastric mucosa was noted solely in infected grafts. Transmission electron microscopy showed adherence of H. pylori organisms to epithelial cell surface. In six animals, intracytoplasmic location of this bacterium was observed in the antrum or the fundus. These results allow us to propose this model as a new ex vivo model for the study of specific H. pylori-gastric cell interactions. (+info)
One-day ex vivo culture allows effective gene transfer into human nonobese diabetic/severe combined immune-deficient repopulating cells using high-titer vesicular stomatitis virus G protein pseudotyped retrovirus.
(24/7715)
Retrovirus-mediated gene transfer into long-lived human pluripotent hematopoietic stem cells (HSCs) is a widely sought but elusive goal. A major problem is the quiescent nature of most HSCs, with the perceived requirement for ex vivo prestimulation in cytokines to induce stem cell cycling and allow stable gene integration. However, ex vivo culture may impair stem cell function, and could explain the disappointing clinical results in many current gene transfer trials. To address this possibility, we examined the ex vivo survival of nonobese diabetic/severe combined immune-deficient (NOD/SCID) repopulating cells (SRCs) over 3 days. After 1 day of culture, the SRC number and proliferation declined twofold, and was further reduced by day 3; self-renewal was only detectable in noncultured cells. To determine if the period of ex vivo culture could be shortened, we used a vesicular stomatitis virus G protein (VSV-G) pseudotyped retrovirus vector that was concentrated to high titer. The results showed that gene transfer rates were similar without or with 48 hours prestimulation. Thus, the use of high-titer VSV-G pseudotyped retrovirus may minimize the loss of HSCs during culture, because efficient gene transfer can be obtained without the need for extended ex vivo culture. (+info)