Intravenous Injection Of Cells Research Articles

Immunohistochemistry And Hormonal Investigation For Spermatogenesis Restoration In Degenerated Testes Of Rats After Bone Marrow Stem Cells Transplantation

Stem cell therapy is considered an important and innovative tool for applied research in andrology, especially in infertility; therefore, it was later adapted as potential therapeutic agents. This study aimed to evaluate rat mononuclear bone marrow cells' ability to recover testis cells in cyclophosphamide (CTX)-treated rats and to assess its effects on hormonal and histopathological changes. The bone marrow cells were harvested from femurs and tibias of rats and purified by a Histopaque gradient. Mononuclear bone marrow cells transplantation was performed by intravenous injection of cells in cyclophosphamide-treated animals. Three weeks after transplantation, blood samples were collected and analyzed for hormonal assay. In addition, the testes were collected for histological and histopathological determination. The results depicted that the serum levels of all tested hormones were significantly different among the three experimental groups G1 (control, healthy animals), G2 (CTX induced infertility and untreated) and G3 (CTX induced infertility and treated with stem cells). FSH and LH levels were significantly increased in G2 (CTX) compared to G1 and G3. Total and free testosterone levels were slightly higher in G3 compared to G2. Mononuclear bone marrow cell transplantation promoted cellular reorganization of the seminiferous epithelium. Also, spermatogenesis regeneration was improved. In conclusion, bone marrow stem cells can regenerate the damaged testicular elements and hence restore hormonal regulation in cyclophosphamide treated rat. Therefore, the treatment of male infertility and testosterone deficiency could be therapeutically treated by using stem cells.

Abstract 3126: Renca RCC syngeneic model to evaluate efficacy of novel antisense oligonucleotides targeting TGF-β isoforms

Abstract Transforming growth factor beta (TGF-β represents a family of cytokines, which function as the primary mediators for TGF-β signaling via TGF-β receptor type II (TβIIR) and both non-canonical and canonical downstream signaling pathways. TGF-β is associated with a wide range of biological processes in oncology, including tumor cell invasion, migration, angiogenesis, immunosuppression, as well as regulation of tumor stem cell properties. Hence, optimal preclinical evaluation of efficacy of TGF-β antagonists is challenging. Isarna Therapeutics has designed and developed selective and potent LNA-modified antisense oligonucleotides targeting the various TGF-β isoforms. In order to adequately evaluate selected preclinical development candidates, Oncodesign has developed customized experimental Renca tumor models in syngeneic and/or immunodeficient mice. The Renca cell line was established from a murine transplantable renal adenocarcinoma of spontaneous origin, and has been used under various experimental conditions: (1) subcutaneous tumor model by inoculating cells into the flanks of the animals; (2) the pulmonary metastatic tumor model by an intravenous injection of cells into the tail vein; and (3) the orthotopic tumor model by injecting cells into the renal subcapsule (and subsequent pulmonary metastasis). Outcome of this development program and preliminary results for selected TGF-β antisense oligonucleotides will be presented and discussed. Citation Format: Hanna Korhonen, Julie-Orlane Redon, Damien France, Guillaume Serin, Francis Bichat, Frank Jaschinski, Katja Wosikowski, Michel Janicot. Renca RCC syngeneic model to evaluate efficacy of novel antisense oligonucleotides targeting TGF-β isoforms. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3126. doi:10.1158/1538-7445.AM2014-3126

Science to Practice: Genetic Engineering Meets Cell Tracking—A Promising Approach for Cell-based Therapies?

Cell-based therapies are gaining increasing importance and have been evaluated in many settings, including cancer, Parkinson disease, myocardial repair, stroke, and diabetes. In this context, it is of major importance that the cells are implanted into an optimal tissue environment and, correspondingly, that they reach the diseased region. Intravenous cell injection often is the route of choice, particularly if the cells are expected to circulate in the blood for longer periods of time and are able to actively migrate to the diseased tissue. Unfortunately, often only a small percentage of intravenously injected cells reach the target area. Higher accumulation can be achieved if the cells are injected into an artery that feeds the diseased area. However, in this case, fast blood velocities and substantial sheer stress make it difficult for the cells to adhere. In the study by Gorelik and colleagues (1), genetic engineering was used to overcome this limitation. Glial precursor cells were transiently transfected with very late antigen-4 (VLA-4) binding to vascular cell adhesion molecule-1 (VCAM-1), which is upregulated in inflamed endothelial cells. After labeling these cells with superparamagnetic iron oxide (SPIO) containing rhodamine, significantly increased binding to inflamed brain endothelial cells was shown and their homogeneous distribution over the inflamed brain tissue was convincingly demonstrated with magnetic resonance (MR) imaging and histologic examination. The authors concluded that transient transfection of SPIO-labeled cells with VLA-4 in combination with their arterial injection and the use of MR imaging monitoring may be an elegant way to increase the efficacy of cell-based treatments of inflammatory brain diseases.

Abstract B18: Champions Tumorgraft™ models demonstrate spontaneous metastases.

Abstract One of the challenges in oncology drug development is the lack of in vivo models that demonstrate the spontaneous metastatic process. Standard subcutaneous implantation of cell-line based xenografts models seldom results in the generation of spontaneous metastatic lesions. In general, experimental metastatic models are utilized which requires the intravenous injection of cells or orthotopic implantation of tumors cells to replicate metastases. These methods fail to capture the full process of metastases and only capture seeding in the distant location. Champions Oncology utilizes its innovative Champions Tumorgraft™ platform where primary human tumors are implanted into immunocompromised mice in a manner that preserves the biological properties of the original human tumor. As part of its extensive characterization, Champions has recently observed an increasing number of Champions Tumorgraft™ models demonstrating spontaneous metastatic lesions following subcutaneous (s.c.) implantation. The tumor types demonstrating spontaneous metastasis include breast, melanoma, colorectal, esophageal adenocarcinoma, and head and neck squamous cell carcinoma. Two Champions Tumorgraft models highlighted in this report are CTG-0033, a breast cancer model, and CTG-0213, a melanoma model. These Champions Tumorgraft models demonstrate frequent metastases to the liver and spleen following s.c. implantation. For model CTG-0033, metastases were seen in greater than 75% of the animals following implantation. The metastatic behavior of this Tumorgraft model also parallels the clinical disease progression. For instance, CTG-0033 was derived from a breast cancer patient who, following the development of the Tumorgraft model, developed extensive metastases. Given that treatment failures clinically are often due to recurrence of the tumor at metastatic locations, there is significant need for in vivo models that replicate the spontaneous metastatic process. Overall, numerous Champions Tumorgraft models implanted s.c. exhibit spontaneous metastatic behavior and are therefore ideal models for oncology drug development focused on inhibition of metastases. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B18.

Human and Mouse Colon Cancer Utilizes CD95 Signaling for Local Growth and Metastatic Spread to Liver

Analysis of clinical colon cancer specimens show alterations in the CD95 (Fas Ag/Fas L) pathway as tumors progress from local to metastatic disease, suggesting that this pathway may play a role in invasive behavior of colon cancer. However, direct causality between these alterations and clinical disease progression has not been shown. Surgically resected metastatic colon cancer samples were evaluated for Fas Ag/L and apoptosis. Alterations in the Fas-signaling pathway found in human samples were recreated through a series of staged transfection experiments in the MC38 mouse colon cancer cell line and the effects on growth tested in vitro and in vivo. Expression of FLICE-like inhibitory protein confers apoptosis resistance, increasing the incidence of primary tumors through a survival advantage by avoiding apoptosis and inducing Fas-mediated proliferation. Coexpression of Fas L enables colon cancer cells to metastasize to the liver from local tumors as well as from intravenous injection of cells. MC38-FasL/FLICE-like inhibitory protein colon cancer cells induce apoptosis in hepatocytes via activation of type II Fas Ag signaling, thus creating a niche conducive to tumor growth and fueling their own growth via Fas proliferative signaling. Alterations in the Fas Ag pathway which inhibit apoptosis and increase Fas-mediated proliferation directly increase local colon cancer growth, and enhance metastasis to the liver. Delineating points in the pathway responsible for growth and metastasis will offer targets that may be exploited for therapy.

A flow cytometric analysis of the expression of adhesion molecules on human renal cell carcinoma cells with different metastatic potentials.

We investigated the relationship between the metastatic potential and the surface expression of adhesion molecules on human renal cell carcinoma (HRCC) cells. The metastatic potential was studied by the intravenous injection of cells from an HRCC cell line SN12C parent and its variants, SN12C-MM3, SN12C-clone2 and SN12C-clone8 into athymic Balb/c nude mice. The surface expression of adhesion molecules on these four HRCC cell lines was studied by a flow cytometric analysis. Of the four cell lines, SN12C-MM3 had the highest ability to produce pulmonary metastatic nodules. In contrast, the SN12C parent, SN12C-clone2 and SN12C-clone8 produced either few or no pulmonary metastatic nodules. A flow cytometric analysis revealed that SN12C-MM3 showed a strong expression of sialyl Lewis X (sialyl Le(x)) carbohydrate antigen and a slightly stronger expression of CD11a (LFA-1 alpha-chain) and CD54 (ICAM-1) compared with the other three cell lines. All cell lines expressed CD29 (beta 1-integrin) and CD44 to the same extent. Sialyl Le(x) is thought to be a ligand for the adhesion molecule called ELAM-1 (endothelial-leukocyte adhesion molecule-1, E-selectin) and mediates the interaction of leukocytes or tumor cells to endothelial cells, followed by the integrin-mediated adhesion. These data suggest that SN12C-MM3 cells may have more of a chance to adhere to endothelial cells in blood vessels and consequently shows a higher metastatic potential when injected intravenously in comparison to the other three lines.

Importance of orthotopic implantation for human tumors as model systems: relevance to metastasis and invasion.

Transplantation of human tumors into immunodeficient athymic nude mice has become an important experimental approach to study the biology and the treatment of human cancer. Most human tumor xenograft experiments have employed subcutaneous injection procedures, but the main limit of this technique is the lack of metastasis from the subcutaneous site. The possibility of producing experimental metastasis by intravenous injection of cells in the animals has been known for a long time, and it has been recently reported that tumorigenic properties and metastatic ability of human cancer can be altered by transplantation of the tumor into its organ or tissue of origin in the recipient animals (orthotopic transplantation). In this paper we review (1) the principal techniques of orthotopic injection of most solid tumors, (2) the most recent techniques to achieve experimental metastases, and (3) the methods for preparing tumor cell suspensions from human surgical specimens suitable for transplantation into animals. These animal models should be used for a more appropriate evaluation of new antitumor treatments including the ones targeted to inhibit metastatic spread.

Growth and metastasis of hypermotile, hyperinvasive cancer cells selectedin vitro by rapid locomotion under various conditions

Cancer cells selected from a cultured murine fibrosarcoma by rapid migration through micropore membranes moved considerably faster through such membranes and invaded biological tissues much more efficiently than did the unselected parent cells. The present data show that populations of cells selected by unstimulated migration or by haptotaxis to laminin moved not only faster, but also in larger numbers than the parent cells. However, the selected cells were far less efficient than the parent cells in forming spontaneous lung metastases in syngeneic mice, although all cell lines were 100 per cent tumorigenic. Analysis of paired data within each group showed no relationship between the primary tumor size at any observation time and the number of lung metastases finally formed. Therefore, although the parent cell line produced primary tumors growing slightly more rapidly than did the various lines of hypermotile cells, this was probably not the main cause of the difference in spontaneous metastasis formation between the groups. Lung colonization experiments performed by intravenous injection of cells could not explain the spontaneous metastasis results. In vitro, the cells selected by rapid haptotaxis to laminin grew considerably better than the other cells in 0.1 per cent fetal bovine serum, but there were no, or only minor, differences in higher serum concentrations. Combined, these results indicate that small subpopulations of cells selected by extreme efficiency in one step of the metastasis process may be so specialized that they perform poorly in other steps. Therefore, the results do not disprove the concept that tumor cell migration plays an important part in metastasis.

A demonstration of a strain related restriction effect in the formation of experimental metastases.

In view of the role of histocompatibility proteins in mediating many types of cell interaction it was decided to investigate their role in the formation of experimental metastatic deposits using the B16 mouse melanoma cell line. The expression of both major histocompatibility complex (MHC) Class I and Class II proteins was studied in vitro. Expression of both MHC Class I and Class II proteins was greater in the highly metastatic F10 cell line as compared with the poorly metastatic F1 line. Intravenous injection of cells into syngeneic and semi-allogeneic animals revealed a strain related restriction effect on tumour growth following intravenous injection. However, this was mediated by a locus other than H-2. No restriction of lung trapping of radiolabelled cells or local growth following intraperitoneal injection was found. It is suggested that non-H-2 Class I proteins may mediate some of the stages of metastatic tumour growth independent of the immune system.

Pattern of spread of bloodborne tumour.

The pattern of bloodborne tumour spread has been studied experimentally in syngeneic rats. A variety of tumour types has been injected intravenously, intraportally and also intra-arterially via the left ventricle. Tumour cell arrest as a factor in the localization of metastases in the lung following intravenous injection of cells, and in the liver following intraportal injection, is emphasized. Once tumour cells have entered the arterial circulation they disseminate to almost all organs in similar proportions to the distribution of cardiac output. The dissemination and arrest of cells is not found to correlate with the later distribution of metastases. For example, certain organs (especially the adrenals) which receive only low fractions of injected cells, are always preferred sites for bloodborne metastasis. A strikingly similar pattern of 'arterial metastasis' is also seen for all the tumours used despite their very different histological and biological natures.

Migration of T lymphocyte precursors into the thymus and factors affecting this process

Migration of FITC-labeled mouse bone marrow cells into the thymus was measured by flow cytometric analysis 3 hours after intravenous injection of cells into irradiated mice. The percentage of cells reaching the thymus diminished when the dose of injected cells increased. The dependence of the number or labeled cells in the thymus on the dose of injected cells was not linear. Pretreatment of cells with anti-SC-I serum, peanut lectin or H-2 incompatibility antigen abolished thymus migration, while treatment with anti-Thy-I serum, soybean lectin, trypsin or Thy-I-incompatibility antigen diminished cellular migration and treatment with neuraminidase enhanced it. It was concluded that the main type of migrating cells is SC-1+ precursors of T-lymphocytes. Penetration of these cells through the blood-thymus barrier is based on the recognition of their partly sialized surface glycoprotein receptors by membrane lectins of the blood-thymus barrier cells.

Disseminated (metastatic) tumors in nude mice produced by intravenous injection of cells of human and nonhuman neurogenic tumor lines.

Cells of two human glioma lines (UC-35AG, UC-58EG), a human medulloblastoma line (UC-51MB), two rat glioma lines C6, RG2), and a rat schwannoma line (Lew-MS) were injected i.v. into male nude mice (BALB/c nu/nu). Each animal was injected with 10(6) viable cells of a particular line; for each line there was a group of eight to nine animals. The mice developed disseminated (metastatic) tumors from all lines. The incidence of mice developing metastatic tumors was different for the various lines: 2/8 for the UC-34AG, 7/9 for the UC-58EG, 1/8 for the UC-51MB, 7/8 for the C6 line, 8/8 for the RG2 and 1/9 for the Lew-MS line. The shortest survival of the mice with tumors was observed with C6 (all died on days 10-14 post injection), and with RG2 (all died from day 32 to day 39 following injection). With the remaining lines, all mice survived until they were killed on day 40 after injection. Most frequently the tumors developed in the lung. Other organs, e.g., kidney or liver, were sometimes also involved, but usually to a considerably lesser degree than the lung. Metastases never developed in the CNS. It was observed that tumors of certain glioma lines tended to grow in the lung in characteristic patterns and involved or spared other organs.

A new class of infectious agents detectable by the production of chorioallantoic membrane lesions by human lymphoblastoid cell lines and their culture supernatants.

Intravenous injection of cells and their tissue culture supernatants (CS) from human lymphoblastoid cell lines (LCL) induced the formation of lesions on the chorioallantoic membrane (CAM) of the chicken embryo. Injection of cells and CS from non-LCL and normal human lymphocytes induced few or no lesions. Irradiated chick embryos were more sensitive to lesion formation than were nonirradiated embryos. The log10 CAM lesions induced in irradiated (500 rads) embryos were a linear function of the log10 cells (from LCL) in the inoculum; the slope was 1.0, within experimental error. The formation of CAM lesions did not depend on the presence of Epstein-Barr virus (EBV) since lesions were also induced by cells and extracts derived from EBV genome-free LCL. Lesion-inducing activity associated with CS was filterable through 0.22-mu filters, sedimented at 78,000 x g, and sensitive to inactivation by heat (56 degrees C for 30 min), UV irradiation, chloroform, sera from chickens immunized against CS, and certain human sera. Lesion-inducing activity associated with cells and extracts was resistant to 5,000 rads of gamma-radiation. B2/B2 embryos (the B locus is the major histocompatibility locus of chickens) were more sensitive to lesion formation than were B15/B21 and outbred embryos; this suggested a genetic influence on lesion formation. Our data suggest that the irradiated chicken embryo may be a highly sensitive and useful means for the detection of an unidentified or unknown agent or agents that may play an important role in human oncogenic lymphocyte transformation or might interact with transforming viruses.

Ditazole and platelets III. Effect of ditazole on tumor-cell induced thrombocytopenia and on bleeding time in mice.

Ditazole (4,5-diphenyl-2-bis-(2-hydroxyethyl)-aminoxazol) inhibits in vivo platelet aggregation induced in mice by intravenous injection of cells derived from an experimental tumor, the Lewis lung carcinoma. Such a protective effect of ditazole could not be observed when the number of circulating platelets dropped slowly following intramuscular implantation and spontaneous dissemination of the same cancer cells. These results support previous observations suggesting a different mechanism for the thrombocytopenia observed after intravenous and intramuscular injection of cancer cells. Tail transection bleeding time of normal mice is significantly prolonged by ditazole, a finding at variance with that reported in rats.

The behaviour of carcinoma of the large bowel in man following transplantation into immune deprived mice.

The growth of carcinoma of the human large bowel was studied in the first 2 passages in immune deprived mice. The tumours were obtained from large bowel resections on 3 people. There was a strong histological similarity between the patient's tumour and the tumour that grew subcutaneously in the mice 2-8 months after implantation. One dissimilarity observed was a higher mitotic index in some of the tumours growing in the immune deprived animals. In the second passage of the bowel tumours, cells were implanted into groups of 8-10 animals in the following sites: subcutaneous, intramuscular, intravenous, intrahepatic, intraperitoneal and intrathoracic. Growth of tumour was observed from all 3 tumours when they were implanted subcutaneously, intramuscularly, intraperitoneally and intrathoracically. Infiltration of muscle by tumour was a frequent finding. Lung metastases developed after intravenous injection of cells in 1 of the 3 tumours. In none of the 3 tumours did growth follow injection of cells directly into the substance of the liver. On no occasions were spontaneous metastases observed.

Hemopoietic colony-forming units in regenerating mouse liver: suppression by anticoagulants.

After hepatic injury induced by carbon tetrachloride, mitotically active hematopoietic cells of nonhepatic origin localize in the liver as judged by an increase in colony-forming nodules in the spleens of lethally irradiated recipient mice on intravenous injection of cells from these livers. The administration of warfarin suppresses the localization of colony-forming units in the regenerating liver by inhibiting the coagulation mechanism of the donor animals.