Search PubMed⌕ Search

Biomedical subjects

L A Fernandez

Publications and source records attributed to L A Fernandez.

At least 37 records · Page 2Linked to original sources
PubMed1995

Growth of B cell colonies independent of T cells: a new perspective.

Growth of human peripheral blood B cells in a B cell colony assay system is a useful technique to study the function of B cell biology. In the initial reports, T and B cells were admixed in the culture system, prior to which the T cells were treated with mitomycin or irradiation to prevent their proliferation. There were reports that optimal growth of B cell colonies required T cells to be in contact with the B cells. However, we were able to grow B cell colonies physically separated from T cells which were placed on a filter. We speculated then that T cells contacted B cells via pseudopods through the pores of the filter. We now report the growth of B cell colonies independent of T cells and conclude that B cell colony growth depends upon a critical number of B cells plated rather than on T cell help.

B-Lymphocytes↗
PubMed1994

Tumor localization and therapeutic potential of an antitumor-anti-CD3 heteroconjugate antibody in human renal cell carcinoma xenograft models.

A heteroconjugate (HC) antibody, constructed with the monoclonal antibody (MoAB) Dal K29 to human renal cell carcinoma (RCC) and an anti-CD3 MoAb, could induce a very high level of lysis of human RCC cells when incubated with human peripheral blood lymphocytes (PBL) in vitro (Kerr et al., 1990, J. Immunol., 144, 4060-4067). We now report that this HC antibody selectively localizes in RCC xenografts in nude mice and could inhibit RCC in an ascites tumor xenograft model when administered intraperitoneally together with PBL.

Animals↗
PubMed1994

Radioimmunotherapy of human B-cell chronic lymphocytic leukemia in nude mice.

After i.v. or i.p. inoculation of 5 x 10(6) D10-1 cells, a subclone of an Epstein-Barr virus transformed human B-cell chronic lymphocytic leukemia (CLL) line, 100% of nude mice developed solid or ascites tumors and died within 17-60 days of tumor inoculation. There was significant tumor inhibition, including tumor cure, when these tumor-inoculated mice were treated with either unmodified or 131I (300 microCi)-linked Dal B02 (50 micrograms/mouse), a monoclonal antibody directed against surface-associated antigens on human CLL B-cells and several histological types of B-lymphoma cells. There was no significant difference between the antitumor activity of unmodified Dal B02 and 131I-linked Dal B02 when the treatment was given 3 days after i.p. or i.v. inoculation of 5 x 10(6) D10-1 cells. However, when the mice were treated 3 days after i.p. inoculation of 15 x 10(6) D10-1 cells, or 7 days after the i.v. inoculation of 5 x 10(6) D10-1 cells, 131I-linked Dal B02 was a more potent tumor inhibitor than was unmodified Dal B02 (P < 0.05 and P < 0.01, respectively). Two injections of 131I (500 microCi) linked to 100 micrograms of a Dal B02 F(ab')2 fragment preparation also prolonged the survival of i.p. or i.v. tumor-inoculated mice (P < 0.05 and P = 0.05, respectively). In nude mice with established s.c. xenografts of D10-1 cells, two injections of 131I (300 microCi) linked to 50 micrograms of Dal B02 led to complete tumor cure in 3 of 4, mice, but two injections of 50 micrograms of unmodified Dal B02 had no effect on the s.c. xenografts. Two injections of 131I (500 microCI) linked to 100 micrograms of Dal B02 F(ab')2 fragment caused significant tumor inhibition but no tumor cure. 131I (300 microCi) linked to 50 micrograms of a nonspecific IgG1 only led to minor tumor inhibition. A mixture of unmodified Dal B02 and 131I-linked nonspecific IgG1 was not a more potent tumor inhibitor than the 131I-linked nonspecific IgG1 preparation by itself. These results suggest that Dal B02 may be an effective carrier for the radioimmunotherapy of human B-cell CLL and other appropriate B-cell lymphomas, especially in the progressive phase of B-cell CLL, which is usually not amenable to currently available therapeutic modalities.

Animals↗
PubMed1994

Inhibition of angiogenesis by the matrix metalloprotease inhibitor N-[2R-2-(hydroxamidocarbonymethyl)-4-methylpentanoyl)]-L-tryptophan methylamide.

The inhibitor N-[2R-2-(hydroxamidocarbonymethyl)-4-methylpentanoyl)]-L- tryptophan methylamide specifically blocks several matrix metalloproteases, enzymes which are thought to be involved in angiogenesis. An extract of Walker 256 carcinoma in Hydron pellets implanted in the corneas of Sprague-Dawley rats was used to stimulate angiogenesis from the vessels of the limbus. Angiogenesis was graded visually as the distance penetrated into the cornea and the number of vessels generated. The vessel area was also measured by image analysis using Image 1 software. Continuous i.v. administration of N-[2-(hydroxamidocarbonymethyl)-4-methylpentanoyl)]- L-tryptophan methylamide at 32 mg/kg/day (n = 17) via syringe pump reduced vessel number [25.06 +/- 5.9 (SEM) compared to 65.33 +/- 9.0] and vessel area (26.14 +/- 3.2 mm2 compared with 40.96 +/- 4.6 mm2), but not distance penetrated, compared to vehicle-treated control eyes after 6 days. These results confirm the suspected role for matrix metalloproteases in angiogenesis and suggest that inhibitors of these enzymes may be angiostatic agents.

Animals↗
PubMed1994

Inhibition of human B-cell chronic lymphocytic leukemia by a monoclonal antibody in xenograft models.

To establish xenograft models of human B-cell chronic lymphocytic leukemia (CLL), we inoculated 5 x 10(6) D10-1 cells, a subline of Epstein-Barr virus (EBV)-transformed human B-cell CLL with a marker chromosomal anomaly, into SCID or irradiated nude mice by the intravenous (i.v.) or intraperitoneal (i.p.) route. All i.p. tumor-inoculated mice developed rapidly progressive, lethal ascites tumor, and 100% of i.v. tumor-inoculated mice developed disseminated CLL. All mice died of tumor within 8 weeks of tumor inoculation. Tumor-inoculated SCID mice died earlier with wider tumor dissemination than the tumor-inoculated nude mice. All the tumor-inoculated mice had histologically confirmed metastases in lymph nodes, and most of them also had metastases in one or more internal organs. Cytogenetic analysis confirmed the origin of these tumors from the xenografted D10-1 cells. The D10-1 cells harvested from the xenografts did not differ from the parent D10-1 cells as regards (i) reactivity with 2 monoclonal antibodies (MAbs) directed against CLL-associated cell-surface antigens; (ii) rate of proliferation in vitro; and (iii) sensitivity to the 2 chemotherapeutic agents, methotrexate and adriamycin. Administration of 50 micrograms/mouse of Dal B02, an IgG1 (kappa) MAb directed against surface-associated antigens of human B-cell CLL, significantly prolonged the survival of D10-1-inoculated nude and SCID mice. The MAb was more effective in D10-1-inoculated nude mice than in SCID mice. In all the D10-1 xenograft models, the effectiveness of Dal B02 decreased with higher tumor load but increased with the amount of MAb injected. Dal B02 F(ab)'2 fragment failed to demonstrate any anti-tumor activity in D10-1-inoculated nude mice. In vitro assays revealed that Dal B02 had no direct inhibitory effect on D10-1 cells, but could be cytotoxic towards D10-1 cells in the presence of splenic cells or peritoneal macrophages from nude and SCID mice, or together with rabbit complement.

Animals↗
PubMed1994

Pharmacokinetics, biodistribution and tumor localization of two anti-human B-cell chronic lymphocytic leukemia monoclonal antibodies and their F(ab)'2 fragments in a xenograft model.

We investigated the pharmacokinetics, biodistribution and tumor localization of intravenously injected Dal B01 and Dal B02, two monoclonal antibodies (MoAbs) directed against tumor associated antigens on human chronic lymphocytic leukemia (CLL) B cells, and their F(ab)'2 fragments in nude mice bearing xenografts of the human B cell CLL line D10-1. More of the percentages of the injected dose (% ID) of these two MoAbs and their F(ab)'2 fragments specifically localized in the tumor xenografts than in normal tissues. Compared to intact MoAbs, their F(ab)'2 fragments had lower % ID in tumors and were cleared from circulation faster. Well-defined tumor images were obtained at 24 and 48 h after administration of [131I]Dal B02 F(ab)'2 fragment and at 96-192 h after administration of [131I]Dal B02. A comparison between intravenous and intraperitoneal routes of administration of [131I]Dal B02 did not reveal any difference in the localization of % ID in tumor or normal tissues.

Animals↗
PubMed1994

Angiotensin AT2 receptor stimulation increases survival in gerbils with abrupt unilateral carotid ligation.

Previous studies showed that angiotensin II (AII) infusion increased survival in gerbils subjected to abrupt unilateral carotid ligation. Recently, stimulation of the AII AT2 receptor, reportedly effectively extended the blood pressure (BP) range of cerebral blood flow (CBF) autoregulation. We evaluated the survival of gerbils treated with PD-123319, a ligand of AT2 receptors, to test the hypothesis that restoration of BF to ischemic cerebral tissue produced by AII is mediated through AT2 receptors. Abrupt unilateral carotid ligation was performed on 300 gerbils. In five experimental groups, animals received no drug pretreatment: (a) saline; (b)-(d) PD-123319 1.0, 3.0, and 10 mg/kg; and (e) losartan 10 mg/kg. In three additional experimental groups, animals were pretreated with enalaprilat: (f) saline; (g) PD-123319, 10 mg/kg, and (h) losartan, 10 mg/kg. Survival for 48 h was significantly improved by PD-123319 (10 mg/kg) (p < 0.05) and by losartan (10 mg/kg) (p < 0.05) as compared with animals injected with saline. Pretreatment with enalaprilat neutralized the protective effect of losartan. PD-123319 is an AT2 agonist and improved survival in this animal model of stroke. Losartan, an AT1 antagonist, also improved survival, possibly through renin release and AT2 stimulation by endogenous AII. This effect was neutralized by enalaprilat.

Animals↗
PubMed1993

Control of anticoagulant and antiplatelet therapy. Managing patients with acute thrombotic disorders.

Indications for using standard anticoagulants, heparin and warfarin; the dosage and route of administration; the importance of monitoring therapy with reliable laboratory indices; and complications of therapy are discussed. Acetylsalicylic acid and ticlopidine can be used as antiplatelet agents. Because their effect on platelet function is not monitored clinically, their clinical indications are emphasized.

Administration, Oral↗
PubMed1992

Immunologic changes after blood transfusion in patients undergoing vascular surgery.

Immunologic changes after blood transfusions cannot be studied ethically in normal individuals. We therefore studied two comparable groups of patients with atherosclerotic cardiovascular disease who received similar drug treatment and experienced a similar degree of surgical trauma, except that one group received an average of 2.5 units of packed red cells at one time period during surgery. We conducted immunologic tests preoperatively and 5, 10, 45 to 60, 90, 180, and 360 days postoperatively. There was no significant difference in all indices tested preoperatively between the two groups. Five and 10 days postoperatively, the absolute numbers of CD3, CD4, CD8, and B cells decreased in both groups; however, the decrease was significantly greater in the transfused group than in the nontransfused group 5 days postoperatively. There was no significant difference in these parameters between the two groups at other time periods tested. At 5 and 10 days postoperatively, the lymphocyte responses to phytohemagglutinin, concanavalin A, and allogeneic lymphocytes in autologous serum were decreased in both groups. However, at 60 days postoperatively, the responses of the nontransfused group became significantly increased, whereas those of the transfused group remained relatively unchanged. By days 90, 180, and 360, the lymphocyte responses of the nontransfused group had dropped to levels seen at earlier time intervals and were comparable to responses in the transfused group. There were no significant differences between the groups in the number of T-cell colonies formed, the number of immunoglobulin-producing cells obtained, and the lymphokine responses (migration inhibitory factor/migration stimulation factor) at all times tested. The major immunologic perturbations attributed to blood transfusions were an exaggerated decrease in the numbers of circulating lymphocytes, particularly those with markers associated with T-helper cells, and failure to demonstrate a rebound increase in the proliferative response 45 to 90 days later.

Adult↗
PubMed1992

Production of interferon by peripheral blood mononuclear cells from normal individuals and patients with chronic lymphocytic leukemia.

Peripheral blood mononuclear cells (PBMC) from normal individuals were studied to identify which cells produce alpha-interferon (IFN-alpha) in response to a virus stimulus. It was found that cells both adherent and nonadherent to plastic formed IFN-alpha after induction by any one of several viruses studied. When nonadherent cells were separated on discontinuous Percoll gradients, only the cells in the less dense Percoll fractions produced IFN, whatever the virus used. By indirect immunofluorescence with monoclonal antibodies to HLA-DR and to Leu 11b, the distribution of the HLA-DR+ cells was shown to resemble most closely that of the IFN-producing population. Elimination of these cells (by complement-mediated lysis with the same antibodies) abrogated the IFN response, but NK cells remained and thus do not produce IFN-alpha. In confirmation, elimination of the Leu 11b+ cells had no effect on the amount of IFN produced. PBMC preparations from patients with chronic lymphocytic leukemia (CLL) appeared incapable of producing IFN-alpha but were shown to contain identifiable IFN-producing cells. The low or absent IFN levels in CLL are probably due to the relative scarcity of IFN-producing cells in their PBMC.

Cell Adhesion↗
PubMed1991

Retardation in tumor growth and metastasis demonstrated in a human CLL B-cell line after the acquisition of an extra chromosome 11.

In this study we described the isolation of a subclonal EBV-transformed human CLL B-cell line with retarded growth rate and metastatic potential from its clonal parent (D10-1), which was karyotyped: 46.XY.dup(1)(q11----q32). The subclone, designated D10-1C, was isolated by limiting dilution of D10-1 following selection in 8-azaguanine-supplemented medium. Chromosome analysis of D10-1C revealed a constitution of 47,XY, + 11.dup(1)(q11----q32). This is the first demonstration that partial trisomy 1q-associated growth advantage in human cancer cells can be retarded by the presence of an additional dose of chromosome 11. Human chromosome 11 had been shown to be responsible for the suppression of tumor development. Whether the same suppressor genes are involved in D10-1C remains to be elucidated. The procedure described here for the enrichment and isolation of a growth-retarded mutant from D10-1 may be applicable in other malignant cell systems.

Animals↗
PubMed1990

Role of 1q trisomy in tumorigenicity, growth, and metastasis of human leukemic B-cell clones in nude mice.

In order to investigate the association between various karyotypes of human tumor cells and biological behavior of tumors such as tumorigenicity, rate of growth, and the capacity to form metastasis, six chromosomally distinctive clones were isolated from an Epstein-Barr virus-transformed human chronic lymphocytic leukemic B-cell line which progressively grew and metastasized in irradiated nude mice. When inoculated into nude mice one clone (D10-1) with the karyotype of 46,XY,dup(1)(q11----q32) was more tumorigenic, grew faster, and produced more metastases than the other five clones. When mixtures of different clone-derived cells were grown in vitro or inoculated s.c. into nude mice the proportion of D10-1 cells was higher than their expected numbers in the cultures, s.c. tumors, and splenic and lymph nodal metastases. The growth and metastatic potential of the D10-1 clone were inhibited when cells from this clone were mixed with one or more clones that had slower growth. Duplication of 1q has been observed as a secondary aberration in human hematological malignancies and solid cancers. Our results demonstrate that duplication of the chromosome segment of 1q11----1q32 is associated with advantages in proliferation and metastasis formation.

Animals↗
PubMed1990

Monoclonal antibodies against Epstein-Barr virus transformed B lymphocytes from a CLL patient.

Immunization of BALB/c mice with EBV-CLL-1 cells, derived from Epstein-Barr virus transformed B lymphocytes from a chronic lymphocytic leukemia (CLL) patient, yielded 2 monoclonal antibodies (IgG1 Kappa and IgG2a Kappa) against a membrane antigen on a subset of normal B lymphocytes and non-Hodgkin's lymphomas. Immunofluorescence revealed strong reactivity of the antibodies with EBV-CLL-1 cells and with most lymphocytes in tonsil follicles, in the intestinal wall, around splenic arterioles and near Hassall's corpuscles in the neonatal thymus as well as with a small proportion of lymphocytes in some large reactive lymph node follicles, weak reactivity with 1/5 of peripheral blood B lymphocytes (PBL), and no reactivity with platelets, granulocytes and non-lymphoid tissues. PBL from 3 CLL patients showed weak staining of only larger cells. Intense fluorescence was observed in several non-Hodgkin's lymphomas of various histological types and in Burkitt's lymphoma lines but not in the 3 T lymphoblastoid and 12 nonlymphoid tumor lines examined. The antibodies precipitated Mr 22,000 and 33,000 bands from surface labeled RAJI or EBV-CLL-1 cells and cross-competed in a binding inhibition assay. The antibodies had approximately 6 million binding sites per EBV-CLL-1 or RAJI cell but were not cytotoxic. This high antigen-density and limited expression in normal cells may permit their use for immunocytological diagnosis and targeting cytotoxic agents and radionuclides against appropriate lymphoma cells.

Animals↗
PubMed1989

Growth of B cell colonies independent of T cell contact.

One of the methods of obtaining B cell colonies involves physically admixing B cells in growth medium containing mitogens with accessory T cells treated with mitomycin C or low dose irradiation to prevent T cell colony formation. However, under these conditions T cells still have the capacity to form colonies in methylcellulose, which is most frequently used for colony assays, and B cells recovered from the colonies are contaminated by the large number of added T cells. Therefore, we have developed a method of growing B cell colonies by physically separating the enriched B cells from the T cells by using millicell-HA inserts (Millipore). The requirement for mitogens, culture conditions, development, and number of B cell colonies formed were similar to those observed with admixture of T and B cells. The advantage of this system is that it permits growth of B cell colonies without the confounding influence of T cell proliferation or treatment of T cells with mitomycin or irradiation, and relatively pure B cell colonies can be recovered for further characterization or functional studies.

Antigens, Differentiation, B-Lymphocyte↗
PubMed1989

Feedback suppression of B cell colony formation in healthy individuals.

Proliferation and differentiation of B cells has been extensively studied and the study of feedback suppression of B cell proliferation has been limited to humoral factors. However, very little is known about feedback suppression of B cell proliferation by cellular influences. We have previously reported on the role of T cells and their subsets on B cell proliferation in that we did not observe suppression of B cell colony growth by T cells. We now report on the role of B cells in limiting B cell proliferation. B cell colonies were grown in methyl cellulose for either 3 days or 5-6 days utilizing 2 x 10(5) T cells irradiated with 9,000 rads, and 2 x 10(5) B cells. The B cells were then obtained from these colonies and increasing numbers of cells were added to fresh autologous B cells that were further cultured for 5 days to form new B cell colonies. At the end of this period, B cell colony numbers were determined. Our data show that addition of CD19- and CD20- positive B cells recovered from mature colonies after 5 days to fresh B cells suppressed further B cell colony growth in all cases tested, whereas addition of CD19-positive B cells recovered from immature colonies after 3 days of culture did not suppress further B cell colony growth. Elimination of CD 19- or CD20-positive cells with monoclonal antibody to CD19 and complement or by the technique of panning enhanced colony growth. Supernatants obtained from B cell colonies did not suppress B cell colony formation. Our data suggest that there is feedback suppression of normal progenitor B cell proliferation by constituent B cells and that this effect develops during maturation of colonies during the growth phase.

Antigens, CD19↗
PubMed1989

Angiogenic properties of normal and psoriatic skin associate with epidermis, not dermis.

Psoriasis is a chronic inflammatory and proliferative epidermal skin disease associated with prominent new vessels in the dermis. Heretofore, unresolved was the question which skin element causes stimulation of vessel formation in this lesion: the epidermis, the dermis, vessels themselves, inflammatory cells, and so forth. It has been argued that the dermis by means of its vessels is responsible for the rapid and prominent epidermal growth and that psoriasis is simply a dermal-induced epidermal disease. Recognizing that vascular proliferations do not occur in the absence of an angiogenic stimulus we asked where in psoriatic skin is that angiogenic element found. By using the rabbit cornea angiogenic assay, the vessel-stimulating properties of epidermis and dermis (separated after cold trypsinization overnight), collected from psoriatic-plaque skin and normal skin both nonpsoriatic and psoriatic patients, were measured. The corneas were examined on days 2, 4, 8, 10, and 12 and graded from 1 to 5+ for vessel growth. Psoriatic plaque epidermis revealed a 4 to 5+ stimulus (8 of 12 corneas). Epidermis from normal subjects or from nonlesional psoriatic patient skin also revealed a 4 to 5+ stimulus (3 of 5 and 5 of 8, respectively). In no case was the dermal implant from normal (7 implants) or psoriatic patients (3 implants of lesional skin) angiogenic. The angiogenic activity of epidermis is stable to freezing and boiling. This study indicates that the vessel-stimulating properties of psoriatic and nonpsoriatic skin are associated with the epidermis and that the developing psoriatic lesion may involve complex epidermal to dermal as well as dermal to epidermal signals.

Adult↗