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Results for “SCID mouse”

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PubMed1995

Equine severe combined immunodeficiency: a defect in V(D)J recombination and DNA-dependent protein kinase activity.

V(D)J rearrangement is the molecular mechanism by which an almost infinite array of specific immune receptors are generated. Defects in this process result in profound immunodeficiency as is the case in the C.B-17 SCID mouse or in RAG-1 (recombination-activating gene 1) or RAG-2 deficient mice. It has recently become clear that the V(D)J recombinase most likely consists of both lymphoid-specific factors and ubiquitously expressed components of the DNA double-strand break repair pathway. The deficit in SCID mice is in a factor that is required for both of these pathways. In this report, we show that the factor defective in the autosomal recessive severe combined immunodeficiency of Arabian foals is required for (i) V(D)J recombination, (ii) resistance to ionizing radiation, and (iii) DNA-dependent protein kinase activity.

Animals↗

PubMed1991

Strong priming of T cells adoptively transferred into scid mice.

We have examined the requirements for activating unprimed T cells in vivo by transferring T cells into scid mice, which lack mature B and T cells. Purified adult thymocytes and a protein antigen, keyhole limpet hemocyanin (KLH), were injected into scid mice. scid mice injected with T cells and KLH developed cellular lymph nodes containing CD4+ and CD8+ T cells. Cells recovered from the lymph nodes of injected scid mice proliferated and secreted interleukin 2 in response to KLH in vitro. The results indicate that T cells can be primed to KLH in the scid mouse in the absence of B cells.

Animals↗

PubMed2003

An essential role of the enhancer for murine cytomegalovirus in vivo growth and pathogenesis.

The transcription of cytomegalovirus (CMV) immediate-early (IE) genes is regulated by a large and complex enhancer containing an array of binding sites for a variety of cellular transcription factors. Previously, using bacterial artificial chromosome recombinants of the virus genome, it was reported that the enhancer region of murine CMV (MCMV) is dispensable but performs a key function for viral multiplication (A. Angulo, M. Messerle, U. H. Koszinowski, and P. Ghazal, J. Virol. 72:8502-8509, 1998). In the present study, we defined, through the reconstitution of infectious enhancerless MCMVs, the growth requirement for the enhancer in tissue culture and explored its significance for steering a productive infection in vivo. A comparison of cis and trans complementation systems for infection of enhancerless virus in permissive fibroblasts revealed a multiplicity-dependent growth phenotype that is severely compromised in the rate of infectious-virus multiplication. The in vivo impact of viruses that have an amputated enhancer was investigated in an extremely sensitive model of MCMV infection, the SCID mouse. Histological examination of spleens, livers, lungs, and salivary glands from animals infected with enhancer-deficient MCMV demonstrated an absence of tissue damage associated with CMV infection. The lack of pathogenic lesions correlated with a defect in replication competence. Enhancerless viruses were not detectable in major target organs harvested from SCID mice. The pathogenesis and growth defect reverted upon restoration of the enhancer. Markedly, while SCID mice infected with 5 PFU of parental MCMV died within 50 days postinfection, all mice infected with enhancerless virus survived for the duration of the experiment (1 year) after infection with 5 x 10(5) PFU. Together, these results clarify the importance of the enhancer for MCMV growth in cell culture and underscore the in vivo significance of this region for MCMV virulence and pathogenesis.

3T3 Cells↗

PubMed1992

In vivo efficacy of B43 (anti-CD19)-pokeweed antiviral protein immunotoxin against human pre-B cell acute lymphoblastic leukemia in mice with severe combined immunodeficiency.

A highly aggressive subclone of the human CALLA+C mu+ pre-B acute lymphoblastic leukemia (ALL) cell line NALM-6 (designated NALM-6-UM1) caused disseminated and fatal leukemia in CB.17 mice with severe combined immunodeficiency (SCID). An intravenous challenge with 1 x 10(6) (NALM-6-UM1 cells caused 15 of 27 (56%) SCID mice to become paraplegic at 31 +/- 2 days (median = 33 days) and 27 of 27 (100%) mice to die of disseminated leukemia at 38 +/- 1 days (median = 39 days). We used this SCID mouse model of aggressive human pre-B ALL to evaluate the in vivo antileukemic efficacy of B43 (anti-CD19)-pokeweed antiviral protein (PAP) immunotoxin. A 3-day treatment with nontoxic doses of B43-PAP markedly reduced the incidence of paraplegia and improved event-free survival (EFS) in SCID mice challenged with 1 x 10(6) NALM-6-UM1 pre-B ALL cells, as reflected by significantly higher cumulative proportions of mice free of paraplegia or alive at 1 to 7 months, as compared with phosphate-buffered saline (PBS) treated control mice. The Kaplan-Meier estimates and standard errors of the probability of developing paraplegia after inoculation of 1 x 10(6) NALM-6-UM1 cells was 64% +/- 10% for PBS-treated mice (median time to paraplegia = 37 days) (N = 27), 18% +/- 8% for mice treated with 15 micrograms B43-PAP (5 micrograms/mouse/d x 3 days) (N = 23) and 5% +/- 5% for mice treated with 30 micrograms B43-PAP (10 micrograms/mouse/d x 3 days) (N = 21). While 27 of 27 PBS-treated control SCID mice died of leukemia at 38 +/- 1 days (range = 24 to 54 days), only 16 of 44 B43-PAP-treated mice developed leukemia at 74 +/- 12 days (range = 30 to 182 days), consistent with greater than or equal to 6 logs kill of clonogenic NALM-6-UM1 cells in 64% of SCID mice. The Kaplan-Meier estimates and standard errors of the probability of long-term EFS after inoculation of 1 x 10(6) NALM-6-UM1 cells were 65% +/- 10% for mice treated with 15 micrograms B43-PAP and 60% +/- 11% for mice treated with 30 micrograms B43-PAP with a median survival time of greater than 7 months for both groups. In contrast, neither unconjugated B43 monoclonal antibody nor the anti-T-cell immunotoxin G17.2 (anti-CD4)-PAP decreased the incidence of paraplegia or improved EFS.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed2006

A novel human osteoblast-derived severe combined immunodeficiency mouse model of bone metastasis.

OBJECT: One of the major difficulties of conducting bone metastasis research is the lack of adequate models for studying the bone-tumor microenvironment. The limitations of current in vivo models include the following: non-human tumor or bone, variable reproducibility, limited supply, and an inability to be easily manipulated. The objective of the present study was to develop a uniform and reproducible model of bone/spine metastasis by utilizing bone derived from human osteoblasts grown subcutaneously in severe combined immunodeficiency (SCID) mice with subsequent introduction of human carcinoma cell lines. METHODS: Human osteoblasts were serially passed in culture and induced to differentiate into mature osteoblasts. They were subsequently loaded on hydroxyapatite-coated collagen sponges and implanted subcutaneously into the SCID mice. After allowing the bone to mature for 8 weeks, tumor cell suspensions were implanted percutaneously into the bone. The bone-tumor complexes were subsequently harvested, decalcified, and prepared for histological examination. CONCLUSIONS: The authors have developed a novel, reproducible SCID mouse model of bone/spine metastasis by using bone derived from human osteoblasts and subsequently introduced human tumor lines. They believe this model will be useful for studying the basic biology of bone metastases.

Animals↗

PubMed2001

The allergen-induced airway hyperresponsiveness in a human-mouse chimera model of asthma is T cell and IL-4 and IL-5 dependent.

The cellular and molecular mechanisms involved in the airway hyperresponsiveness (AHR) of patients with allergic asthma remain unclear. A role for Th2 inflammatory cells was suggested based on murine asthma models. No direct evidence exists on the role of these cells in human asthma. The development of a mouse-human chimera might be useful, allowing the in vivo study of the components of the human immune system relevant to asthma. We investigated the role of allergen-reactive T lymphocytes in a human-mouse SCID model. SCID mice were reconstituted intratracheally with human PBMC from healthy, nonallergic, nonasthmatic donors and exposed to an aerosol of house dust mite allergen after i.p. injection with Dermatophagoides pteronyssinus I Ag and alum. The donor T lymphocytes had a Th1 cytokine phenotype. The reconstituted and allergen-challenged mice developed AHR to carbachol. The mouse airways and lungs were infiltrated with human T lymphocytes. No eosinophils or increases in human IgE were observed. The intrapulmonary human T lymphocytes demonstrated an increase in intracytoplasmic IL-4 and IL-5 and a decrease in IFN-gamma after exposure to allergen adjuvant. Antagonizing human IL-4/IL-13 or IL-5 resulted in a normalization of the airway responsiveness, despite a sustained intracellular Th2 cytokine production. These results provide evidence that the activated human allergen-reactive Th2 cells producing IL-4 or IL-5 are pivotal in the induction of AHR, whereas no critical role for eosinophils or IgE could be demonstrated. They also demonstrate that human allergen-specific Th1 lymphocytes can be driven to a Th2 phenotype.

Allergens↗

PubMed1997

Human cytotoxic CD8+ T-lymphocyte clones engraft in severe combined immunodeficient (SCID) mice but show diminished function.

Mice with severe combined immunodeficiency (SCID) provide an in vivo model for studying interactions between human tumor cells and effector cells of the immune system. We studied the behavior of human alloreactive cytotoxic T lymphocytes (CTLs) in SCID mice, including the migration pattern of CD8+ or CD4+ CTL clones to various murine tissues, their engraftment in the absence or presence of recombinant human interleukin 2 (rhIL-2) compared with the engraftment of lymphokine activated killer (LAK) cells, and the in vitro as well as the in vivo function of the engrafted CTL clones. The polymerase chain technique using T-cell-receptor-gamma specific primers revealed the presence of human CD8+ CTL clones in the blood, lungs, and liver of mice receiving rhIL-2 for 14 days, for at least 18 days after intravenous inoculation. Human T cells were transiently detected in the bone marrow, lymph nodes, thymus, and spleen. Although the three CD8+ CTL clones and two CTL lines tested required rhIL-2 for their engraftment, the four LAK cell populations also engrafted in the absence of rhIL-2. In contrast to CD8+ T cells, only low frequencies (<1%) of CD4+ cells could be detected in the blood of the SCID mouse. Engrafted human T cells recovered from the murine blood showed absent or diminished cytotoxicity in vitro against human target cells in five or six experiments, respectively. In addition, when the antitumor activity of engrafted CD8+ clones was investigated in vivo using a xenotransplantation model of human B-cell lymphoma in SCID mice, no significant prolongation of the mean survival time of six treated animals was observed compared with animals treated with rhIL-2 alone. Our results illustrate that although in vitro primed and cultured human CD8+ T cells engraft in SCID mice, their in vitro and in vivo function is impaired.

Animals↗

PubMed2000

Human dendritic cells in the severe combined immunodeficiency mouse model: their potentiating role in the allergic reaction.

Dendritic cells (DCs) are present in the lungs and airways of healthy and allergic subjects where they are exposed to inhaled antigens. After the uptake of antigens, DCs migrate to lymphoid organs where T cells initiate and control the immune response. The migratory properties of DCs are an essential component of their function but remain unclear in the situation of allergic diseases. To better understand the role of DCs in response to allergens, we first investigated their presence in an original experimental model of allergic asthma: the humanized severe combined immunodeficiency (SCID) mouse reconstituted with peripheral blood mononuclear cells from patients sensitive to Dermatophagoides pteronyssinus (Dpt). Human DCs were detected in lungs of mice developing an inflammatory pulmonary infiltrate and appeared to be mainly located in the alveolar spaces. In a second step, human DCs were generated in vitro from monocytes and injected into naive SCID mice exposed or not exposed to Dpt aerosols. Their migratory behavior was explored, as well as their potential role in modulating the IgE production after exposure to Dpt. After exposure to Dpt, the number of DCs present in airways decreased, while it increased into the spleen and thymus of the mice. The IgE production increased in the presence of DCs as compared with mice not injected with DCs. These results suggest that DCs may play a role in the pulmonary allergic reaction developed in response to Dpt in SCID mice.

Adoptive Transfer↗

PubMed1998

Interleukin-4 and interleukin-10 are chondroprotective and decrease mononuclear cell recruitment in human rheumatoid synovium in vivo.

We used the severe combined immunodeficient (SCID) mouse model to assess the effect of interleukin-4 (IL-4) or IL-10 injection on cartilage degradation and mononuclear cell (MNC) recruitment to human rheumatoid synovium in vivo. Human rheumatoid synovium and cartilage from five rheumatoid arthritis patients, obtained after joint replacement surgery, were engrafted subcutaneously to 6-8-week-old SCID CB17 mice. Synovial tissues were injected with recombinant human IL-4 (rhIL-4, 100 ng; rhIL-10, 100 ng), both cytokines, or tumour necrosis factor-alpha (TNF-alpha) (1000 U), or phosphate-buffered saline twice a week for 4 weeks. The graft was removed and immunochemical analysis was carried out to assess intracellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-selectin expression. Moreover, cartilage degradation was assessed through the quantification of the erosion surface on a computerized image of the engrafted cartilage at high power view. MNC recruitment in the synovial tissue was determined by labelling blood MNC with indium-111 before their intraperitoneal injection. The activity obtained in the region of the graft were determined with a gamma camera 72 hr postinjection. The results are expressed as a percentage of initial injected activity. After 4 weeks we observed a decrease of cartilage area in controls (77 +/- 8%), inhibited after injection of IL-4, IL-10, or both cytokines (90 +/- 3%, 89.1 +/- 4%, 89.2 +/- 5% respectively), and 57 +/- 17% after TNF-alpha injection. The % MNC activity in the graft decreased to 77 +/- 81% (NS), 9 +/- 4% (P < 0.003) and 19 +/- 6% (P < 0.007) compared with untreated synovial tissue after treatment with IL-4, IL-10, or both cytokines, respectively. Moreover, IL-10 but not IL-4 decreased the expression of ICAM-1 but not VCAM-1 or E-selectin by synovial cells. These results suggest that IL-10 and IL-4 could have chondroprotective properties, and that IL-10 but not IL-4 inhibits MNC traffic towards the synovial tissue efficiently.

Animals↗

PubMed1993

Adoptive transfer of cytotoxic T lymphocytes for the treatment of transplant-associated lymphoma.

BACKGROUND: Immunocompromised organ transplant recipients have a high incidence of B cell lymphomas (BCL). Severe combined immunodeficient (SCID) mice develop human BCL when engrafted with Epstein-Barr virus (EBV) transformed and immortalized B lymphoblastoid cell lines (BLCL). Because a lack of effective EBV-specific cytotoxic T lymphocytes (EBV-CTL) is thought to lead to lymphoma development, the SCID mouse model was used to determine the relationship between EBV-infected B cells and EBV-specific CTL in BCL development in vivo. METHODS: EBV-CTL were generated by in vitro stimulation of peripheral blood leukocytes with autologous BLCL. CD8+ CTL were isolated from CTL populations by depletion of CD4+ cells. SCID mice were engrafted with BLCL, EBV-CTL were adoptively transferred into engrafted SCID mice either immediately or 7 days after engraftment, and the animals were monitored for the development of BCL. Statistical significance was determined by the log rank test. RESULTS: SCID mice engrafted with BLCL rapidly developed BCL (mean, 20 days). SCID mice engrafted with BLCL and human leukocyte antigen-identical EBV-CTL or CD8+ EBV-CTL had a significant delay in BCL development (p < 0.05), whereas some mice did not develop BCL. In contrast, human leukocyte antigen-nonidentical EBV-CTL did not significantly delay BCL development. CONCLUSIONS: This study showed the role of EBV-CTL in inhibiting the development of BCL. A greater understanding of the cellular and viral interactions leading to B-cell transformation and malignancy may allow the development of specific interventional therapies in patients who have received immunosuppressants.

Adult↗

PubMed1997

Tallimustine, an effective antileukemic agent in a severe combined immunodeficient mouse model of adult myelogenous leukemia, induces remissions in a phase I study.

Despite progress in leukemia therapy, only 20-30% of patients with acute myelogenous leukemia (AML) are cured. 1-beta-D-arabinofuranosylcytosine- and topoisomerase II-reactive drugs are the primary therapeutic agents used. The aim of this study was to evaluate the potential activity of tallimustine in leukemia. In this study, we first investigated the efficacy and toxic effects of tallimustine, a distamycin-A derivative, in a human leukemia model in severe combined immunodeficient (SCID) mice. On the basis of its dramatic activity in this preclinical study, a Phase I study of tallimustine at a starting dose of 300 microgram/m2/day for 3 days every 3-4 weeks was conducted in patients with refractory or relapsed leukemia. In SCID mice grafted with a human myelomonocytic leukemia cell line, tallimustine resulted in complete remission of disease in most mice at tolerable dosages ranging from 0.86 to 3.0 mg/kg/day for 3 days and was combined effectively and safely with a 2-day schedule of high-dose ara-C. In the Phase I study, 26 patients with refractory or relapsed leukemia were treated. The maximum tolerated dose was 900 microgram/m2/day for 3 days every 3-4 weeks. This dose was 3 times higher than the maximum tolerated dose in solid tumors and was limited by severe mucositis. Magnesium and potassium wasting were also observed, but other side effects (fatigue and gastrointestinal) were minor. Two (8%) patients with AML achieved complete remission and two achieved hematological improvement with persistent thrombocytopenia. The results of this study indicate that tallimustine has promising activity in AML. Future studies may combine tallimustine with other agents known to be active against AML, and investigate its activity in other hematological malignancies. The recommended Phase II single-agent dose of tallimustine is 750-900 microgram/m2/day for 3 days, and combination studies may start at 50-66% of this dose schedule. The SCID mouse model of human leukemia may be promising in the preclinical evaluation and selection of potential antileukemic agents.

Adolescent↗

PubMed1996

Human T-cell lymphotropic virus type-I infection in the severe combined immunodeficiency mouse.

Human T-cell lymphotropic virus type-I (HTLV-I) is the etiologic agent of HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and adult T-cell leukemia (ATL). HAM/TSP and ATL occur infrequently among HTLV-I-infected individuals, and rarely develop in the same individual. To study host and viral factors involved in the induction, tissue tropism, as well as pathogenesis of HAM/TSP, peripheral blood lymphocytes (PBL) from 14 patients with HAM/TSP and from 9 controls were introduced into severe combined immunodeficiency (SCID) mice by intraperitoneal injection. Mice were followed for up to 26 weeks. Human IgG was produced from 2 to 14 weeks after reconstitution in all animals. Thirty-two of 44 mice (72%) showed circulating human antibody against the major viral protein products of HTLV-I. Analysis of viral sequences by polymerase chain reaction (PCR) demonstrated HTLV-I sequences in 21/38 (55%) brains and in 7/17 (41%) spinal cords from HTLV-I-hu SCID mice. No animal had clinical evidence of neurological impairment or pathological findings similar to those seen in HAM/TSP. Seven mice who received PBL from Epstein Barr virus (EBV)-seropositive patients developed an intraperitoneal lymphoma. In 2 mice an infiltration of brain by a lymphoblastic tumor of B/T cell type was observed. By PCR, all the tumors were EBV-positive; HTLV-I sequences were detected in 5 of them. Our study suggests that the HTLV-I-hu-SCID mouse provides a potentially valuable system for studying the production, kinetics, and pathogenicity of anti-HTLV-I antibody, and may help clarify the interaction of EBV and retroviruses in the development of disease.

Adult↗

PubMed1988

Transcription of unrearranged antigen receptor genes in scid mice.

The scid mouse mutant is severely deficient in lymphocytes; cells of the B or T lymphocyte lineage cannot be detected by either serological or functional assays. However, as shown here, germ-line transcripts of B cell immunoglobulin (Ig) constant and variable region genes and of T cell receptor (TCR) genes are detectable in lymphopoietic tissues of scid mice, as well as B and T lineage-specific lambda 5 and T3 delta transcripts. We conclude that B and T lineage-committed cells do arise in scid mice and that their Ig and TCR genes are accessible to enzymes involved in their recombination. This suggests that scid impairs lymphopoiesis at the stage at which antigen receptor genes normally undergo rearrangement.

Animals↗

PubMed1995

Sex-dependent liver metastasis of human melanoma lines in SCID mice.

The liver metastasis formation of two human melanoma cell lines were compared in male and female SCID mice. The intrasplenic injection of both tumour lines resulted in a significantly higher number of liver metastases in male than in female mice; the incidence and weight of spleen tumours, as well as the incidence of metastases were similar. Both melanoma cell lines bound fluorescent oestradiol, progesterone and testosterone conjugates, and proved to be positive for oestrogen receptor-related protein by immunocytochemistry. These observations support the view that endocrine factors influence the progression of human melanomas. This SCID mouse model could be useful in studying the effects of hormonal manipulations on human melanoma metastases.

Adult↗

PubMed2004

Methotrexate (MTX) and albumin coupled with MTX (MTX-HSA) suppress synovial fibroblast invasion and cartilage degradation in vivo.

OBJECTIVE: To investigate the effect of methotrexate (MTX) and albumin coupled with methotrexate (MTX-HSA) on cartilage invasion and induction of perichondrocytic degradation by rheumatoid arthritis synovial fibroblasts (RA SF) in vivo. METHODS: Human cartilage and RA SF were co-transplanted in three groups of severe combined immunodeficient mice (SCID), which received 1 mg/kg free MTX (n = 9), 1 mg/kg MTX-HSA (n = 6), or 0.9% NaCl (n = 5), respectively, intraperitoneally twice a week. After 4 weeks' treatment, the mice were killed and the implants analysed histologically. RESULTS: The control group had a mean (SEM) score for cartilage invasion of RA SF of 2.0 (0.26) and for perichondrocytic cartilage degradation of 1.5 (0.34). In contrast, mice which received MTX showed a significantly reduced invasion (0.78 (0.14), p<0.01) and a reduction in perichondrocytic cartilage degradation scores (0.69 (0.2), p<0.05) in comparison with the control group. Mice treated with MTX-HSA also had significantly reduced scores for RA SF invasion into the cartilage (0.92 (0.41), p<0.05) and for cartilage degradation (0.83 (0.44), p<0.05) compared with controls. The effects of MTX and MTX-HSA were not significantly different between these two groups. CONCLUSION: Treatment with MTX or MTX-HSA significantly ameliorates cartilage destruction in the SCID mouse model for human RA.

Albumins↗

PubMed1998

Chemically modified tetracyclines inhibit human melanoma cell invasion and metastasis.

Recent work has shown that chemically modified tetracyclines (CMTs) are potent inhibitors of matrix metalloproteinase (MMP) activity, both in vitro and in vivo, which is distinct from their antimicrobial activities (Golub et al. Crit Rev Oral Biol Med, 2, 297-321, 1991; Ryan et al. Curr Opin Rheumatol, 8, 23847, 1996). The process of tumor cell invasion requires MMP-mediated degradation of extracellular matrix barriers as a key step in the metastasic cascade. In this study, we examined the effect(s) of doxycycline and CMTs on extracellular levels of gelatinase A and B activity from a highly invasive and metastatic human melanoma cell line C8161, and correlated these observations with changes in the cells' biological behavior in an in vitro invasion assay and in an in vivo SCID mouse model. The results indicate that coincident with the ability of these compounds to differentially suppress extracellular levels of gelatinase activity, C8161 cells treated with doxycycline, CMT-1, CMT-3, or CMT-6 were less invasive in vitro in a dose-dependent manner (3-50 microg/ml). Furthermore, data derived from the in vivo model indicate that SCID mice dosed orally with CMT-1 or CMT-3 contained a reduced number of lung metastases following i.v. injection of C8161 cells via tail vein inoculation. These observations suggest that careful screening of different CMTs could lead to the identification of compounds which suppress the formation and magnitude of metastases associated with certain cancers, and if used as an adjunct to other treatment regimes, lead to greater efficacy in the treatment of metastatic cancers.

Animals↗

PubMed2005

Low pH immobilizes and kills human leukocytes and prevents transmission of cell-associated HIV in a mouse model.

BACKGROUND: Both cell-associated and cell-free HIV virions are present in semen and cervical secretions of HIV-infected individuals. Thus, topical microbicides may need to inactivate both cell-associated and cell-free HIV to prevent sexual transmission of HIV/AIDS. To determine if the mild acidity of the healthy vagina and acid buffering microbicides would prevent transmission by HIV-infected leukocytes, we measured the effect of pH on leukocyte motility, viability and intracellular pH and tested the ability of an acidic buffering microbicide (BufferGel) to prevent the transmission of cell-associated HIV in a HuPBL-SCID mouse model. METHODS: Human lymphocyte, monocyte, and macrophage motilities were measured as a function of time and pH using various acidifying agents. Lymphocyte and macrophage motilities were measured using video microscopy. Monocyte motility was measured using video microscopy and chemotactic chambers. Peripheral blood mononuclear cell (PBMC) viability and intracellular pH were determined as a function of time and pH using fluorescent dyes. HuPBL-SCID mice were pretreated with BufferGel, saline, or a control gel and challenged with HIV-1-infected human PBMCs. RESULTS: Progressive motility was completely abolished in all cell types between pH 5.5 and 6.0. Concomitantly, at and below pH 5.5, the intracellular pH of PBMCs dropped precipitously to match the extracellular medium and did not recover. After acidification with hydrochloric acid to pH 4.5 for 60 min, although completely immotile, 58% of PBMCs excluded ethidium homodimer-1 (dead-cell dye). In contrast, when acidified to this pH with BufferGel, a microbicide designed to maintain vaginal acidity in the presence of semen, only 4% excluded dye at 10 min and none excluded dye after 30 min. BufferGel significantly reduced transmission of HIV-1 in HuPBL-SCID mice (1 of 12 infected) compared to saline (12 of 12 infected) and a control gel (5 of 7 infected). CONCLUSION: These results suggest that physiologic or microbicide-induced acid immobilization and killing of infected white blood cells may be effective in preventing sexual transmission of cell-associated HIV.

Acids↗

PubMedDate not supplied

Orthotopic transplant mouse models with green fluorescent protein-expressing cancer cells to visualize metastasis and angiogenesis.

There have been major efforts in metastasis research in recent years, especially on the role of angiogenesis in the metastatic process. Much of the information in this area has been obtained from model systems that are not representative of clinical cancer. The technique of surgical orthotopic implantation (SOI) has allowed the development of clinically relevant metastatic models of human cancer in immunodeficient rodents such as the nude and SCID mouse. In order to allow direct visualization of the metastatic process, we took advantage of the green fluorescent protein (GFP) of the jellyfish, Aequorea victoria. A series of cancer cell lines have been stably transfected with vectors containing humanized GFP cDNA. To utilize GFP expression for metastasis studies, fragments of subcutaneously growing tumor, which were comprised of GFP-expressing cells, were implanted by SOI in nude mice. Subsequent metastases were visualized in systemic organs by GFP fluorescence in the lung, liver, bones, brain and other organs down to the single-cell level. With this fluorescent tool, we detected and visualized for the first time tumor cells at the microscopic level in fresh viable tissue in their normal host organs even in the live animal. Angiogenesis is readily visualized in the transplanted GFP-expressing tumors in real time in situ in the live animal using simple laparotomy and fluorescent techniques. The results with the GFP-transfected tumor cells, combined with the use of SOI, demonstrate a fundamental advance to visualize and study cancer metastasis and the role of angiogenesis and other factors in the metastatic process.

Animals↗