Evaluation of the in vitro and in vivo antiplasmodial effect of water treated with Photonic Multiphase Modulators (PMM) designed with Advanced Physics System Engineering (APSE™) and BioPhoton-X™ technology.

BACKGROUND: In vitro and in vivo testing of new technology was performed to evaluate the antiplasmodial activity of Photonic Multiphase Modulators (PMM) in cultures and in mice previously infected with Plasmodium falciparum and Plasmodium berghei parasites. METHODS: Cultures of P. falciparum infected-erythrocytes were exposed overnight to two generations of different APSE™ and BioPhoton-X™ PMM (C#1, R#1, R#2, D8 and D9). Growth of parasites was determined through flow cytometry or microscopy. Mice of the strain C57BL/6 were infected and treated with water exposed to second-generation APSE™ and BioPhoton-X™ PMM plus one previously untested first-generation PMM (AGN10). Parasitemia and weight loss were monitored throughout the infection until death or point of euthanasia was reached. After death, necropsy was performed on all animals and the number of days each survived was recorded. RESULTS: In vitro and in vivo testing using different APSE™- and BioPhoton-X™-designed PMM revealed an effect of D8 in lowering the growth of the parasite in vitro, while the best effect in mice was observed with D9 PMM, with a reduced weight loss and an increase in survival, although the results in lowering the parasitemia were inconclusive. D9 PMM did not generate ROS in vitro. CONCLUSIONS: APSE™ and BioPhoton-X™ optic circuit technologies can affect the growth of parasites and show protective effects in mice drinking from water treated with their PMM.

Mitochondria, Energy and Cancer: The Relationship with Ascorbic Acid.

Ascorbic Acid (AA) has been used in the prevention and treatment of cancer with reported effectiveness. Mitochondria may be one of the principal targets of ascorbate's cellular activity and it may play an important role in the development and progression of cancer. Mitochondria, besides generating adenosine triphosphate (ATP), has a role in apoptosis regulation and in the production of regulatory oxidative species that may be relevant in gene expression. At higher concentrations AA may increase ATP production by increasing mitochondrial electron flux, also may induce apoptotic cell death in tumor cell lines, probably via its pro-oxidant action In contrast, at lower concentrations AA displays antioxidant properties that may prevent the activation of oxidant-induced apoptosis. These concentration dependent activities of ascorbate may explain in part the seemingly contradictory results that have been reported previously.

RNA interference: a novel and physiologic mechanism of gene silencing with great therapeutic potential.

The post-genomics scientific era has evolved rapidly while achieving advanced understanding of the structure and function of the genes responsible for both the phenotypic characteristics of higher organisms and the pathophysiology of several genetic diseases. Researchers in the fields of oncology and infectious diseases have become more convinced of the great potential of molecular biology approaches to further develop highly specific diagnostic and less toxic therapeutic strategies. During the last two decades, approaches for the specific silencing of essential viral genes and cellular oncogenes were evaluated with optimism for developing directed therapies. However, there were drawbacks in the use of antisense oligonucleotides as a practical mechanism of achieving gene silencing both in vitro and in vivo. Recently, a novel role for post-transcriptional gene silencing mediated by double-stranded RNA (dsRNA) was discovered in the experimental model of C. elegans. This mechanism, termed RNA interference (RNAi) has also been found in other eukaryotes, from plants to mammals, including humans. RNAi is presently being explored both in vitro and in vivo in functional genomics studies and possible therapeutic uses due to its highly specific and physiologic mode of gene silencing. This article focuses on the most current information available regarding the RNAi mechanism and its uses in models of cancer and infectious diseases.

Why do cells die in HIV infection? Potential mechanisms inducing programmed cell death/apoptosis

This work reviews the suggested mechanisms which result in programmed cell death in human HIV infection. Here we present state-of-the-art scientific information related to the newly rediscovered phenomenon of Apoptosis, and to its biological relevance in the pathogenesis of HIV disease. General features of this phenomenon are reviewed, as well as available evidence for its occurrence and possible role in AIDS pathogenesis. A complex series of cellular and molecular events leading to cellular apoptosis are also reviewed and discussed. They include events which take place at the cell membrane level and those which occur at the intramembrane level and cytoplasmic locations, which result from the immunological activation of affected cells. Cellular events which follow and occur within the mitochondrial space and at the nuclear level are also discussed. The biological significance of all these phenomena is summarized in a theoretical scheme, which attempts to integrate all cellular events leading a primed cell into its HIV-induced programmed death

Multifunctional immunological monitoring of HIV positive patients: a novel staging system

A tri-functional in vitro evaluation has been utilized to analyze peripheral blood mononuclear cells (BMNC) from HIV-infected patients, which allows for the classification of these individuals into convenient stages, according to the number of in vitro parameters affected. The classifying functional parameters are: the mitochondrial metabolic activity of freshly isolated BMNC, measured by an MTT reduction assay, the detection of apoptosis in 72 hour cultures of these cells assessed by propidium iodide staining and dual parametric flow cytometric analysis, and their proliferative response to pokeweed mitogen. Our results indicate that HIV-infected patients at different stages of their clinical disease, can present dysfunctions in one, two or three of the above-mentioned parameters. Based on these results, patients can be classified into four newly-described stages which are Stage 0, including uninfected controls and all patients with unaffected parameters, and Stages 1, 2 and 3, including patients having one, two or all three parameters affected, respectively. This type of immunological evaluation and classification of HIV-infected patients has the potential of becoming a predictive tool in the longitudinal follow-up of their HIV infection

Effect of heat-shock on rhesus monkey lymphocytes and IL-2-dependent cells

Treatment of Rhesus monkey peripheral blood lymphocytes and IL-2 dependent cell lines with heat prior to incubation with mitogens or IL-2, respectively, induces significant cell changes at the nuclear level, detected by DNA staining with Vindelov's propidium iodide and the simultaneous measurement of its red fluorescence and 90 degrees light scatter. These changes are an increase in their nuclear granularity and in apparently fragmented DNA which shows less fluorescence intensity than DNA from nuclei in the G0G1 phase, a phenomenon suggestive of apoptosis. Treated cells also show an increased number of nuclei in G1 or early S phase, with a reduction in those reaching the G2 or M phases. After heat-shock treatment, CTLL-2 cells show an increase in their response to low doses of recombinant IL-2 and an impaired ability to proliferate at higher IL-2 concentrations. These results provide further evidence for the regulatory role of stress-induced events

Breve historia del SIDA en los primates no humanos y su contribución al estudio del síndrome de inmunodeficiencia adquirida / Brief history of AIDS in non-human primates and its contribution to the study of acquired imunodeficiency syndrome

Infection of the rhesus monkey (Macaca mulatta) with retroviruses originating from African non human primates (SIV) induces in this species an acquired immunodeficiency syndrome (SAIDS) closely resembling AIDS in humans. Analogies between the SIV-rhesus system and AIDS in humans are described in this work, analyzing the close relationship existing between the HIV and SIV viruses, and the similarities between SIV disease in the rhesus and HIV disease in humans. A review of current advances in AIDS vaccine research, using the SIV-rhesus model, is also included

Effect of various microbial preparations on P-388 mouse lymphocytic leukemia

Four bacteria-derived immunopotentiators were tested for their protective effect on a P-388 mouse lymphocytic leukemia model. The microbial test products were prepared from the following bacterial strains: ATCC 35983 Staphylococcus epidermidis isolated from a patient with IV catheter; ATCC 31874, a patented strain listed as Staphylococcus epidermidis isolated from the urine of a cancer patient; ATCC 25615 Staphylococcus hominis obtained from a child with lymphocytic leukemia, and ATCC 25614 Staphylococcus warneri, an isolate from a patient with adenocarcinoma of the breast. A limited degree of protection and prolongation in survival time was observed in the animal group treated with the bacterial strain ATCC 31874

Multiparametric analysis of nuclei from activated B and T lymphocytes of rhesus monkeys

Los parámetros físicos de tamaño y granularidad nuclear, y los biológicos de contenido proteico y de ADN nuclear se midieron simultáneamente a través del análisis flujocitométrico de linfocitos provenientes de monos Rhesus activados com mitógenos. El propósito de estos ensayos fué el establecimiento de valores comparables entre el modelo simio y el ya definido modelo humano de activación mitogénica de linfocitos. Se observaron respuestas proliferativas significativas con los mitógenos PHA, PWM, Con A y LPS. Las determinaciones simultáneas de: contenido de ADN/tamaño, contenido protéico/granularidad, contenido de ADN/granularidad; y contenido protéico/contenido de ADN, demostraran la existencia de correlaciones positivas entre estos parámetros. Estos resultados ilustran la validez de su medición en estudios in vitro de activación de linfocitos de simios, y su valor potencial en la evaluación de cambios en función linfocítica, causados por algunas infecciones virales en este modelo experimental

Production of monoclonal antibodies to Sp 2/0-AG14 myeloma cells

La técnica de hibridomas, desarrollada por Kohler y Milstein, permite la obtención de anticuerpos de especificidad única (p.e. anticuerpos monoclonales). Esta metododlogía fue introducida en el Laboratorio de Immunología del Departamento de Microbiología de la Escuela de Medicina, Recinto de Ciencias Médicas, UPR en 1986. La producción de anticuerpos monoclonales contra Sp2/0-Ag 14 que aquí se reporta, describe la metodología utilizada y es de importancia porque constituye el primer trabajo sobre anticuerpos monoclonales realizado en el Recinto de Ciencias Médicas y en Puerto Rico

Analysis by fluorescence microscopy and flow cytometry of Monoclonal antibodies produced against cell surface antigens

La reactividad de siete anticuerpos monoclonales contra antígenos de superficie de las células de mieloma Sp2/O-Ag14 fue analizada simultáneamente por microscopía de fluorescencia y por citometría de flujo. Se incubaron 1 x 10**6 células de Sp2/O-Ag14 con 200 micronl de anticuerpo monoclonal a 4-C por 30 min. Luego de lavar 2 veces con PBS las células fueron incubadas a 4-C por 30 minutos con una dilución 1:400 de anticuerpos de carnero anti-immunoglobulinas de ratón, conjugados a isotiocianato de fluoresceína. Luego de lavar tres veces con PBS, las células se utilizaron para el análisis. Por microscoía de fluorescencia se detectaron diferentes patrones de reactividad con los anticuerpos monoclonales. Estos patrones fueron denominados: homogéneo anular, parchos anulares, parchos putiformes, difuso y homogéneo. Estos patrones podrían representar diferentes espítopes reconocidos monoclonales contra los antígenos de superficie de Sp2/O-Ag14 varió entre un 79 a un 90% al analizarse por cimetría de flujo con el sistema EPICS V. Al compararla con la microscopía de fluorescencia, la citometria de flujo determinó de una forma mas rápida, sensitiva y cuantitativa la reactividad de los diferentes anticuerpos monoclonales contra antígenos de superficie de las células Sp2/O-Ag14