Appearance Of Transformation Research Articles

Prediction of Transformation of Acute Disseminated Encephalomyelitis into Multiple Sclerosis

Prediction of transformation of acute disseminated encephalomyelitis (ADEM) into multiple sclerosis is of great clinical importance, as it enables timely determination of the tactics for treatment of a patient as well as volume of respective therapeutic interventions.This work is aimed at ascertainingthe prognostic factors that determine the risk of transformation of acute disseminatedencephalomyelitis into multiple sclerosis. We have examined 101 patients with the diagnosis ADEM, namely: 28 men and 73 women in the age from 17 up to 53 years (average value31.7 ± 1.01years). To ascertain the prognostic meaning of clinic-paraclinic indices corresponding to patients with ADEM, we estimated the cumulative part of absence of relapses in the group of patients by using the Kaplan-Meyer method with estimating the Fisher criterion and using the most important clinic-paraclinic data. Development of transformation of ADEM into multiple sclerosis is reliably related to the following prognostic signs: degree of disability in accord with the EDSS scale and sizes of demyelination focuses determined using MRT. Criteria for congenial prediction in disease development with delayed appearance of transformation of ADEM into multiple sclerosis are as follows: slight degree (in EDSS scale) of disability and large sizes of demyelination focuses (MRT data).Our analysis of the main clinic-paraclinic indexes obtained using the Kaplan-Meyer method indicates reliability of results and enables us to find a number of important prognostic criteria for development of transformation of ADEM into multiple sclerosis.

Induction and Maintenance Therapy for Patients with Mycosis Fungoidus (MF) and Sezary Syndrome (SS) in Large Cell Transformation Phase.

Abstract 4758 AimTo determine the frequency of transformation of cutaneous T-cell tumors (CTCL) and effect of different therapies. Materials and methodsRetrospective analysis of treatment results for 68 patients with two types of CTCL, MF and SS, selected from 263 patients T-cell lymphoma/leukemia treated at NCH during 9 years. All 68 patients have CD 3 +, CD 4+, CD 7-, CD 8-, CD 30- immunophenotype and T-cell clonality (rearranged genes for T-cell receptor). MF was diagnosed in 48 patients (24 men, 24 women) with median age 52 years (26 - 77), SS in 20 patients (12 men, 8 women), median age 61 years (23 -87). Half of the MF patients started surveillance at III - IV stages, 15 of 20 patients with SS - at stage IV. Treatment of CTCL in the indolent phase was conducted in accordance with their stage: monochemotherapy (leukeran or methotrexate) and biologically active agents (interferons, retinoids, monoclonal antibody) were used. Appearance of transformation to large cell lymphoma was consistent with tumor progression as determined by presence of two or more of the following: 1. rapid growth of tumors in MF or leukemization in SS; 2. presence of large anaplastic cells in blood or tumor biopsy; 3. increase in expression of Ki-67 (10-70%); 4. appearance of activation markers (CD30, CD25) in tumor cells, and 5. loss of certain linear T-cell markers. For treatment of patients in large cell transformation phase Promace-Cytabom or FNC protocols were used in 7 patients. Five patients received A-CHOP-14 (with alemtuzumab) and subsequent maintenance therapy (methotrexate 20 mg per week, 6-mercaptopurine 50-100 mg/day in combination with alpha interferon 3 million daily). Results and DiscussionThe overall median time of observation (before and during treatment) in 48 patients with MF was 96 months (8-264). For 13 patients who died it was 76 months (8-264), mortality rate was 27%. The overall median observation (before and during treatment) in 20 patients with SS was 60 months (6-123), for 8 who died, it was 62 months (6-94), mortality 40%. The median observation for treatment of MF was 30 months (from 3-122) for the SS, it was 18 months (1 - 85). Restraining aggressive therapy for an indolent process resulted in complete or partial remission, while maintaining a good quality of life. Large cell transformation phase was observed in 12 patients (7 of 48 patients with MF (14.5%) and 5 of 20 patients with SS (25%). Seven of the 12 patients died within 2-4 months due to septic complications during Promace-Cytabom or FNC protocols. The remaining 5 patients received A-CHOP-14 (6-7 courses) which reduced tumor formation and /or leukemization. However, the persistence of patches, plaque or erythrodermia was the reason for maintenance therapy. This combination led to further regression of skin manifestations and full or partial remission within 4-12 months. ConclusionBecause of the lack of a generally accepted treatment for GM and SS in Large Cell Transformation phase, even small successes should be discussed. The effectiveness of combining of polychemotherapy with maintenance therapy may be effective due to the presence of several distinct clones of tumor cells with different properties. Keywords: treatment of cutaneous T-cell lymphoma, Mycosis Fungoidus, Sezary Syndrome, Large Cell Transformation Phase, immunohistochemical markers of T-cell lymphomas. Disclosures:No relevant conflicts of interest to declare.

Stimulatory effects of compounds present in the rhizosphere on natural transformation of Acinetobacter sp. BD413 in soil

The stimulation of natural transformation of Acinetobacter sp. BD413(pFG4) cells by various organic compounds naturally found in the rhizosphere of crop plants was determined in sterile and non-sterile soil. Stimulation of bacterial competence by the organic acids, amino acids, sugars and inorganic fertilizers, was monitored by the transformation frequencies obtained with added cell lysates of Acinetobacter sp. carrying a kanamycin resistance gene ( nptII). Clear differences in the capability of the organic compounds to induce gene capture were detected. In sterile soil, most of the organic compounds used stimulated the appearance of transformants. Fewer compounds were able to induce transformation in non-sterile soil, and a high-P salt supplementation was found to be required for detectable gene transfer. The transformation frequencies in non-sterile soil were, however, if found, on average less than 10-fold reduced compared to those obtained under sterile soil conditions. Of the organic acids and amino acids applied in non-sterile soil, acetate, lactate and alanine gave the highest transformation frequencies, i.e. 1 to 2×10 −6. Of the sugars used, only glucose produced a few transformants in non-sterile soil (2.5×10 −7). Mixtures of the organic and amino acids, or the sugars, and the high-P salts produced the highest transformation frequencies in non-sterile soil, of 4.4 to 6.6×10 −6. The addition of fertilizers had a less pronounced effect on the transformation frequencies. We conclude that many of the compounds exuded into the rhizosphere of agriculturally grown plants are able to stimulate natural transformation of Acinetobacter sp. BD413(pFG4) cells in sterile silt loam soil, and that the presence of both carbon sources and inorganic salts is required for detectable gene transfer in non-sterile silt loam.

Restriction enzyme-mediated DNA integration in Coprinus cinereus.

Restriction enzyme-mediated DNA integration (REMI) has recently received attention as a new technique for the generation of mutants by transformation in fungi. Here we analyse this method in the basidiomycete Coprinus cinereus using the homologous pabI gene as a selectable marker and the restriction enzymes BamHI, EcoRI and PstI. Addition of restriction enzymes to transformation mixtures results in an earlier appearance of transformants and influences transformation rates in an enzyme- and concentration-dependent manner. Low concentrations of restriction enzyme result in increased numbers of transformation rates decrease with higher enzyme concentrations. If protoplasts are made from cells stored in the cold, the transformation rates drop drastically even in the presence of low amounts of enzyme. In several transformants, plasmid integration directly correlated with the action of restriction enzyme at random chromosomal restriction sites. In some cases, restriction enzymes appear to reduce the number of integration events per transformant. Simultaneously, mutation rates can be enhanced due to the presence of restriction enzymes. Although restriction enzymes clearly promote plasmid integration into the host genome they also have cytotoxic and possibly mutagenic effects that result from processes other than plasmid integration. In consequence, for any given enzyme used in REMI mutagenesis, the enzyme concentration that gives the highest number of transformants must be defined experimentally. Such optimal transformation conditions should give the highest probability of obtaining mutations caused by a single restriction enzyme-mediated integration of the selection marker.

The effect of magnesium of transfection and transformation in Bacillus subtilis.

The influence of Mg2+ on phage SPP1 DNA-mediated transfection as compared with chromosomal transformation was studied. A differential influence of this cation in both processes was detectable by analyzing the competence development and the kinetics of appearance of transformants and transfectants. Binding and uptake of DNA and release of acid-soluble products by competent cells in high- and low-Mg2+ media was measured for 3H-labelled SPP1 and 3H-labelled Bacillus subtilis DNAs. Phage SPP1 DNA was shown to be subjected to endonucleolytic cleavage after exposure to competent cells.

Enhancement of outgrowth of EB virus-transformed cells from normal human peripheral blood by a tumor promoter, TPA.

Effect of 12-0-tetradecanoyl-phorbol-13-acetate (TPA) on the establishment of lymphoblastoid cell lines (LCL) following spontaneous tranformation of B cells by Epstein-Barr virus (EBV) from peripheral lymphocytes of healthy EBV-seropositive adults was examined. In the lymphocyte culture with TPA (0.5 ng/ml), the frequency of establishment of LCL increased and the time required for appearance of transformation decreased. A larger number of EBV-positive cells in the peripheral blood were detected in the assay by co-cultures with cord blood lymphocytes in the presence of TPA than in the absence of it. The enhancement of LCL establishment by TPA was largely abolished in the culture with EBV neutralizing antibody-containing human umbilical cord serum. This suggests that the enhancement by TPA may be caused mostly by transformation of coresident B lymphocytes infected with active virus induced from the EBV genome-positive cells by TPA.

Identification of the target cells in human B lymphocytes for transformation by epstein-barr virus

Abstract Lymphocytes from human umbilical cord blood were purified and then separated into rosette-forming cells (T cells) and non-rosette-forming cells (non-T cells). Non-rosette-forming cells were further divided by the technique of rosette formation, after neuraminidase treatment, into surface immunoglobulin (SIg)-carrying cells (B cells) and cells lacking SIg but carrying Fc receptors (null cells). The three cell subsets, T, B, and null cells, were examined for susceptibility to transformation by the B95-8 strain of Epstein-Barr virus (EBV) using the criteria of colony formation by transformed cells and/or transformation efficiency as judged by the days required for the first appearance of transformation. Not only the T cells but also the null cells were unsusceptible to transformation by EBV. In contrast, B cells were highly susceptible. In a study of the quantitative relationship between the target cells for viral transformation and those B cells which possessed SIg after an acid pH treatment, 10 lymphocyte preparations from three cord blood samples and seven adult peripheral blood samples were tested individually. The fraction size of transformable cells was determined by the growth-curve procedure for transformed cells while the procedure of direct membrane immunofluorescence with anti-IgM (μ-specific) serum and polyvalent anti-Ig serum was used to determine the fractions of SIg and SIg(M) cells. The actual fraction of EBV target cells was nearly equal to that of SIg(M) B cells but not to that of the total SIg B cells. Thirty-four lymphocyte preparations from eight cord and 26 adult peripheral blood samples were examined for the percentage of SIg B cells and for susceptibility to EBV transformation as assayed by the colony-formation procedure. EBV susceptibility and the fraction of SIg(M) B cells, but not of total SIg B cells, correlated nicely, suggesting that SIg(M)-bearing cells were probably the major target among the B lymphocytes for transformation by EBV.

Intercellular effects on development of competence in Bacillus subtilis.

The intercellular transfer of competence during growth under the conditions specified by the transformation procedure of Spizizen was investigated with Bacillus subtilis 168. The rate of competence development as assayed uniformly in medium B was not affected by variations in the cell concentration, although the first appearance of transformants occurred earlier with high cell densities in medium A, approximately in proportion to the onset of the stationary phase in the culture. Growth in the presence of Pronase enhanced the frequency of transformation, but did not detectably alter the kinetics of competence development. The rate of competence increase in physiologically noncompetent cultures was not changed by mixing with competent cultures either in medium A or in medium B; however, an early appearance of transformants was noted in mixed cultures in which the proportion of competent to noncompetent cells prevented exponential growth of the noncompetent strain. These experiments indicate that the normal development of competence in B. subtilis is not mediated by a soluble or loosely bound protein factor capable of transmitting competence directly via cell contact. The onset of competence is thus a function of internal physiological changes which are induced by the overall metabolic state of the culture.

Sequential entry of transforming markers into Neisseria meningitidis after chromosome alignment.

The kinetics of appearance of transformants as a function of time of exposure to deoxyribonucleic acid (DNA) was examined in Neisseria meningitidis. Incubation with chloramphenicol for as long as 2 hr, which probably leads to chromosome alignment, resulted in augmentation of the lag period before the appearance of the first transformants. The lag periods thus found were dependent upon the marker tested. This permitted the construction of a time map according to the lag periods observed for individual markers. This map was in general agreement with the chromosome map of the recipient strain as determined by marker frequency analysis. Transformation of recipient cells with chromosomes aligned by growth to the stationary phase showed the same type of increased lag in the appearance of transformants before the logarithmic phase of growth had again been reached. These results support the assumption that the nature of the marker accepted by a recipient cell corresponds to the marker present at the replication point of the chromosome. In the absence of DNA and protein synthesis, the uptake of one marker seems to be successively followed by other markers in a linear order determined by the chromosome of the recipient cell.

LAG PERIOD CHARACTERIZING THE ENTRY OF TRANSFORMING DEOXYRIBONUCLEIC ACID INTO BACILLUS SUBTILIS.

Levine, James S. (Yale University, New Haven, Conn.), and Norman Strauss. Lag period characterizing the entry of transforming deoxyribonucleic acid into Bacillus subtilis. J. Bacteriol. 89:281-287. 1965.-The kinetics of appearance of transformants as a function of time of exposure to deoxyribonucleic acid (DNA) has been studied in Bacillus subtilis. A short lag period of approximately 1 min at 37 C is evident when the transformation is terminated with deoxyribonuclease. The length of this lag is independent of the genetic trait transferred. Moreover, the lag is unaffected by transforming DNA concentration, by the presence of homologous unmarked DNA, and by shearing and cross-linking of the transforming DNA. The lag shows a strong inverse temperature dependence. The energy of activation is 13.9 kcal. The lag is abolished when the transformation is terminated by washing instead of by addition of deoxyribonuclease. These results are taken to indicate an immediate adsorption of DNA to cells, followed by a deoxyribonuclease-sensitive period of 1 min, during which time the genetic trait is entering the cell. These results make feasible an investigation of the configuration assumed by the DNA molecule during entry into the cell.