PubMed Health⌕ Search

Biomedical subjects

J H Pavlovitch

Publications and source records attributed to J H Pavlovitch.

At least 19 recordsLinked to original sources
PubMed1999

Distribution of cycling T lymphocytes in blood and lymphoid organs during immune responses.

Proliferation of murine T lymphocytes in blood, lymph nodes, and spleen was studied in four in vivo stimulation systems, using BrdU pulse-labeling of DNA-synthesizing cells. The T cell response to the superantigen Staphylococcus enterotoxin B (SEB) was studied in detail. Vbeta8+ T cells showed a peak of DNA synthesis 16-24 h after SEB injection, and the percentage of BrdU+ CD4 and CD8 T cells was higher in blood than in lymph nodes and spleen. DNA synthesis was preceded by massive migration of Vbeta8+ cells from blood to lymphoid organs, in which the early activation marker CD69 was first up-regulated. SEB-nonspecific Vbeta6+ cells showed minimal stimulation but, when cycling, also expressed a high level of CD69. The other systems studied were injection of the IFN-gamma inducer polyinosinic:polycytidylic acid, infection by the BM5 variants of murine leukemia virus (the causative agent of murine AIDS), and T cell expansion after transfer of normal bone marrow and lymph node cells into recombinase-activating gene-2-deficient mice. In each case, a peak of T cell proliferation was observed in blood. These data demonstrate the extensive redistribution of cycling T cells in the first few hours after activation. Kinetic studies of blood lymphocyte status appear crucial for understanding primary immune responses because cycling and redistributing T lymphocytes are enriched in the circulating compartment.

Acute Disease↗
PubMed1996

Resistance to murine AIDS in offspring of mice infected with LP-BM5. Role of CD8 T cells.

The murine-acquired immunodeficiency syndrome (MAIDS) is caused by a mixture of murine leukemia viruses (LP-BM5 MuLV). The influence of perinatal contact with retroviruses or their Ags on the response to infection was tested by infecting with LP-BM5 (MuLV) the adult offspring of mice with MAIDS. These offspring were resistant to disease after virus challenge. Most of them were free of defective viral DNA, and even those with molecular evidence of infection had lymphoid cells with a lower infectious capacity to cause MAIDS in naive recipients. No ecotropic, xenotropic, or mink cell focus-forming (MCF) virus expression was found at the age of 5 wk, which is the time of LP-BM5 (MuLV) challenge. However, at 22 wk of age, one-half of the offspring from MAIDS mothers had ecotropic virus-expressing cells in their spleens. At the time of suckling, offspring from infected mothers had enhanced percentages of B cells and CD4 and CD8 T cells in the spleen, possibly followed by a slight persistent splenomegaly. These results suggest that immune reactivity, rather than tolerance to the virus, is responsible for resistance to disease after challenge. The offspring of MAIDS mice could clear the virus after challenge. This clearance was mediated by CD8 T cells, as continuous CD8 T cell depletion initiated at the time of viral challenge abrogated the resistance of these mice to MAIDS.

Animals↗
PubMed1994

Murine AIDS protects mice against experimental cerebral malaria: down-regulation by interleukin 10 of a T-helper type 1 CD4+ cell-mediated pathology.

The retrovirus LP-BM5 murine leukemia virus induces murine AIDS in C57BL/6 mice that has many similarities with human AIDS; Plasmodium berghei ANKA causes experimental cerebral malaria in the same strain of mice. The outcome of malaria infection was studied in mice concurrently infected with the two pathogens. The retrovirus significantly reduced the gravity of the neurological manifestations associated with Plasmodium berghei ANKA infection. The protection against experimental cerebral malaria induced by murine AIDS increased with duration of viral infection and, hence, with the severity of the immunodeficiency. Interleukin 10, principally from splenic T cells, was shown to play a crucial role in this protection.

Animals↗
PubMed1994

The significance of the pre-challenge immune status of mice for development of retrovirus-induced immunodeficiency syndrome (MAIDS).

The effects of vaccination with RNA-free viral pseudoparticles, preinfection with non-pathogenic ecotropic virus, and induction of tolerance to viral proteins in newborns on the outcome of murine immunodeficiency syndrome (MAIDS) were studied. The parameters used to follow disease progression were: lymphopenia, circulating B and T8 cells, serum IgG and IgM levels, lymphoproliferation and skin graft rejection. Immunization with RNA-free viral pseudoparticles had no effect on any of these parameters. Preinfection of adults with ecotropic virus and the induction of tolerance in newborns to virus antigens both attenuated the early symptoms of viral infection and delayed the onset of immunodeficiency and lymphoproliferation in some mice, but did not significantly alter the number of deaths due to MAIDS. Failure of immune-based therapy to produce successful protection against MAIDS suggests that immune destruction caused by the persistent virus rather than hyperimmune activity is the main pathogenic factor in this disease.

Animals↗
PubMed1993

Epidermal calcium-binding protein: a marker of early differentiation of basal layer keratinocytes of rats.

Epidermal calcium-binding protein (ECaBP) is present in the cells of the basal layer of the epidermis and other stratified epithelia. Since the basal layer compartment contains at least two types of cells: slow-cycling, poorly-differentiated, and actively proliferating, more differentiated cells, it was of interest to determine whether they both contained ECaBP. Basal and nearly suprabasal layer keratinocytes from newborn rat epidermis were fractionated into three fractions on the basis of cell size, using low-gravity sedimentation. The cell differentiation in each subgroup was estimated by cell size, morphology, cell cycle stage, RNA/DNA content, and the presence of specific keratins. The presence of ECaBP in these fractions was detected by immunocytochemistry and immunoblotting. Double staining with ECaBP antibodies and propidium iodide followed by flow cytometry was used to correlate ECaBP production and the stage of cell cycle. The relative cell size, measured by the light scattering was used to study the relationship between cell size and ECaBP production. The results show that small keratinocytes with low DNA and RNA content (G0 cells) do not express ECaBP. ECaBP was found only in intermediate size basal keratinocytes with higher DNA and RNA contents, corresponding to actively proliferating S phase cells. Large keratinocytes, which express suprabasal keratin and have low DNA and high RNA content, cease to express ECaBP. ECaBP may, therefore, be a useful marker for assessing the movement of cells from poorly differentiated reserve compartment towards proliferation and further differentiation in both physiological and pathological situations.

Animals↗
PubMed1991

Characteristics of homogeneously small keratinocytes from newborn rat skin: possible epidermal stem cells.

The aims of the present study were to characterize the phenotype, growth kinetics, and proliferative activation in culture of a population of poorly differentiated homogeneously small (HS) keratinocytes. These slow-cycling cells were separated by unit gravity sedimentation from a population of actively proliferating basal keratinocytes in newborn rat skin. This population (approximately 1% of the total basal keratinocytes) consisted of extremely small cells with little cytoplasm or RNA. Their positive KL4 staining demonstrates that they were keratinocytes. HS keratinocytes did not, however, contain epidermal calcium binding protein. Acridine orange, bivariate Hoechst, and ethidium bromide flow cytometry of in vitro bromodeoxyuridine-labeled cells as well as Ki67 staining showed that HS keratinocytes were in the G0 stage of the cell cycle and did not actively proliferate in vivo. [3H]thymidine label-retaining cells were found only in the HS cell population, showing that HS cells may originate from a central position in the epidermal proliferative unit. Growth of HS cells in vitro was characterized by a delayed but progressive increase in RNA before entry into the cell cycle. The clonogenic efficiency of HS cells in primary culture was much less than that of larger cells. Subclones of HS cell colonies exceeded primary colonies in their cloning efficiency and proliferative potential, suggesting that HS cells, although normally prevented from dividing, retain a high self-renewal capacity. They also maintain the ability to differentiate. The results are consistent with the concept that HS cell population may represent the epidermal-specific progenitor cells which act as stem cells in this tissue.

Animals↗
PubMed1990

Cell size and RNA content correlate with cell differentiation and proliferative capacity of rat keratinocytes.

Keratinocytes from rat skin were separated according to their size in a specially designed unit-gravity sedimentation chamber. The fractions obtained with this technique showed clear morphological differences, and analysis of size distribution confirmed that size was the criterion for separation. Simultaneous DNA and RNA staining of the fractions with acridine orange and subsequent flow cytometric analysis enabled one to classify cells into resting, proliferating, and differentiating stages. Cell size was not directly correlated with proliferation in situ as determined with acridine orange flow cytometry, nor with proliferative capacity in culture as assayed by BrdU/Hoechst flow cytometry. The smallest cells, exhibiting low DNA and RNA content, which do not proliferate in vivo, required a prolonged period of serum stimulation in vitro to initiate RNA and DNA synthesis. Cells of intermediate size exhibited early RNA synthesis and maximal proliferative capacity, whereas the largest cell population displayed no RNA synthesis in culture and the least proliferative capacity. In conclusion, these results suggest that RNA synthesis early after serum stimulation, in addition to a specific, optimal cell size, correlates with the proliferative capacity of keratinocytes in cell culture.

Animals↗
PubMed1989

Expression of alpha and beta tropomyosin subunits during early myogenesis in somites and limb buds of chick embryos.

The appearance of alpha and beta subunits of skeletal tropomyosin in early myogenesis was studied histochemically using monoclonal antibody to alpha tropomyosin and affinity-purified polyclonal antibody to beta tropomyosin. In muscle cells, in both somites and limb buds, the alpha and beta subunits are simultaneously expressed and first appear in the somites at the 30-36 somites. The relatively greater amount of beta than alpha tropomyosin found in early myogenesis is thus likely to result from a higher rate of beta tropomyosin synthesis.

Animals↗
PubMed1989

Cell subpopulations within proliferative and differentiating compartments of epidermis.

The heterogeneous population of newborn rat keratinocytes was separated into different subgroups according to their cell size. The relation between cell size, position in the cell cycle, RNA content, and proliferative potential in culture was examined. A reserve stem cell population of Go/G1 cells, low in RNA, giving rise to colonies of undifferentiated phenotype in cell culture, has been separated from more differentiated transit basal cells. In the fractions of the larger cells, several subgroups, probably corresponding to different stages of differentiation, were identified: G2M cells with low RNA content, large S-phase cells rich in RNA, and small Go/G1 cells low in RNA. The clonogenic cells from these fractions have limited growth potential and give rise to moderately or terminally differentiated colonies. The selective sorting of stem cell populations may be useful for elucidating the mechanism of carcinogenesis in epidermis and other proliferative tissues. Analysis of the relative proportions of cell subpopulations represents a novel approach leading to the refinement of the concepts of epidermal structure in physiological and pathological states. It also could, by extension, shed new light on the behavior of other proliferative tissues.

Animals↗
PubMed1988

Effect of vitamin D deficiency and 1,25-dihydroxycholecalciferol treatment on epidermal calcium-binding protein (ECaBP) RNA activity.

We have previously reported that the amount of epidermal calcium binding protein (ECaBP) in the skin decreases in the absence of vitamin D. Since vitamin D influences epidermal differentiation, and the synthesis of ECaBP may vary with cell differentiation, it was necessary to know whether vitamin D acts directly on the translational or post-translational level of ECaBP synthesis or indirectly by its action on epidermopoiesis. The cell-free translation technique was used to demonstrate the presence of mRNA coding for ECaBP. The activity of this mRNA has been evaluated in the skin of vitamin D-fed and in vitamin D-deficient rats with or without treatment with 1,25-dihydroxycholecalciferol (1,25(OH)2D3). Vitamin D deficiency decreased the ECaBP mRNA activity. The latter was selectively increased in animals given a single dose of 1,25(OH)2D3. These results suggest that 1,25(OH)2D3 stimulates the production of ECaBP mRNA or stabilizes this mRNA.

Animals↗
PubMed1987

Calcium-binding proteins in rat skin.

Skin Ca2+-binding protein (SCaBP) was reported to be distinct from the Ca2+-binding parvalbumin (PV), however, more recently its amino acid sequence was shown to be identical to PV. We purified a protein (Mr 12,000; pI4.5) from isolated epidermis (free of other cell layers) of adult rats and whole skin (containing no PV) of newborn rats. This protein is referred to as epidermal protein (EP-12), distinct from PV in its hydrophobicity, amino acid composition and immunological properties. Previously isolated SCaBP was shown to be a mixture of EP-12 and PV. The localization and possible functions of EP-12 and of PV in skin of adult and newborn rat are discussed.

Amino Acids↗
PubMed1987

Cell free translation of rat epidermal calcium-binding protein (EP-12) messenger RNA.

The primary step in the biosynthesis of 12 KDa rat epidermal calcium binding protein was studied by cell-free protein synthesis. Poly(A)+ rich RNA was extracted and purified from whole newborn rat skin and translated in a lysate system in the presence of labeled methionine. Immunoprecipitation of translation products with a monospecific antibody directed against this protein, which did not react with parvalbumin yielded a product migrating as a single band of molecular weight 12 KDa on polyacrylamide gel electrophoresis. Thus, a mRNA coding for this protein is present in rat skin. The presence of this messenger RNA opens the way for further studies on the regulation of epidermal expression during epidermal cell proliferation and differentiation.

Animals↗
PubMed1986

Skin calcium binding protein: effect of ultraviolet B irradiation.

Skin calcium binding protein (SCaBP) has been found in the epidermal basal layer but not in the suprabasal layers, suggesting that its presence is correlated with the position of the cell and/or the cells' low degree of differentiation and high proliferation capacity. Rats were exposed to ultraviolet B (UVB) irradiation (280-320 nm, 0.1 J/cm2) to dissociate these two main characteristics of basal layer cells. Skin biopsy specimens were taken 24 and 48 h after UVB exposure. The [3H]thymidine labeling index, SCaBP localization (indirect immunofluorescence), and SCaBP levels monitored by radioimmunoassay were investigated. The [3H]thymidine labeling index was significantly increased after UVB treatment, and the [3H]thymidine-labeled cells were present not only in the basal layer but also in the suprabasal epidermal layers. Anti-SCaBP immunofluorescence was observed in both suprabasal and basal layers (rather than exclusively in the basal layer, as in control conditions). The radioimmunoassay indicated an overall increase in skin SCaBP content. No evidence was obtained that this increase was due to humoral factors, as no changes in SCaBP concentration on cell proliferation were observed in nonirradiated epidermal areas. Topical application of a single dose of vitamin D3, equivalent to the amount synthesized by UVB exposure, was also without effect. Thus the presence of this marker is correlated with the low degree of cell maturity and the cells' ability to proliferate rather than their basal position.

Animals↗
PubMed1984

Skin calcium-binding protein: effect of vitamin D deficiency and vitamin D treatment.

The amount of skin calcium-binding protein, evaluated using a sensitive radioimmunoassay and indirect immunofluorescence, was decreased in vitamin-D deficient rats and increased after one week vitamin D3 or 1 alpha-hydroxyvitamin D3 treatment. In vitamin D replete and in vitamin D-deficient animals, skin calcium-binding protein was not sensitive to changes in dietary and/or serum calcium concentrations. These results indicate that this protein is different from other calcium-binding proteins such as parvalbumin and calmodulin which are not vitamin D-dependent, and also different from intestinal calcium-binding protein which, in D replete animals, is sensitive to changes in dietary and serum calcium concentrations. Skin calcium-binding protein may, therefore, represent a new class of vitamin D-dependent protein.

Animals↗
PubMed1984

Alterations in rat epidermis provoked by chronic vitamin D deficiency.

Long-term effects of vitamin D deficiency on epidermis were studied using histometric techniques, [3H]thymidine incorporation into DNA (labeling index), estimation of epidermal acid phosphatase activity, and one-dimensional gel electrophoresis of keratin proteins. The decrease in epidermal thickness due to a reduced number of granular cell layers and a lower level of epidermal acid phosphatase activity were observed in vitamin D-deficient rats. The number of nuclei in the basal layer was increased. No changes in labeled index due to chronic vitamin D deficiency or to 'single injection of 1,25-dihydroxycholecalciferol to vitamin D-deficient rats were observed. A comparative study of the keratin composition revealed differences in the keratin polypeptide pattern: vitamin D-deficient epidermis specifically lacked two low-molecular-weight components and presented several quantitative differences among other keratin polypeptides. The changes in epidermal morphology and metabolism that took place with vitamin D deficiency were independent of plasma calcium levels because similar modifications were present in vitamin D-deficient but normocalcemic rats (fed a diet rich in calcium and supplemented with lactose). These findings suggest that vitamin D may be one of the important factors for maintaining normal epidermal structure and metabolism through an effect on cell differentiation and formation of granular cell layers. They offer the possibility of using epidermal modification as an additional marker of vitamin D deficiency.

Acid Phosphatase↗
PubMed1984

Skin calcium-binding protein in squamous metaplasia of human uterine cervix.

The distribution of skin calcium-binding protein in squamous cell metaplasia of human endocervix, in normal human skin, and in ovarian cancer was determined by the immunofluorescence technique. A rabbit antiserum specific to rat SCaBP was characterized by Ouchterlony immunodiffusion and by immunoprecipitation of 125I-labeled SCaBP. The specificity of antibody labeling was demonstrated by using preimmune rabbit serum and SCaBP antiserum competitively absorbed with purified SCaBP. In normal human skin SCaBP was found exclusively in the basal layer cell cytoplasm. This protein was not detected in normal columnar epithelium of endocervix. Epithelial tissues in the zone of transition between the cylindrical epithelium of the endocervical mucosa and the stratified squamous epithelium of the exocervix were obtained from 14 patients with a wide variety of squamous cell metaplasia. In the early stage of metaplasia SCaBP was detected exclusively in the cytoplasm of reserve undifferentiated cells. In the terminal stage of metaplasia the SCaBP was present only in the basal cell layer. SCaBP was found in several layers of dysplastic tissue, and this distribution appeared to be related to the loss of normal maturation of the epithelium. SCaBP was also present in squamous cell carcinoma of endocervix especially in the least differentiated regions of the tumor. No SCaBP was detected in any ovarian cancer cells. These findings are potentially useful as a means of early detection of squamous metaplasia and of distinguishing premalignant anaplastic lesions from those that are benign and nonproliferative. In addition, the presence of SCaBP in tumors derived from metaplastic epithelia and its absence in the ovarian cancer indicate that immunohistochemical search for this protein might be of value in tumors in which an epidermoid origin is a possibility.

Animals↗