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J Varani

Publications and source records attributed to J Varani.

At least 199 records · Page 11Linked to original sources
PubMed1984

Demonstration of C5 cleaving activity in bronchoalveolar fluids and cells: a mechanism of acute and chronic alveolitis.

Recently, we have demonstrated that preformed chemotactic fragments derived from C5 (C5fr), when instilled intratracheally, induce an intense acute alveolitis. An intense intrapulmonary inflammatory reaction, characterized by accumulation of large numbers of neutrophils in the alveolar spaces, also resulted from intratracheal instillation of purified human C5. Since purified C5 does not induce PMN chemotaxis in vitro and presumably native C5 is not phlogistic in vivo, a mechanism for C5 cleavage productive of chemotactic fragments may be operative within the lung. To explore the possibility that pulmonary-mediated fragmentation of C5 occurred, both in vitro and in vivo studies were undertaken. As described in this paper, extensive cleavage of 125I-labeled C5 was demonstrated in the bronchoalveolar lavage fluids (BALF) of hamster lungs previously instilled with 125I-labeled C5. In vitro incubation of 125I-labeled C5 with normal hamster lung lavage components, cells, and lavage fluid also cleaved 125I-labeled C5. Bioassay of such in vitro reactions also revealed potent chemotactic activity for polymorphonuclear leukocytes. A potent C5-cleaving activity was detected in fluid of normal bronchoalveolar lavage (BAL) recovered from normal hamster lungs. This BALF "enzyme activity" selectively cleaved C5 in vitro, producing potent C5-related chemotactic fragmentation products, but it did not fragment C3 nor did it produce C3 chemotactic factors in vitro. Cleavage of C5 by alveolar macrophages that produces chemotactic factors has previously been attributed to an acid protease of lysosomal origin. This "enzyme activity" is apparently distinct from elastase, collagenase, and trypsin since it is not inhibited by alpha 1-antitrypsin. In separate studies, we have recently demonstrated the ability of isolated lung cells to synthesize and secrete C3 and C5 in vitro. Thus, these studies suggest that the lung may intrinsically have the capacity to regulate inflammatory reactions by the localized production of both complement components (C3 and C5) as well as complement-activating systems (e.g., C5-cleaving activity of BALF).

Acute Disease↗
PubMed1984

Phorbol ester binding to chemotactically responding and nonresponding Walker 256 carcinosarcoma cells.

Phorbol esters induce, in the chemotactically responsive Walker 256 carcinosarcoma cells, functional responses that are similar to those induced by peptide chemotactic factors. These responses are presumed to result from ligand binding to cellular receptors, although this has not been formally demonstrated. In this study, it was shown that tritium-labeled phorbol dibutyrate [( 3H]PDB) bound to the Walker cells in a time- and concentration-dependent manner. Binding was inhibited by excess unlabeled PDB and was reversible. Half-maximal binding was achieved with a 31 nM concentration of [3H]PDB and occurred within 15 min after addition of the ligand. This is in accord with biological activity (i.e., cell-to-substrate adherence). Half-maximal cell adherence was observed with 25 nM PDB. Increased adhesiveness was detected as early as 15 min after exposure of the cells to the ligand. The response peaked at 30 to 45 min after exposure and fell off rapidly thereafter. A number of phorbol esters successfully competed with [3H]PDB binding to the cells. There was a direct relationship between the ability of these agents to compete for binding and their ability to induce biological activity. Using cell-to-substrate adherence as an indicator of biological activity allowed separation of responding and nonresponding populations. The biologically responsive cells and the nonresponsive cells both bound high levels of [3H]PDB. This suggests that receptor occupancy is, by itself, not sufficient for biological activity and that, in Walker cells, one or more points of control exist subsequent to ligand binding.

Animals↗
PubMed1983

Enzyme-linked lectin assay (ELLA): use of alkaline phosphatase-conjugated Griffonia simplicifolia B4 isolectin for the detection of alpha-D-galactopyranosyl end groups.

Alkaline phosphatase has been coupled to Griffonia simplicifolia I B4 isolectin using a one-step glutaraldehyde conjugation procedure. This enzyme-lectin conjugate (AP-GS I-B4) has been used to specifically detect plastic-bound natural and synthetic glycoproteins bearing alpha-D-galactopyranosyl end groups. The extent of reactivity of the AP-GS I-B4 with the glycoproteins appears to be proportional to the number of terminal galactosyl residues present. Furthermore, this assay, termed ELLA (enzyme-linked lectin assay), is specifically inhibitable by low-molecular-weight sugars containing terminal alpha-D-galactosyl groups. The ELLA reactions may be assayed rapidly and objectively by the use of commercially available ELISA-plate using standard filters.

Alkaline Phosphatase↗
PubMed1983

Suppressor cells and increased primary tumor growth rate induced by thiopental.

A single dose of thiopental (37-42 mg/kg) sufficient to achieve anesthesia induction increased the growth rate of a 3-methylcholanthrene-induced syngeneic murine fibrosarcoma in C57B1/6 mice. While no alterations in in vitro growth kinetics of tumor cells cultured with thiopental could explain these data, significant alterations in cell-mediated immunity were observed. Spleen cells from C57B1/6 mice treated with thiopental were impaired in their ability to respond in a mixed leukocyte culture (MLC) to BALB/c stimulator cells. Spleen cells from thiopental-treated mice suppressed the ability of cells from untreated animals to respond in a third-party MLC. The degree of suppression was directly proportional to the number of cells from thiopental-treated mice in the third party. When these cells were added to an MLC at various times during the culture period, suppression of the early phases of the MLC was seen. Removal of plastic adherent cells from the third party resulted in loss of suppression, but treatment of third-party cells with anti-thy 1.2 serum or irradiation did not prevent suppression. Transfer of spleen cells from thiopental-treated mice enhances growth of the B-16 melanoma. Suppressor-cell activity may be one mechanism by which thiopental promotes tumor growth.

Animals↗
PubMed1983

Modulation of tumor cell adherence by prostaglandins.

The initiation of tumor cell metastasis is thought to involve the adherence of circulating tumor cells to endothelial cells and/or basement membranes. The adherence of Walker carcinosarcoma tumor cells to foreign surfaces induced by phorbol myristate acetate and the chemotactic formyl peptide, formylmethionyl-leucyl-phenylalanine is inhibited by prostacyclin and 15-S-15-methyl PGE1 in a dose-dependent manner. The inhibition of tumor cell adherence by prostacyclin and 15-S-15-methyl PGE1 is associated with a 3- to 5-fold increase in intracellular cyclic adenosine monophosphate (cAMP) concentrations. The data suggests that the local production of prostaglandins may be an important modulator of tumor cell metastasis due to its direct effect on the tumor cells.

Animals↗
PubMed1983

Hyperimmune human ABO blood-typing sera: reactivity with murine laminin and cytotoxicity for metastatic murine tumour cells.

Commercially prepared ABO blood-typing antisera have been tested for their ability to bind to murine laminin and their cytotoxic effects upon high and low metastatic variants of a murine fibrosarcoma. Previous studies have shown that alpha-D-galactopyranosyl end-groups comprise the major antigenic determinants on type B erythrocytes and that these same end-groups are present on murine laminin purified from the EHS sarcoma and on a laminin-like glycoprotein on the surface of the high, but not low, metastatic fibrosarcoma cells. In the present study we found that all sera containing anti-B activity were cytotoxic to the high, but not the low, metastatic cells and that all of these sera reacted strongly against immobilized murine laminin in an enzyme-linked immunosorbent assay. Sera lacking anti-B activity, i.e. anti-A antisera, were much less cytotoxic to either cell line and three of the four anti-A sera did not bind to murine laminin. The laminin reactivity and cytotoxic effect of the anti-B sera were specifically abrogated by preincubation of the sera with water-soluble blood group B substance or with murine laminin but not with water-soluble blood group A substance.

ABO Blood-Group System↗
PubMed1983

Differential expression of a lamininlike substance by high- and low-metastatic tumor cells.

High-metastatic murine fibrosarcoma cells readily attached to Type IV (basement membrane) collagen, whereas low-metastatic cells isolated from the same tumor did not. The addition of laminin--a glycoprotein that facilitates the adherence of epithelial cells to their basement membranes--enhanced the attachment of the low-metastatic cells, but not the high-metastatic cells. Using anti-laminin antibodies and a laminin-binding lectin as probes, the authors were able to identify by immunofluorescence a moiety associated with the high-metastatic cells, but not the low-metastatic cells, which cross-reacted with murine laminin purified from the EHS sarcoma. When extracts from the high-metastatic cells were separated by affinity chromatography, with the laminin-binding lectin as the affinity substrate, a substance was isolated that had an apparent molecular weight of 56,000 daltons. The affinity-purified material reacted strongly with anti-laminin antibodies by enzyme-linked immunosorbent assay.

Animals↗
PubMed1983

Cell surface alpha-D-galactopyranosyl end groups: use as markers in the isolation of murine tumor cell lines with different cancer-causing potentials.

Cell lines from 2 3-methylcholanthrene-induced murine tumors were established in culture and examined for reactivity with Griffonia simplicifolia isolectin B4 (GS I-B4), a lectin that has strict specificity for terminal alpha-D-galactopyranosyl residues. Virtually all of the cells in both populations were strongly reactive, indicating the presence of this carbohydrate on the cell surface. Both tumor lines were exposed to human serum with antibodies to the blood group B antigen. More than 99.99% of the exposed cells were killed by this treatment. This is not surprising, because terminal alpha-D-galactopyranosyl residues comprise the blood group B antigen. From the few surviving cells, it was possible to establish cell lines resistant to the cytotoxic effects of the anti-blood group B antibodies. A total of 4 cell lines were independently obtained in this way. The human serum-resistant lines showed no detectable reactivity with GS I-B4, indicating that alpha-D-galactopyranosyl-deficient cell lines had been obtained. The parent and variant cells were compared for reactivity with anti-laminin antibodies. Both parent lines showed strong reactivity by immunofluorescence in the viable state, whereas the alpha-D-galactopyranosyl-deficient lines showed no reactivity. The parent and variant lines were also compared with regard to in vitro and in vivo growth. The alpha-D-galactopyranosyl-deficient lines had reduced in vitro growth capacity relative to the parent lines. More importantly, in contrast to the parent lines, these lines were significantly less tumorigenic than the parent lines when injected into syngeneic mice and did not metastasize spontaneously.

ABO Blood-Group System↗
PubMed1983

Actin changes in normal human and rat leukocytes and in transformed human leukocytic cells.

The DNase I inhibition assay was used for the determination of the relative amounts of monomeric and total actin in normal human peripheral blood lymphocytes, normal Sprague-Dawley rat peritoneal leukocytes, and 6 transformed human cell lines of lymphoid and myeloid origins. In the normal lymphocytes and leukocytes, greater than 50% of the total actin was monomeric. In contrast, only 1 of the 6 transformed lines had greater than 50% of its actin in the monomeric form. In the other 5 lines, the percentage of actin in the monomeric form ranged from 23 to 39%. The normal lymphocytes, peritoneal leukocytes, and 2 of the transformed cell lines (CEM and K562) were examined in greater detail. The total actin (as a percentage of the total protein) was found to be much lower in the transformed cell lines than it was in the normal lymphocytes. The total amount of actin in the normal rat leukocytes was very similar to the amount in the normal human lymphocytes. In addition to these differences between the normal and transformed human cells, treatment of the normal lymphocytes with mitogenic doses of concanavalin A was found to significantly reduce the amount of monomeric actin in the cells. Similar treatment of the transformed cells produced no significant reduction in the monomeric actin.

Actins↗
PubMed1982

Chemotaxis of metastatic tumor cells.

A number of factors have been identified which are chemotactic for tumor cells. Recent studies have shown that, in addition to inducing directional motility in the Boyden chamber assay, these factors also induce a number of other responses. Included among these responses are cell swelling and foreign surface adhesiveness. The adherence response has been studied in detail using the Walker 256 carcinosarcoma cells and several other cell types. In the Walker cells, treatment with the C5a-derived tumor cell chemotactic peptide, the synthetic tripeptide, N-formyl-methionyl-leucyl-phenylalanine or with 12-O-tetradecanoyl phorbol ester induces a rapid, transient adherence response. The response is completely inhibited by several agents known to block the activity of phospholipase A2 or the metabolism of arachidonic acid through the lipoxygenase pathway but is not inhibited by inhibition of the cyclooxygenase pathway. This suggests that lipoxygenase metabolites of arachidonic acid may actually mediate the adherence response. It has been shown that chemotactic factor treatment of animals that are bearing circulating tumor cells induces a localization of these cells at the site of chemotactic factor injection. On the basis of these observations it has been hypothesized that tumor cells respond to chemotactic factors in much the same way that leukocytes do and that tumor cell localization at metastatic sites in vivo may be influenced by chemotactic factors in much the same way that leukocyte localization at inflammatory sites is.

Animals↗
PubMed1982

Nonspecific protease and elastase activities in rat leukocytes.

Extracts were prepared from rat peritoneal leukocytes obtained 4 h after glycogen injection and assayed for proteolytic enzyme activities against various substrates. The substrates used included acid-denatured bovine hemoglobin, bovine serum albumin, a partially purified preparation of rat pulmonary basement membrane, bovine neck ligament elastin, and an artificial substrate with elastase specificity. A high level of activity was observed when hemoglobin was used as the substrate. The serum albumin and basement membrane preparation were also readily hydrolyzed by the leukocyte extract. In contrast, the native elastin and synthetic elastase substrate were much more resistant. Although the leukocyte extract demonstrated little intrinsic elastase activity, when it was mixed with a commercial hog pancreatic elastase preparation, it greatly potentiated the elastolytic activity, suggesting the activation of a latent enzyme.

Animals↗
PubMed1982

CAMP differences between clones of high and low malignant fibrosarcoma cells.

The levels of 3',5'-cyclic adenosine monophosphate (cAMP) were measured in a high malignant and a low malignant clone of murine fibrosarcoma cells isolated from a common parent in a normal mouse fibroblasts at various stages of growth. There was an inverse correlation between the degree of malignant potential and the cAMP levels. The normal fibroblasts had the highest cAMP at all time periods measured. The high malignant cells had the lowest levels and the low malignant cells were intermediary. The degree of susceptibility of each population to the growth inhibiting effects of dibutyryl cAMP, a cAMP analogue, was also measured. There was a direct correlation between degree of malignant potential and susceptibility to growth inhibition. These findings strongly support our belief that the low malignant cells represent a state of malignancy that is intermediary between the high malignant cells and the normal fibroblasts. cAMP differences may contribute to this.

Animals↗
PubMed1982

Actin polymerization induced by chemotactic peptide and concanavalin A in rat neutrophils.

Changes in the state of actin in rat neutrophils were studied after chemotactic peptide and concanavalin A stimulation by using the DNase I inhibition assay. Actin polymerization occurred within seconds after stimulation with F-Met-Leu-Phe and concanavalin A. Pretreatment of cells with cytochalasin D prevented chemotactic peptide-induced actin polymerization. The addition of F-Met-Leu-Phe to lysed cells did not produce any change in actin state. These data offer strong evidence for receptor-induced actin polymerization and support the models implicating actin microfilament formation as a crucial event in cell activation. The observations on platelets, lymphocytes, neutrophils, and islets of Langerhans from different species suggest that actin polymerization might be a universal intracellular event accompanying cell surface receptor perturbation in eukaryotic cells.

Actins↗
PubMed1982

Phenotypic stability of murine tumor cells in vitro and in vivo.

Murine fibrosarcoma cells that vary in their transplantability in syngeneic mice were isolated from a heterogeneous parent tumor (induced in a female C57BL/6 inbred mouse) and maintained in culture by serial passage. Several biologic properties that discriminate between the high- and low-transplantable lines (including adhesiveness, motility, and levels of chymotrypsin-like esterase activity), as well as properties that do not separate these lines (e.g., in vitro growth rates and levels of protease and glycosidase activities), were measured at periodic intervals over 2 years. Ability to induce primary tumors and to induce metastases from these tumors were evaluated at the same intervals. The high- and low-transplantable lines were also transplanted into syngeneic mice at tumorigenic doses. Isolates from primary and metastatic tumors induced in the animals were reestablished in culture and examined for the various characteristics of passages 5, 10, 20, and 30. The properties of the cells maintained continually in culture remained stable throughout the 2-year observation period. Tumor isolates showed some evidence of modulation immediately after reestablishment in culture, but by passage 10 they appeared to be identical to the prototype parent lines. These data show that the fibrosarcoma cells do not undergo continual phenotypic "drift" as has been suggested to occur with other tumor lines maintained by serial passage in animals or in culture.

Animals↗
PubMed1982

Phorbol myristate acetate-induced adherence of Walker 256 carcinosarcoma cells.

Treatment of nonadherent Walker 256 carcinosarcoma cells with phorbol myristate acetate (PMA) causes these cells to become adherent to noncellular foreign surfaces such as nylon fibers and plastic culture dishes and to monolayers of endothelial cells. Increased adherence is first observed after a short lag period (5 to 15 min) and is transient. Other tumor-promoting analogs of PMA also induce this response, while inactive analogs of PMA do not. Simultaneous treatment of the cells with 2-deoxyglucose, colchicine, cytochalasin B, and cycloheximide indicates that the adherence response of the cells is an energy-dependent process that requires an intact cytoskeleton but does not require protein synthesis. Inhibitors of phospholipids and arachidonic acid metabolism including indomethacin, nordihydroguaiaretic acid, and p-bromophenacyl bromide greatly inhibit PMA-induced adherence, but acetylsalicylic acid is much less effective. PMA also increases the rate of attachment to plastic dishes of cells which would normally attach, although slowly, and grow as substrate-attached cells. However, PMA treatment has no effect on the subsequent degree of susceptibility of these cells to release from plastic dishes mediated by proteolytic enzymes. These findings suggest (a) that PMA may be useful in delineating the initial events involved in the adherence of cells to cellular and noncellular surfaces and (b) that PMA may stimulate tumor cell adherence in a manner similar to that of chemotactic peptides may be useful in delineating the events associated with chemotactic factor stimulation of these cells.

Animals↗
PubMed1981

A model of tumor cell dormancy: effects of anesthesia and surgery.

Three cloned populations of tumor cells obtained from a murine fibrosarcoma were able to remain viable for a long period of time in syngeneic mice which failed to exhibit clinically evident tumors following tumor cell inoculation. Viable tumor cells under such conditions can be considered to be in a dormant state. On the basis of past studies, two of the lines were shown to have low malignant potential, while the third line was shown to have a higher degree of malignant potential. The rates of spontaneous reactivation of tumor growth in animals carrying the low malignant cells were 3 and 4%, while a rate of 30% was observed in animals with the more malignant cells. Treatment of animals carrying the low malignant cells in a dormant state with anesthesia alone, (thiopental) or with anethesia and surgery, increased the rate of reactivation to 20-22%.

Anesthesia↗