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Biomedical subjects

S H Barsky

Publications and source records attributed to S H Barsky.

At least 19 recordsLinked to original sources
PubMed1992

A study of the heterogeneity of the mucoepidermoid tumor and the implication for future therapies.

Mucoepidermoid carcinomas are common head and neck neoplasms whose wide distribution includes the major salivary glands, especially parotid, the minor salivary glands within the oropharynx, sites within the larynx, and the submucosal bronchial glands. These tumors are heterogeneous neoplasms composed of intermediate, squamous, and mucous cell subpopulations and have been traditionally classified as low or high grade based on their histological nature and tendency for invasion and metastasis. To gain a better understanding of the heterogeneity of these tumors, we conducted in-depth studies of the different cellular subpopulations obtained from primary human mucoepidermoid carcinomas from various sites, and a derived cell line grown in monolayer culture and as a nude mouse xenograft. Flow cytometric and clonal dilution studies indicate that the intermediate cell behaves as a reserve cell capable of differentiating into either the squamous cell or mucous cell. Immunocytochemical studies of bromodeoxyuridine uptake and Ki-67 antigen expression indicate that whereas the squamous and mucous cells are terminally differentiated and permanently arrested in G0, the intermediate cell actively enters G1 and S. Studies of invasion utilizing a human amnionic membrane indicate that only the intermediate cell is capable of active invasion. Because the mucous and squamous cells are incapable of further division or invasion, therapeutic strategies aimed at differentiating the intermediate cell in vivo into either mucous or squamous directions would serve as useful adjuncts to surgery by reducing the rate of recurrence and the incidence of metastasis.

Animals↗
PubMed1992

Intratubular germ cell neoplasia in infantile yolk sac tumor. Verification by tandem repeat sequence in situ hybridization.

The strong association of intratubular germ cell neoplasia (ITGCN) with adult germ cell testicular tumors is well known, but studies noting the absence of ITGCN in certain germ cell neoplasms such as spermatocytic seminoma, childhood teratoma, and infantile yolk sac tumor (YST) have raised the issue of whether these latter neoplasms follow a different path of tumorigenesis, accounting for their more benign behavior. A case study illustrating the association of ITGCN with infantile YST is presented to challenge this hypothesis. In addition to the usual characteristic features that included strong cytoplasmic glycogen deposits, and focal placental alkaline phosphatase immunoreactivity, the atypical intratubular germ cells manifested triploidy by in situ hybridization using as probe a telomeric tandem repeat sequence, p1-79, specific to chromosome 1. The invasive YST cells, in contrast, showed evidence of tetraploidy by both in situ hybridization and flow and image cytometric studies, excluding the possibility that the atypical intratubular germ cells represented intratubular invasion by adjacent YST. These findings challenge the belief that the infantile YST follows a different path of tumorigenesis than its adult germ cell counterpart and suggest other hypotheses that might better explain its more benign behavior.

Blotting, Southern↗
PubMed1988

Two human tumors with high basement-membrane-producing potential.

Two human tumors, an adenoid cystic carcinoma and a yolk sac tumor, were found by immunocytochemical, ultrastructural, and biochemical studies to contain abundant basement membrane matrix in contrast to the vast majority of human tumors which contained either an absent or scant basement membrane matrix. These tumors were established as xenografts in athymic (nude) mice. Both xenografts maintained characteristic histologic features, immunocytochemical localization of basement membrane components, and reasonable yields of native laminin and Type IV collagen throughout three successive transplant generations. Although only a small fraction of the yield of that of the murine Engelbreth-Holm, Swarm (EHS) sarcoma, the yield of the human basement membrane-producing tumors could be increased by rendering the mice lathyritic. The human basement membrane proteins so extracted were identical on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) to their murine counterparts. These human tumors then represent a potential source of human basement membrane proteins.

Animals↗
PubMed1988

High metalloproteinase inhibitor content of human cirrhosis and its possible conference of metastasis resistance.

Human cirrhotic livers exhibited a strong resistance to metastasis and demonstrated high levels of both soluble and 3 M NaCl extractable metalloproteinase inhibitor(s) directed against tumor type I and type IV collagenases. This inhibitory activity was detected in human cirrhosis from diverse causes, including alpha-1-antitrypsin deficiency, alcoholic liver disease, postviral hepatitis, and biliary cirrhosis, but was nearly undetectable in normal liver. The inhibitory activity was able to be purified by carboxymethyl cellulose chromatography and gel filtration, had a molecular weight of approximately 40,000, and was relatively heat stable. Such metalloproteinase inhibitory activity was also present in conditioned media of human myofibroblast primary cultures obtained from cirrhotic liver explants but absent in conditioned media of primary normal liver explants. Purified fractions of the metalloproteinase inhibitor activity obtained from both myofibroblast conditioned media and extracts of cirrhotic liver blocked invasion but not attachment of MCF-7 cells on human amnion in vitro. The results further support the role of myofibroblasts in inhibiting tumor invasion and metastasis and provide one possible mechanism by which cirrhotic livers resist metastasis.

Enzyme Inhibitors↗
PubMed1988

Tumor promoter-induced membrane-bound protein kinase C regulates hematogenous metastasis.

A strong correlation was found between the basal levels of membrane-bound protein kinase C and the ability of B16 melanoma cell sublines (F10, F1, and BL6) to metastasize to the lung after intravenous injection. By treating with tumor-promoting phorbol esters for 1 hr, the low-metastasizing F1 cells exhibited both translocation of protein kinase C from cytosol to plasma membrane and an increase in metastasis to a level comparable to the (untreated) highly metastatic subline F10. Prolonged treatment of melanoma sublines with phorbol 12-myristate 13-acetate for 24 hr resulted in both inactivation of protein kinase C activity and loss of their metastasizing capabilities. Under conditions that induced only the activation of protein kinase C but not its membrane association, no increase in metastasis occurred, suggesting that activation of protein kinase C alone is insufficient to promote metastasis and that its membrane association is also necessary. Exposure of B16 melanoma sublines to phorbol esters for 1 hr had (i) no effect on the growth and morphology of these cells in vivo and in vitro and (ii) a short-term effect (approximately equal to 5 hr) on membrane association of protein kinase C. Nonetheless, in this period, the membrane-bound protein kinase C, probably by influencing cell-surface and cell-attachment properties, increased the retention of circulating melanoma cells in the lung, which eventually led to an increased number of metastatic nodules. The membrane-bound protein kinase C activity also correlated with metastatic ability in rapidly growing cells, growth-arrested cells, and cells growing in a low-Ca2+ medium. The results strongly suggest that the membrane-bound protein kinase C influences hematogenous metastasis of tumor cells and show that tumor promoters like phorbol esters have an additional role in promoting hematogenous spread of cancer in the body.

Animals↗
PubMed1987

Isolation of a myofibroblast growth factor from human breast carcinoma cell lines.

Conditioned media of a series of well-established human breast carcinoma cell lines were screened for mitogenic activity on human myofibroblasts. Whereas all carcinoma lines derived from desmoplastic ductal breast carcinoma primaries exhibited moderate to high levels of mitogenic activity, the single line derived from a non-desmoplastic (medullary) carcinoma exhibited low activity. Levels of mitogenic activity were independent of estrogen receptor status and estrogen/antiestrogen treatment. Fractionation of the conditioned media revealed a cationic, hydrophobic mitogenic factor of M.W. 25,000. The factor did not stimulate the growth of endothelial or carcinoma cells nor the growth of NRK fibroblasts in soft agar.

Animals↗
PubMed1987

Increased invasion and spontaneous metastasis of BL6 melanoma with inhibition of the desmoplastic response in C57 BL/6 mice.

BL6 melanoma cells injected s.c. in 18-month C57 BL/6 mice elicit a markedly fibrotic response similar in myofibroblast and collagen composition to that characterizing the desmoplastic response of human breast carcinoma. This host response can be quantitated by measuring hydroxyproline (total collagen) and incorporation of i.p.-injected [14C]proline into collagenase-sensitive protein (new collagen synthesis). Inhibition (70%) of the desmoplastic response can be achieved by daily injections of L-3,4-dehydroproline. Inhibiting the response in this manner promotes local invasion of tumor and increases the incidence of spontaneous pulmonary metastasis. 10(5) BL6 melanoma cells produce tumor nodules with a mean diameter of 1.5 +/- 0.5 cm and mean collagen content of 36 +/- 15 mg/g wet tissue at 4 weeks and 10% incidence of pulmonary metastasis at 7 weeks. L-3,4-dehydroproline produces nodules with a mean diameter of 2.3 +/- 0.5 cm and mean collagen content of 12 +/- 2 mg/g with a 40% incidence of metastasis. L-3,4-dehydroproline exerts a selective effect on myofibroblast collagen synthesis in vitro and no effect on [3H]thymidine uptake, doubling time, and viability of BL6 cells and myofibroblasts. Furthermore, this drug exerts no effect on BL6 invasion and metastasis in 6-week C57 BL/6 mice, hosts which exhibit a negligible desmoplastic response.

Animals↗
PubMed1987

An experimental model for studying the desmoplastic response to tumor invasion.

BL6 melanoma cells injected subcutaneously in 18-month-old syngeneic C57BL/6 mice elicit a marked fibrotic response highly similar in myofibroblast composition and Type V collagen content to that characterizing the desmoplastic response of human carcinomas. This host response can be quantitated in vivo by measuring both hydroxyproline content (total collagen) and incorporation of intraperitoneally injected [14C] proline into collagenase-sensitive protein (new collagen synthesis). 70% inhibition of the response can be achieved with daily L-3,4-dehydroproline. The response can be similarly quantitated in vitro in explants of desmoplastic tumor tissue. The model allows for subsequent investigations of the effects of the desmoplastic response on tumor invasion and metastasis.

Animals↗
PubMed1987

Extracellular hyaline bodies are basement membrane accumulations.

Diverse hyaline bodies occurring in various diseases were investigated immunocytochemically with antibodies to type IV collagen and laminin. These hyaline bodies included those found predominantly extracellularly in adenoid cystic carcinomas, endodermal sinus tumors, Spitz nevi, lichen planus, diabetic glomerular nodular sclerosis, and odontogenic cysts, and those found predominantly intracellularly in alcoholic liver disease, Kaposi's sarcoma, and malignant fibrous histiocytoma. The hyaline bodies found in the first six diseases displayed intense immunoreactivity for the basement membrane components, type IV collagen, and laminin, whereas the hyaline bodies occurring in the latter three diseases were unreactive. These findings suggest that a common histogenesis of the extracellular hyaline body is the stimulatory effect of an altered growth state (hyperplasia, neoplasia, or endocrinopathy) on basement membrane synthesis in cells that normally produce a basement membrane.

Basement Membrane↗
PubMed1987

Desmoplastic basal cell carcinomas possess unique basement membrane-degrading properties.

Desmoplastic basal cell carcinomas (fibrosing or morphea types) were studied ultrastructurally, immunocytochemically, and biochemically for basement membrane-degrading activity and compared with the common varieties of basal cell (superficial and nodular-ulcerative types). Whereas the latter lesions demonstrated intact basement membranes as evidenced by extracellular laminin and type IV collagen immunoreactivity and the presence of an unusually thickened basal lamina, desmoplastic basal cell carcinomas showed large defects and absences in basal lamina and basement membrane immunoreactivity. Intense tumor cytoplasmic immunoreactivity for type IV collagenase was present in 13 of 15 cases of desmoplastic basal cell but absent in the superficial and nodular-ulcerative varieties. Whereas explant cultures of all the types of basal cell carcinoma studied gave rise to high levels of interstitial (type I) collagenase activity in conditioned media, only the desmoplastic variety exhibited high type IV collagenase activity. These findings suggest that the mechanisms by which the desmoplastic and the common varieties of basal cell carcinoma infiltrate host tissues may be fundamentally different.

Basement Membrane↗
PubMed1986

Factors influencing chelator-stable, detergent-extractable, phorbol diester-induced membrane association of protein kinase C. Differences between Ca2+-induced and phorbol ester-stabilized membrane bindings of protein kinase C.

One of the early events associated with the treatment of cells by tumor promotor phorbol esters is the tight association of protein kinase C to the plasma membrane. To better understand the factors that regulate this process, phorbol ester-induced membrane binding of protein kinase C was studied using homogenates, as well as isolated membranes and purified enzyme. Addition of 12-O-tetradecanoylphorbol 13-acetate (TPA) to the homogenates of parietal yolk sac cells and NIH 3T3 cells in the presence of Ca2+ resulted in plasma membrane binding of protein kinase C which subsequently remained bound to the membrane independent of Ca2+. Although protein kinase C was activated by TPA in the absence of Ca2+ and by diolein in the presence of Ca2+, both these agents when added to homogenates under these respective conditions had no effect on membrane association of protein kinase C. However, under these conditions relatively weak binding of protein kinase C was found if purified protein kinase C was used with isolated membranes. Binding studies using purified protein kinase C and washed membranes showed that the binding of the TPA-kinase complex to membranes required phospholipids and reached saturation at 0.1 unit (24 ng of protein kinase C)/mg of parietal yolk sac cell membrane protein. Phorbol ester treatment of cells in media with and without Ca2+ showed that the TPA-induced increase in membrane-associated protein kinase C was regulated by Ca2+ levels even in intact cells. TPA-stabilized membrane binding of protein kinase C differs in several aspects from the previously reported Ca2+-induced reversible binding. TPA-stabilized binding of protein kinase C to isolated membranes is temperature dependent, relatively high in the plasma membrane-enriched fraction, saturable at physiological levels of protein kinase C, requires the presence of both membrane protein(s) and phospholipids, and further requires the addition of phospholipid micelles. In contrast, Ca2+-induced reversible binding is more rapid, not appreciably influenced by temperature, not selective for a particular subcellular fraction, not saturable with physiological amounts of protein kinase C, exhibits trypsin-insensitive membrane binding sites, and requires membrane phospholipids but not added phospholipid micelles.

Animals↗
PubMed1986

Hydrophobic association of calpains with subcellular organelles. Compartmentalization of calpains and the endogenous inhibitor calpastatin in tissues.

Calpains I and II isolated from diverse tissues possess both Ca2+-independent, and Ca2+-dependent accessible hydrophobic regions. Possible subcellular organelle association of calpains involving these hydrophobic regions was studied. By homogenizing rat tissues directly in Ca2+ (50 microM), about 30-60% of the cytosolic calpain I and II activity reversibly associated with isolated subcellular fractions (microsomal greater than plasma membrane greater than nuclear). After binding to the particulate fraction, calpain II converted to a calpain I-like form exhibiting stronger Ca2+-independent binding to phenyl-Sepharose and a lower Ca2+ requirement for optimal activity. However, it retained its DEAE-cellulose chromatographic pattern, and precipitated with monospecific anti-calpain II antibodies. Although purified calpastatin (endogenous inhibitor) is known to form a Ca2+-dependent complex with calpains, it was not able to reverse the binding of calpains to the particulate fraction upon short incubation. It was, however, effective in blocking calpain binding when the isolated cytosolic fraction or a mixture of purified calpain and calpastatin was preincubated in the presence of Ca2+, and then added to the particulate fraction. Extraction of tissues under controlled conditions revealed that in fact calpains are already loosely associated with subcellular organelles even in the absence of Ca2+. This is the reason why in the crude homogenates with the addition of Ca2+, calpains strongly bind to the particulate fraction without interference by cytosolic calpastatin. Although calpastatin by complexing initially to calpain can prevent the association of this protease with subcellular organelles, it cannot dissociate calpains already bound to these subcellular fractions. By prior Ca2+-independent association with the hydrophobic proteins present in the subcellular fractions, calpains overcome the 3- to 30-fold inhibitory excess of calpastatin in tissues.

Animals↗
PubMed1986

The extracellular matrix of pulmonary scar carcinomas is suggestive of a desmoplastic origin.

Pulmonary scar carcinomas and noncarcinomatous apical scars were subjected to collagen extraction, ultrastructural, and immunocytochemical studies designed to investigate the nature of their extracellular matrix. These studies revealed marked differences in both cellular and biochemical composition of scar carcinomas, compared with apical scars. Myofibroblasts, identified by antimyosin antibodies and confirmed by electron microscopy, constituted over 90% of the stromal cells of the scar carcinomas, compared with 0-10% in the apical scars. Collagen extraction studies revealed both an absolute and relative increase in Type V collagen in the scar carcinomas, compared with that found in the apical scars. The extracellular matrix of the pulmonary scar carcinomas was, however, identical to that of scirrhous carcinomas of the breast. These findings suggest that pulmonary scar carcinomas are probably desmoplastic carcinomas, rather than scar-arising tumors.

Adenocarcinoma↗
PubMed1985

Quantitation of tissue calpain activity after isolation by hydrophobic chromatography.

A rapid and reliable method for quantitating tissue calpains (Ca2+-activated, neutral, thiol proteases) was developed using hydrophobic chromatography with phenyl-Sepharose. Calpains I and II isolated by this method are free of endogenous inhibitor(s) (calpastatin), activator(s), and nonspecific proteases. These calpains expose hydrophobic regions in the presence of Ca2+ and bind tightly to phenyl-Sepharose. Inactivation of bound calpain is prevented by the addition of leupeptin (20 microM). Calpains I and II bound initially by phenyl-Sepharose in a Ca2+-dependent manner are then eluted successively on the basis of their Ca2+-independent binding to phenyl-Sepharose. Because calpastatin may prevent binding of calpain to phenyl-Sepharose by forming a protease-inhibitor complex in the presence of Ca2+, preadsorbing the protease to a suspension of phenyl-Sepharose beads initially in the absence of Ca2+ separates most of the calpain present in tissue extracts from calpastatin. The isolated calpains obtained are assayed by casein digestion. This quantitation procedure is suitable for measuring calpain activity in various tissues and cells including erythrocytes.

Animals↗
PubMed1985

Isolation of S-100 binding proteins from brain by affinity chromatography.

S-100-binding proteins, and calmodulin-binding proteins were isolated from S-100- and calmodulin-depleted bovine brain extract by Ca2+-dependent affinity chromatography using S-100- and calmodulin-coupled Sepharose columns respectively. The majority of the protein (80 to 90%) including calcineurin that bound to S-100 also bound to calmodulin and vice versa, suggesting both proteins may regulate common targets. However these two regulatory proteins also bind few other proteins specific for each. These include cyclic nucleotide phosphodiesterase, 55k, and 220k proteins for calmodulin and 24k, 42k, and 90k proteins for S-100. Certain proteins also specifically bound to S-100 both in Ca2+-dependent and independent ways. In glial cells S-100 protein may replace calmodulin in regulating Ca2+-influenced functions.

Animals↗
PubMed1985

Monoclonal antibodies to the human laminin receptor recognize structurally distinct sites.

Laminin, a glycoprotein of basement membranes, binds to a specific receptor on the surface of neoplastic and non-neoplastic cells. The laminin receptor purified from human breast carcinoma plasma membranes was used as an antigen to generate two types of monoclonal antibodies (mAbs). Both types of mAbs bind to (a) the purified receptor coated on a solid phase; (b) isolated breast carcinoma plasma membranes; and (c) the surface of cultured MCF-7 human breast carcinoma cells by immunohistology. Using immunoblotting, both types of mAbs recognize a single 67 000 Dalton protein among all the proteins extracted from breast carcinoma plasma membranes. The mAbs differed in their ability to block binding of laminin to the plasma membrane receptor. Antibody LR1 inhibited virtually 100% of the specific binding of laminin to both the isolated human breast carcinoma plasma membranes or the living MCF-7 cells. In contrast, antibody LR2 had no effect on laminin binding under identical conditions. Thus, the two types of mAbs may recognize structurally distinct sites on the laminin receptor. These mAbs should be useful to dissect the biology and the molecular genetics of the laminin receptor.

Antibodies, Monoclonal↗