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Biomedical subjects

J G Krikorian

Publications and source records attributed to J G Krikorian.

At least 19 recordsLinked to original sources
PubMed1992

Treatments that extract the 43K protein from acetylcholine receptor clusters modify the conformation of cytoplasmic domains of all subunits of the receptor.

The nicotinic acetylcholine receptor (AChR) of Torpedo electric organ and vertebrate skeletal muscle is closely associated with a Mr 43,000 protein (43K). In this study, we have examined the effects on the AChR of treatments which remove the 43K protein. We used semiquantitative fluorescence techniques to measure the binding of antibodies to clustered AChR in cultured rat myotubes. We found that labeling by antibodies to the cytoplasmic portions of each of the four receptor polypeptides increased significantly upon extraction of the 43K protein. Labeling by an antibody to an extracellular epitope of the alpha subunits was not affected by removal of the 43K protein, suggesting that changes were restricted to the cytoplasmic domains of the AChR. Increases in labeling by antibodies were more limited following protease treatment, which removes most cytoskeletal structures but leaves the 43K protein bound to the membrane. Competition between an antibody to the beta subunit and an antibody to the gamma and delta subunits suggests that the cytoplasmic portion of the AChR still retains a degree of native structure in the absence of the 43K protein. Our results suggest that, although some of these changes may be due to simply exposing additional epitopes on the AChR, the cytoplasmic portions of all the subunits of the AChR undergo significant conformational changes upon extraction of the 43K protein.

Animals↗
PubMed1990

Association of cytoskeletal proteins with newly formed acetylcholine receptor aggregates induced by embryonic brain extract.

Aggregates of acetylcholine receptors (AChR) in muscle cell membranes are associated with accumulations of certain cytoskeletal and peripheral membrane proteins. We treated cultured rat myotubes briefly with embryonic brain extract (EBX) to promote AChR aggregation and determined the distribution of several of these proteins at early stages of aggregation. EBX-treated and control cultures were stained with tetramethylrhodamine-alpha-bungarotoxin to identify AChR aggregates and were then frozen and sectioned on a cryostat. These sections were stained with primary antibodies and fluoresceinated secondary antibodies to localize cytoskeletal proteins. The distributions of AChRs and cytoskeletal proteins was examined qualitatively and analyzed by a semiquantitative assay. Qualitatively, the 43K protein had a distribution that was virtually identical to that of AChR in both control and EBX-treated cultures, and it always colocalized with early AChR aggregates. The 58K protein similarly colocalized with early AChR aggregates, but it was also in aggregate-free areas of muscle membrane. The association of vinculin with the aggregates was quantitatively similar to that of the 43K and 58K proteins, but, qualitatively, its distribution did not follow that of the AChR as closely. Like the 58K protein and vinculin, alpha-actinin, filamin, and actin were concentrated in AChR aggregates and were also enriched elsewhere. However, they were less closely associated with the aggregates, both quantitatively and qualitatively. These results show that AChR aggregates induced by EBX tend to be enriched in the same cytoskeletal proteins that are present at the neuromuscular junction in vivo and at AChR clusters formed at sites of cell-substrate adhesion in vitro. Semiquantitative analysis also revealed that the fractional area of the cell surface associated with vinculin, alpha-actinin, and the 58K protein was the same in controls and EBX-treated myotubes, although the area enriched in AChR and the 43K protein increased about three-fold upon EBX treatment. These results suggest that AChR aggregates may form preferentially in membrane regions that are already enriched in these proteins.

Actinin↗
PubMed1989

Microfilaments and actin-associated proteins at sites of membrane-substrate attachment within acetylcholine receptor clusters.

Rat myotubes in tissue culture form broad areas of close contact with the substrate. These areas often display two distinct, interdigitating sets of membrane domains. One, the "contact domain", is close to the substrate; the other, termed the "AChR domain", is further from the substrate and is rich in acetylcholine receptors (AChR). We have used fluorescence techniques to study the organization of the cytoskeleton in these areas. Substrate-apposed membrane of the myotubes was exposed either by shearing or by permeabilizing the cells with a neutral detergent. Phalloidin derivatives and affinity-purified polyclonal or monoclonal antibodies specific for cytoskeletal proteins were then applied to the samples. Sheared samples were observed by epifluorescence microscopy; detergent-permeabilized samples were observed by total internal reflection fluorescence microscopy. We found that, like antivinculin, fluorescent phalloidin derivatives and antibodies to alpha-actinin, filamin, and talin preferentially labeled the contact domains. This suggests that bundles of microfilaments associate with the membrane at sites of myotube-substrate attachment. In contrast, a 43K protein, closely associated with AChR, was present only at AChR domains. A monoclonal antibody to actin labeled both AChR and contact domains, suggesting that actin is enriched over both regions. Our results suggest that, like the plasma membrane of AChR clusters, the underlying membrane skeleton is organized into at least two distinct domains.

Actin Cytoskeleton↗
PubMed1989

Reorganization and stabilization of acetylcholine receptor aggregates on rat myotubes.

Aggregation of acetylcholine receptors (AChRs) is an important early feature of the postsynaptic development of the vertebrae neuromuscular junction. At later stages of differentiation, aggregates are remodeled and stabilized. Aggregation of AChRs can be induced on rat myotubes in culture within 4 hr by treatment with embryonic pig brain extract (EBX). In this study, further sequential changes in the distribution of AChRs were followed by video-intensified fluorescence microscopy. These studies have revealed that groups of AChR aggregates that have formed after 4 hr in EBX are reorganized during the exposure to EBX for 20 additional hr to form a smaller number of larger, oval-shaped aggregates. We have named these two types of aggregates "4-hr aggregates" and "24-hr aggregates". This reorganization occurs by the expansion and merging of individual aggregates within a group, and by the incorporation of newly inserted AChRs. The 24-hr aggregates are an average of 15 times greater in area than 4-hr aggregates, and contain regions with an apparent AChR site density (fluorescence intensity) that is more than twice that of 4-hr aggregates. Electron microscopy of mapped 24-hr aggregates revealed that folded plasma membrane is associated with these regions, probably accounting for the elevated fluorescence. The 24-hr aggregates are more stable than 4-hr aggregates, as determined by their significantly slower disassembly after removal of EBX, elevation of temperature (38 degrees C), reduction of extracellular calcium levels (0.1 mM), or the addition of sodium azide (7 mM). This was determined by following disassembly both statistically (using fixed cultures) and by direct observations of living myotubes. These findings were confirmed by measuring the sequential changes in relative AChR site density over time in individual living myotubes. Thus, 24-hr aggregates form by the reorganization of 4-hr aggregates; exhibit a more regular, compact shape; and are more stable than 4-hr aggregates. These changes in AChR organization and aggregate stability resemble the changes occurring after the initial formation of junctional AChR aggregates during embryonic development, demonstrating additional similarities between this model system and the developing neuromuscular junction.

Animals↗
PubMed1987

Lymphocyte predominance Hodgkin's disease. A clinicopathologic reassessment.

A clinicopathologic analysis of 35 cases of lymphocyte predominance Hodgkin's disease (LPHD) was performed, and the relationship between various histologic features and stage at presentation was examined. The series comprised 31 male and 4 female patients with a median age of 32 years. Twenty patients had Ann Arbor Stage I disease; 5, Stage II; 8, Stage III; and 2, Stage IV. Eight cases showed a multifocal epithelioid histiocytic reaction; six of these had advanced-stage (III or IV) disease (P = 0.003). Five cases contained relatively frequent atypical mononuclear cells; four of these were Stage III or IV (P = 0.017). A purely diffuse growth pattern was also associated with high stage (P = 0.031). Presence of nodularity correlated with low stage (P = 0.031). During follow-up (median, 4 years; range, 2.5 months-18 years) there was a high incidence of second neoplasms (2 large cell lymphomas, 3 non-hematologic malignancies). LPHD manifests greater diversity of stage at presentation than originally thought. A high content of epithelioid histiocytes, relatively frequent atypical mononuclear cells, and diffuse histology appear to define subsets associated with advanced-stage disease. The correlation of nodular growth pattern with low-stage disease is consistent with the distinctive behavior and derivation of nodular LPHD.

Adolescent↗
PubMed1986

Early stages in the formation and stabilization of acetylcholine receptor aggregates on cultured myotubes: sensitivity to temperature and azide.

We have studied the effects of temperature and sodium azide on the formation and stability of embryonic brain extract (EBX)2-induced acetylcholine receptor (AChR) aggregates on myotubes. Sequential changes in AChR distribution were studied on living myotubes in culture by video-intensified fluorescence microscopy. Aggregate formation was temperature dependent, increasing sharply from 24-36 degrees, maximal at 36-37 degrees, and virtually blocked at 38-40 degrees. Whereas aggregate size increased rapidly with time (up to 4 hr) at 36 degrees, at 18-24 degrees small (less than or equal to 1 micron) "microaggregates" formed and accumulated for up to 10 hr. Aggregates formed within 1.5 hr at the sites of microaggregates (formed after 4 hr at 23 degrees) if the temperature was raised to 36 degrees. However, if EBX was removed, the microaggregates on 50% of myotubes disassembled within 1.5 hr. The formation of microaggregates at 23 degrees and aggregates at 36 degrees was reversibly inhibited by sodium azide. These results show that clusters of microaggregates are the precursors of aggregates, and suggest that microaggregate clouds represent a discrete, labile, ATP-dependent stage in aggregate formation. Aggregates that had formed after 4 hr in the presence of EBX disassembled slowly (within 12-14 hr) following removal of EBX at 36 degrees, and even more slowly at 23-30 degrees. However, a temperature shift to 38 degrees, or the addition of azide, resulted in a rapid but reversible disassembly of aggregates (within 4 hr). Thus, newly formed aggregates appear to be relatively stable structures, while microaggregate clouds are labile, tending to disassemble or evolve into aggregates.

Animals↗
PubMed1985

Central nervous system pharmacokinetics of high-dose cytosine arabinoside.

The cerebrospinal fluid (CSF) pharmacokinetics of cytosine arabinoside (Ara-C) was determined in 8 patients with metastatic cancer in the central nervous system. High dose (3 gm/M2) one-hour intravenous infusions of Ara-C were given with serial CSF sampling obtained from indwelling Ommaya reservoirs. CSF was analyzed by high pressure liquid chromatography. Mean Ara-C elimination half life of 140 minutes in CSF was eight times longer than that in plasma. The peak mean CSF concentration of Ara-C (2.1 micrograms /ml) was about 7% of the plasma concentration (30 micrograms/ml). A total of 28 treatment courses were administered with minimal hematopoietic, gastrointestinal and neurological toxicities. A schedule of administration of 2 high-dose treatments 12 hours apart repeated every two weeks should maintain cytotoxic CSF concentrations which could prove useful in the management of CNS leukemia and lymphoma.

Adult↗
PubMed1985

Clinical pharmacology of intraperitoneal cisplatin.

The clinical pharmacology of cisplatin was determined in six patients with malignant ascites secondary to ovarian cancer, and in one patient with peritoneal mesothelioma, following intraperitoneal administration of cisplatin (25-60 mg/m2). The drug was administered in 1 liter of normal saline as a 15- to 30-min infusion. Total, and in some patients free (ultrafilterable), platinum concentrations were determined in plasma, urine, and ascitic fluid by flameless atomic absorption spectrometry. The peak total platinum concentrations in ascitic fluid at the end of infusion were related to dose, a 50 mg/m2 dose producing a 20 to 80 micrograms cisplatin/ml concentration. Filterable platinum represented between 3 and 59% of total platinum in the peritoneum at 4 to 6.5 hr following its administration. Plasma platinum concentrations ranged between 0.2 to 1.6 micrograms/ml 4 hr following administration, and reached a plateau for the next 24 to 48 hr largely in the form of protein-bound platinum. The urinary excretion of cisplatin was consistent with variation in absorption from the peritoneum. Minimal gastrointestinal, bone marrow, and renal toxicities during therapy suggest that sustained free platinum concentrations in ascites may be obtained without significant toxicity and support the intraperitoneal route of administration as an effective strategy for cisplatin therapy of intra-abdominal malignancies.

Aged↗
PubMed1985

Etoposide and cisplatin salvage chemotherapy for small cell lung cancer.

Thirty patients with previously treated small cell lung cancer received salvage combination chemotherapy with etoposide and cisplatin. Two complete and six partial responses were observed, for a major response rate of 27%. Responses occurred promptly and sustained palliation was achieved among responders. Myelosuppression was the major dose-limiting toxic effect. A schedule of etoposide (115 mg/m2 iv on Days 1-3) and cisplatin (25 mg/m2 iv on Days 1-3 every 28 days) is recommended for further clinical trials.

Aged↗
PubMed1983

Cerebrospinal fluid and plasma pharmacokinetics of high doses of 1-beta-D-arabinofuranosylcytosine in nonhuman primates.

The pharmacokinetics of 1-beta-D-arabinofuranosylcytosine (ara-C) in plasma, cerebrospinal fluid (CSF), and urine was studied in nonhuman primates. Conventional and high-dose schedules of ara-C were administered i.v. and intraventricularly through indwelling Ommaya reservoirs. ara-C and its metabolite 1-beta-D-arabinofuranosyluracil (ara-U) were measured by high-pressure liquid chromatography. Due to rapid peripheral deamination (230 nm ara-C/hr/ml plasma), the half-life of ara-C in plasma after a 140-mg/kg i.v. 1-hr infusion was short (3.7 min). Peak plasma concentrations of ara-C, ranging between less than 0.1 and 49 micrograms/ml, were dose dependent (ara-C, 15 to 140 mg/kg). Under these conditions, ara-C was undetectable in CSF. About 53% of the administered dose (140 mg/kg) was excreted in urine during the first 5 hr mostly as ara-U. Intraventricular administration of 50 and 250 mg of ara-C resulted in peak lumbar CSF levels of 435 and 2235 micrograms/ml, respectively, at about 155 and 80 min postinjection. Concomitant ara-U levels were one-tenth of those of ara-C and increased progressively, suggesting deamination of the drug in the central nervous system. The half-life of ara-C in CSF ranged between 50 and 60 min. Administration of 50 mg of ara-U intraventricularly resulted in peak lumbar ara-U levels of 595 micrograms/ml at about 180 min with a prolonged clearance. Concomitant plasma levels throughout the study were less than 0.1 micrograms/ml, suggesting slower equilibrium. No hematological or nervous system toxicity was observed during these studies. The clinical implications of these findings are discussed.

Animals↗
PubMed1982

Errors in measurement and reporting of serum markers in patients with germ cell tumors.

Elevation of the serum value of either human chorionic gonadotropin or alpha-fetoprotein in patients with germ cell testicular tumors is often accepted as evidence for persistent malignancy in the absence of histologic confirmation. These laboratory values are not subject to review and confirmation, unlike histologic diagnosis. We report 3 cases in which serum marker values were incorrectly reported to be elevated by commercial laboratories over a thirteen-month period and suggest a method for handling serum samples to avoid errors in diagnosis.

Adult↗
PubMed1980

Spontaneous regression of non-Hodgkin's lymphoma: a report of nine cases.

Seven previously untreated patients and two previously treated patients with advanced non-Hodgkin's lymphoma (Stages III and IV) and favorable histologic subtypes had spontaneous regression of their lymphomas. Regressions were either complete or partial and were frequently durable. Six of the seven patients who had spontaneous regression of their lymphomas prior to any therapy have yet to require treatment. Seven of the nine spontaneous regressions occurred in a group of 44 patients who were followed with initial therapy deferred. Six patients had regression of their lymphomas prior to any therapy and one patient had previously received a small field of radiation therapy. Temporary spontaneous regression of lymphoma may be common in selected patients with favorable histologies and advanced disease in whom initial therapy is deferred.

Adult↗