A tool provisioning problem in an FMS

A fundamental decision in designing a flexible manufacturing system (FMS) is the number and types of tools (cutters) to provide for system operation. This tool provisioning decision becomes especially important when space and equipment must be provided for refurbishing and storing tools within the FMS. In this article, we describe in detail the tool provisioning problem for a particular application, discuss approaches to making the tool provisioning decision, and explain why analytic models for this problem may be difficult to develop.     

DNA methylation alters chromatin structure and regulates Thy-1 expression in EL-4 T cells

Thy-1 exhibits marked differences in expression in various tissues in many species; therefore, it is of interest to define possible mechanisms that may regulate Thy-1 expression. We produced Thy-1 negative variants of the murine T cell lymphoma EL-4 by mutagenesis with ethylmethanesulfonate (EMS), negative selection with anti-Thy-1 monoclonal antibodies (mAb) plus complement, and fluorescence-activated cell sorting (FACS). Thy-1 surface negative (Thy-1-) mutants produced in this manner were shown to produce no detectable Thy-1 mRNA, but contained an intact Thy-1 gene as determined by Southern blotting. 5' CG sequences, which had been demethylated in the parent EL-4 clone, were completely methylated in the EMS-induced Thy-1-variant. In addition, a DNase I hypersensitive site that mapped to the 5' end of the Thy-1 gene in EL-4 was absent in the Thy-1- variant. Treatment of this Thy-1- clone with 5-azadeoxycytidine (5-dAZA) resulted in re-expression of surface Thy-1, demethylation of the 5' CG sequences, and regeneration of the DNase I hypersensitive site. These studies indicate that methylation of certain critical DNA sequences in the 5' region of the Thy-1 gene can alter local chromatin structure and regulate expression of this gene.     

The response of multilamellar liposomes to freezing and thawing

The use of liposomes as a model system for investigating the mechanism of freezing injury was investigated. Modification of the liposome phospholipid and cholesterol content allows a correlation to be made between the composition of a membrane system and its response to the stresses of freezing and thawing. The data on phase transitions are contradictory in the sense that liposomes become more sensitive to freezing injury following treatments which both increase or decrease phase transition temperature. In contrast the effect of cholesterol in sensitizing membranes to the stresses of freezing and thawing appears to be more fundamental. Direct cryomicroscope observations of liposomes during slow cooling indicate that they are osmotically active at low temperatures and upon thawing morphological alterations to the membranes occur. The response of liposomes following cooling at a range of rates to ?196 °C and the effects of cryoprotective additives are similar to those observed with many cell types. These results indicate that liposomes are a valid model for investigating the biochemistry of membrane damage induced by the stresses of freezing and thawing.     

Frontiers in mammalian cells culture

Summary For the past 60 years, fundamental discoveries in eukaryotic biology using mammalian cell cultures have been significant but modest relative to the enormous potential. Combined with advances in technologies of cell and molecular biology, mammalian cell culture technology is becoming a major, if not essential tool, for fundamental discovery in eukaryotic biology. Reconstruction of the milieu for cells has progressed from simple salt solutions supporting brief survival of tissues outside the body to synthesis of the complete set of structurally defined nutrients, hormones and elements of the extracellular matrix needed to reconstruct complex tissues from cells. The isolation of specific cell types in completely defined environments reveals the true complexity of the mammalian cell and its environment as a dynamic interactive physiological unit. Cell cultures provide the tool for detection and dissection of the mechanism of action of cellular regulators and the genes that determine individual aspects of cell behavior. The technology underpins advances in virology, somatic cell genetics, endocrinology, carcinogenesis, toxicology, pharmacology, hematopoiesis and immunology, and is becoming a major tool in develomental biology, complex tissue physiology and production of unique mammalian cell-derived biologicals in industry. This article is the first of a series of invited reviews aimed at identifying fundamental contributions and current challenges associated with research activities in subdiscriplines of cell and developmental biology in vitro. This treatise is dedicated to Dr. Brian Kimes, Program Director at the National Cancer Institute, whose vision, encouragement and support have contributed significantly to modern developments in mammalian cell culture.     

Isolation and characterization of an adriamycin-resistant breast tumor cell line

Summary An adriamycin-resistant human breast tumor cell line MDA-A1 ^R was generated by step-wise selection in increasing concentrations of drug from the parent cell line MDA-MB-231. MDA-A1 ^R cells grow as loosely attached cell aggregates with a doubling time of 28–32 h; the MDA-MB-231 parent cell line grows as a standard monolayer culture with a 20-h doubling time. The MDA-A1 ^R cell line is highly resistant to adriamycin compared to the parent cell line, and is cross-resistant to velban and colchicine suggestive of a multidrug resistance (MDR) phenotype. MDA-A1 ^R cells exhibit reduced net adriamycin conent as compared to the parent cell line. The MDR-associated P-glycoprotein gene is amplified approximately 10-to 30-fold in MDA-A1 ^R cells. P-glycoprotein sequences are overexpressed in the resistant cells and are stable for up to 13 wk after drug removal. Moreover, MDA-A1 ^R cells show the presence of very high levels of P-glycoprotein. MDA-A1 ^R is thus an in vitro model system to study the mechanism of MDR in human breast cancer. This work was supported in part by grant C30195 from the National Institute of health, Bethesda, MD. Portion of this study appeared as a poster presentation at the Tissue Culture Association meeting, Las Vegas, 1988.     

Hemorrhagic Shock—Influence of Hyperbaric Oxygen on Metabolic Parameters

Oxygen therapy at atmospheric and increased pressure was used in the treatment of experimental hemorrhagic shock. Arterial gases, pH, lactate and pyruvate were determined and compared. Survival was carefully followed and complete pathological evaluation was carried out. The results showed no difference between control animals and those in the hyperbaric oxygen therapy group. It was concluded that hyperbaric oxygen therapy post facto did not influence the metabolic or survival data in this experiment.