Cytochemical and scanning electron-microscopic analysis of apoptotic cells and their phagocytosis in mucoid epithelium of the mouse stomach

Summary The formation of apoptotic cells and their phagocytosis by viable neighbouring cells in the gastric epithelium of 2-to 6-day-old mice was analysed. In order to observe the topographic relationship between apoptotic and normal epithelial cells using scanning electron microscope, the critical-point dried tissues was cracked before coating with gold. Cytochemical methods for the identification of surface carbohydrates and different tracers for apical and lateral cell membranes were applied for the analysis using the transmission electron microscope. Apoptotic cells were found on apical and lateral surfaces; this indicates the presence of tight connections with viable cells at some points. Ruthenium red strongly stained all accessible surfaces of normal cells and of apoptotic bodies. The quantity of neutral mucosubstances, as revealed by staining with tannic acid-uranyl acetate, seemed to decrease in the glycocalyx of apoptotic cells. The scanning and transmission electronmicroscopic results suggest that the phagocytotic vacuoles arise at the lateral side of the cells. The phagocytotic activity is not dependent upon a definite differentiation step of the mucoid cell.     

Two Types of Apoptotic Cell Death of Rat Central Nervous System-Derived Neuroblastoma B50 and B104 Cells: Apoptosis Induced During Proliferation and After Differentiation

Abstract: We describe here two types of apoptotic cell death observed in the rat CNS-derived neuroblastoma B50 and B104 cells. One type was induced by dibutyryl cyclic AMP (DBcAMP) after differentiation, and the other was induced by treatment of proliferating cells with cycloheximide. When B50 and B104 cells were treated with 1 m M DBcAMP in the presence of 0.5% fetal calf serum, they began to extend neurites within 12 h and differentiated into neurons at 24 h, as reported previously. However, further cultivation with DBcAMP for up to 72 h led to flotation and, finally, death. Death was by apoptosis as shown by chromatin condensation and DNA fragmentation. Addition of a protein kinase A inhibitor or removal of DBcAMP after differentiation suppressed apoptosis, indicating the involvement of cyclic AMP and protein kinase A in apoptotic cell death. Cell death was also induced in proliferating cells without neurite outgrowth by treatment with cycloheximide. The death was also judged to be by apoptosis based on chromatin condensation and apoptotic body formation, although DNA fragmentation into small sizes was not detected. Both types of cell death showed similar responses to inhibitors for protein kinases and protein phosphatases.     

Molecular Regulation of Renal Phosphate Transport

Received: 18 April 1996/Revised: 26 June 1996     

Induction of apoptosis and c-myc in L1210 lymphocytic leukemia cells by adenosine

High concentrations of adenosine (Ado), when added to L1210 lymphocytic leukemia cells, resulted in apoptosis or programmed cell death. The apoptotic process was accompanied by distinct morphological changes including chromatin condensation and blebbing of plasma membranes. Extensive DNA fragmentation was correlated with Ado concentrations. Furthermore, apoptosis in these cells was preceded by an early but transient expression of c-myc proto-oncogene, and was not influenced by homocysteine thiolactone added to the cells. Since severe combined immunodeficiency (SCID) is associated with a deficiency of adenosine deaminase, leading to defects in both cellular and humoral immunity, Ado-induced apoptosis may thus be a contributing factor in the pathology of SCID.     

Cell Cycle Arrest of Proliferating Neuronal Cells by Serum Deprivation Can Result in Either Apoptosis or Differentiation

Abstract: Apoptotic cell death plays a critical role in the development of the nervous system. The death of mature nondividing neurons that fail to receive appropriate input from the target field has been extensively studied. However, the mechanisms mediating the extensive cell death occurring in areas of the developing brain where proliferating neuroblasts differentiate into mature nondividing neurons have not been analyzed. We show here that the cell cycle arrest of a proliferating cell of neuronal origin by removal of serum results in either apoptotic cell death or differentiation to a mature nondividing neuronal cell. The proportion of cells undergoing death or differentiation is influenced in opposite directions by treatment of the cells with cyclic AMP and retinoic acid. This suggests that following the withdrawal of signals stimulating neuroblast cell division, neuronal cells either can cease to suppress a constitutive suicide pathway and hence die by apoptosis or, alternatively, can differentiate into a mature neuronal cell. Regulation of the balance between apoptosis and neuronal differentiation could therefore play a critical role in controlling the numbers of mature neurons that form.     

Aurintricarboxylic Acid Protects Hippocampal Neurons from NMDA-and Ischemia-Induced Toxicity In Vivo

Abstract: The polymeric dye aurintricarboxylic acid (ATA) has been shown to protect various cell types from apoptotic cell death, reportedly through inhibition of a calcium-dependent endonuclease activity. Recent studies have indicated that there may be some commonalities among apoptosis, programmed cell death, and certain other forms of neuronal death. To begin to explore the possibility of common biochemical mechanisms underlying ischemia-or excitotoxin-induced neuronal death and apoptosis in vivo, gerbils or rats subjected to transient global ischemia or NMDA microinjection, respectively, received a simultaneous intracerebral infusion of ATA or vehicle. As a biochemical marker of neuronal death, spectrin proteolysis, which is mediated by activation of calpain I, was measured in hippocampus after 24 h. ATA treatment resulted in a profound reduction of both NMDA-and ischemia-induced spectrin proteolysis, consistent with the possibility of some common mechanism in apoptosis and other forms of neuronal death in vivo.     

mTOR in programmed cell death and its therapeutic implications

Mechanistic target of rapamycin (mTOR), a highly conserved serine/threonine kinase, is involved in cellular metabolism, protein synthesis, and cell death. Programmed cell death (PCD) assists in eliminating aging, damaged, or neoplastic cells, and is indispensable for sustaining normal growth, fighting pathogenic microorganisms, and maintaining body homeostasis. mTOR has crucial functions in the intricate signaling pathway network of multiple forms of PCD. mTOR can inhibit autophagy, which is part of PCD regulation. Cell survival is affected by mTOR through autophagy to control reactive oxygen species production and the degradation of pertinent proteins. Additionally, mTOR can regulate PCD in an autophagy-independent manner by affecting the expression levels of related genes and phosphorylating proteins. Therefore, mTOR acts through both autophagy-dependent and -independent pathways to regulate PCD. It is conceivable that mTOR exerts bidirectional regulation of PCD, such as ferroptosis, according to the complexity of signaling pathway networks, but the underlying mechanisms have not been fully explained. This review summarizes the recent advances in understanding mTOR-mediated regulatory mechanisms in PCD. Rigorous investigations into PCD-related signaling pathways have provided prospective therapeutic targets that may be clinically beneficial for treating various diseases.     

The role of apoptosis in antibody-dependent cellular cytotoxicity

Apoptosis in three lymphoma cell lines has been studied following cytotoxicity induced in vitro by normal human blood lymphocytes utilizing either natural killer (NK) or antibody-dependent cellular cytotoxic (ADCC) mechanisms. Guinea-pig L₂C leukaemic lymphocytes, but not the human cell lines Daudi and Jurkat, revealed a degree of time- and temperature-dependent apoptotic death upon simple culture in vitro. NK cytotoxicity at low effector: target ratios (E: T) induced both release of⁵¹Cr and apoptosis. However NK cytotoxicity at higher E : T, and ADCC at all E : T, increased the level of⁵¹Cr release while reducing the level of apoptosis. The findings were consistent with the apoptotic process being cut short by intervention of necrotic death. The same characteristics accompanied ADCC whether the effectors were recruited by Fc regions of antibody coating the targets, or by bispecific antibodies attaching one arm to the targets and the other to Fc receptors type III on effectors. This finding, and the high level of cytotoxicity elicited by the bispecific method, confirm the belief that NK cells, in addition to exerting NK cytotoxicity, represent the principal effectors for ADCC among blood mononuclear cells. Our results suggest that NK cells have both apoptotic and necrotic mechanisms available for killing their targets, but use only the latter for ADCC.