摘要:

AbstractNon‐insulin dependent diabetes mellitus (NIDDM) is characterized by a specific defect in glucose recognition by the pancreatic islet beta cell. This is in clear distinction to patients with insulin dependent diabetes mellitus (IDDM) who undergo pancreatic islet beta cell death and no longer have the ability to synthesize, store, and release insulin. Defective glucose‐induced first phase insulin responses in patients with NIDDM can be partially restored by exogenous insulin treatment and by other pharmacologic therapy. These observations provide strength for the theory of glucose desensitization of the pancreatic beta cell as an important secondary defect in the pathogenesis of abnormal insulin secretion in NIDDM. However, even though defective insulin secretion is an essential part of the pathogenesis of NIDDM, in itself it is not sufficient. A multiplicative effect is required involving interaction between tissue resistance to insulin action and defective insulin secretion whose product is the syndrome of NI

ISSN:0730-2312    

DOI:10.1002/jcb.240480302

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractIon channels in beta cells regulate electrical and secretory activity in response to metabolic, pharmacologic, or neural signals by controlling the permeability to K+and Ca2+. The ATP‐sensitive K+channels act as a switch that responds to fuel secretagogues or sulfonylureas to initiate depolarization. This depolarization opens voltage‐dependent calcium channels (VDCC) to increase the amplitude of free cytosolic Ca2+levels ([Ca2+]i), which triggers exocytosis. Acetyl choline and vasopressin (VP) both potentiate the acute effects of glucose on insulin secretion by generating inositol 1,4,5‐trisphosphate to release intracellular Ca2+; VP also potentiates sustained insulin secretion by effects on depolarization. In contrast, inhibitors of insulin secretion decrease [Ca2+]i by either hyperpolarizing the beta cell or by receptor‐mediated, G‐protein‐coupled effects to decrease VDCC activity. Repolarization is initiated by voltage‐ and Ca2+‐activated K+channels. A human insulinoma voltage‐dependent K+channel cDNA was recently cloned and two types of alpha1 subunits of the VDCC have been identified in insulin‐secreting cell lines. Determining how ion channels regulate insulin secretion in normal and diabetic beta cells should provide pathophysiologic insight into the beta cell signal transduction defect characteristic of non‐insulin depen

ISSN:0730-2312    

DOI:10.1002/jcb.240480303

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractPlatelet‐derived growth factor (PDGF) stimulates the expression of a number of genes associated with entry of quiescent Balb/c‐3T3 fibroblasts into the cell cycle. We determined that two of these genes, c‐mycand c‐fos, are induced equivalently in medium supplemented with platelet‐poor plasma (PPP) and either PDGF‐BB or PDGF‐AA. The rate at which fibroblasts entered S phase was also similar in PDGF‐BB– and AA–treated cells as was the expression of the late G1gene, thymidine kinase (TK). However, PDGF‐AA must be present for a period of 16 h to stimulate the proliferation of 90% of the cells, whereas PDGF‐BB was required for only 4 h. Exposure of cells to PDGF‐AA for 4 h, a time during which maximum expression of c‐fosand c‐mycoccurred, only induced 20% of the cells in a quiescent population to enter the cell cycle. Therefore, PDGF‐AA–mediated expression of the immediate early genes c‐fosand c‐mycmay be necessary but is not sufficient to rapidly stimulate density‐arrested Balb/c‐3T3 fibroblasts into the competent state. Thus, these data suggest that PDGF‐AA and PDGF‐BB initiate trav

ISSN:0730-2312    

DOI:10.1002/jcb.240480304

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractWe have obtained expression of the β‐N‐acetylglucosamine‐binding receptor from chicken hepatocytes inXenopusoocytes by injecting mRNA synthesized in vitro from a full length cDNA cloned into an expression vector (Mellow et al: J. Biol Chem 263:5468–5473, 1988). Immunoprecipitation of the receptor after labeling of oocytes with [35S]‐methionine for times ranging from 6 to 72 h revealed 4–5 closely spaced bands of 25–30 kDa after SDS‐PAGE. Although these bands were largely resistant to endoglycosidase H cleavage, endoglycosidase F reduced the size of all bands to a single species at 23–24 kDa, indicating that they resulted from heterogeneity in glycosylation of a single polypeptide. Incubation of oocytes expressing this receptor with [125I]‐GlcNAc‐BSA resulted in 1.8 to 10 × higher levels of cell‐associated ligand in mRNA‐injected vs. water‐injected control oocytes, 2–35% of cell‐associated counts was removed by EGTA rinse at 20°C, suggesting that most ligand was inaccessible (presumably intracellular). Immunoprecipitation of sucrose gradient fractions detected receptor molecules predominantly in a light organelle at 1.09–1.12 g/cc (the density of early endosomes and plasma membrane vesicles), with no evidence of the receptor in much heavier yolk platelet fractions even in the presence of ligand. In contrast, internalized [125I]‐GlcNAc‐BSA was found either at the top of the gradients or in organelles at 1.09–1.17 g/cc and in yolk platelets. TCA precipitation indicated that much intracellular ligand was degraded to acid‐soluble fragments. Addition of vitellogenin (the yolk protein precursor) to the medium together with the [125I]‐GlcNAc‐BSA shifted much of the ligand into yolk platelets. These data indicate that the chicken glycoprotein receptor expressed in oocytes mediates binding and internalization of this ligand into an organelle in which ligand‐receptor dissociation occurs, allowing for separation of these two molecules into different compartments. The behavior of ligand inXenopusoocytes expressing the chicken rece

ISSN:0730-2312    

DOI:10.1002/jcb.240480305

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractIn cloned osteoblast‐like cells, MC3T3‐E1, prostaglandin F2α(PGF2α) stimulated arachidonic acid (AA) release in a dose‐dependent manner in the range between 1 nM and 10 μM. 12‐O‐tetradecanoylphorbol‐13‐acetate (TPA), a protein kinase C (PKC) activator, which by itself had little effect on AA release, markedly amplified the release of AA stimulated by PGF2αin a dose‐dependent manner, 4 α‐phorbol 12, 13‐didecanoate, a phorbol ester which is inactive for PKC, showed little effect on the PGF2α‐induced AA release. 1‐oleoyl‐2‐acetylglycerol (OAG), a specific activator for PKC, mimicked TPA by enhancement of the AA release induced by PGF2α. H‐7, a PKC inhibitor, markedly suppressed the effect of OAG on PGF2α‐induced AA release. Quinacrine, a phospholipase A2inhibitor, showed partial inhibitory effect on PGF2α‐induced AA release, while it suppressed the amplification by OAG of PGF2α‐induced AA release almost to the control level. Furthermore, TPA enhanced the AA release induced by melittin, known as a phospholipase A2activator. On the other hand, TPA inhibited the formation of inositol triphosphate stimulated by PGF2α. Under the same condition, PGF2αindeed stimulated prostaglandin E2(PGE2) synthesis and TPA markedly amplified the PGF2α‐induced PGE2synthesis as well as AA release. These results indicate that the activation of PKC amplifies PGF2α‐induced both AA release and PGE2synthesis through the potentiation

ISSN:0730-2312    

DOI:10.1002/jcb.240480306

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractThe interaction of125I‐asialoeythropoietin (asialoepo) with receptors has been characterized both by binding assay and affinity cross‐linking. Purified spleen cells from mice infected with the anemia strain of Friend virus (FVA cells) have receptors for123I‐asialoepo with two classes of affinity constant: one with Kd = 0.02–0.03 nM and 300–400 per cell, the other with lower affinity (kd = 0.9–1.2 nM) and 1,000–1,200 per cell. The Kd value for the high affinity site is one third of that for the binding of native125I‐erythropoietin (125I‐epo) to the same FVA cells (Kd = 0.08–0.1 nM). Using125I‐asialoepo or125I‐epo affinity cross‐linking methods, we find two components with apparent molecular weights of 88 kDa and 105 kDa in FVA cells, and in the transformed mouse cell lines, 201, IW32, and NN10, in agreement with earlier studies using125I‐epo. These results indicate that125I‐asialoepo binds to the same receptors as125I‐epo, but with greater affinity for the high affinity site. Since 201 cells contain only a single class of lower affinity receptors for erythropoietin (epo), finding the same two components as found for FVA cells by cross‐linking experiment indicates that the two components do not repr

ISSN:0730-2312    

DOI:10.1002/jcb.240480307

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractThe progressive differentiation of both normal rat osteoblasts and HL‐60 promyelocytic leukemia cells involves the sequential expression of specific genes encoding proteins that are characteristic of their respective developing cellular phenotypes. In addition to the selective expression of various phenotype marker genes, several members of the heat shock gene family exhibit differential expression throughout the developmental sequence of these two cell types. As determined by steady state mRNA levels, in both osteoblasts and HL‐60 cells expression of hsp27, hsp60, hsp70, hsp89α, and hsp89β may be associated with the modifications in gene expression and cellular architecture that occur during differentiation.In both differentiation systems, the expression of hsp27 mRNA shows a 2.5‐fold increase with the down‐regulation of proliferation while hsp60 mRNA levels are maximal during active proliferation and subsequently decline post‐proliferatively. mRNA expression of two members of the hsp90 family decreases with the shutdown of proliferation, with a parallel relationship between hsp89α mRNA levels and proliferation in osteoblasts and a delay in down‐regulation of hsp89α mRNA levels in HL‐60 cells and of hsp89β mRNA in both systems. Hsp70 mRNA rapidly increases, almost twofold, as proliferation decreases in HL‐60 cells but during osteoblast growth and differentiation was only minimally detectable and showed no significant changes. Although the presence of the various hsp mRNA species is maintained at some level throughout the developmental sequence of both osteoblasts and HL‐60 cells, changes in the extent to which the heat shock genes are expressed occur primarily in association with the decline of proliferative activity. The observed differences in patterns of expression for the various heat shock genes are consistent with involvement in mediating a series of regulatory events functionally related to the control of both cell grow

ISSN:0730-2312    

DOI:10.1002/jcb.240480308

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractZymosan and phorbol ester induced in liver macrophages the release of arachidonic acid, prostaglandin E2, and superoxide; the calcium ionophore A 23187 elicited a release of arachidonic acid and prostaglandin E2but not of superoxide, and exogenously added arachidonic acid led to the formation of prostaglandin E2only. The zymosan‐ and phorbol‐ester‐induced release of arachidonic acid, prostaglandin E2, and superoxide was dose‐dependently inhibited by staurosporine and K252a, two inhibitors of protein kinase C, and by pretreatment of the cells with phorbol ester which desensitized protein kinase C. The release of arachidonic acid or prostaglandin E2following the addition of A 23187 or arachidonic acid was not affected by these treatments. Zymosan and phorbol ester but not A 23187 or arachidonic acid induced a translocation of protein kinase C from the cytosol to membranes in intact cells. These results demonstrate an involvement of protein kinase C in the zymosan‐ and phorbol‐ester‐induced release of arachidonic acid, prostaglandin E2, and superoxide; the release of arachidonic acid and prostaglandin E2elicited by A 23187 and the formation of prostaglandin E2from exogenously added arachidonic acid, however, is independent of an activation of pro

ISSN:0730-2312    

DOI:10.1002/jcb.240480309

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractPlasma membrane samples from rat brain, heart, and liver were examined for biochemical changes with age. A rise in superoxide radical (SOR) levels was followed by increases in thiobarbituric acid reactive substances and decreases in membrane fluidity with age. The earliest rise in SOR formation appeared in the plasma membrane from the brain. With age, protein synthesis also decreased significantly in tissue homogenates from brain and heart but was unchanged in the liver. Exposure of plasma membrane samples to in vitro‐elevated SOR levels stimulated formation of lipid peroxides, as indicated by the thiobarbituric acid test, and resulted in a decrease in membrane fluidity in each tissue and in a decline in protein synthesis in brain and heart. Changes in brain lipid peroxidation and in membrane fluidity in brain and heart as a result of SOR supplementation were further enhanced due to age. In addition, the mechanism of SOR formation was examined in plasma membrane samples from the brain. SOR generation was Ca2+‐sensitive, blocked by superoxide dismutase or vitamin E and inhibited by both indomethacin, a cyclooxygenase inhibitor, and bromophenacyl bromide, a phospholipase A2inhibitor. These results show significant increases in SOR formation and biochemical alterations in plasma membranes from brain, heart, and liver in aging rats. SOR formation appears to be enzyme‐mediated and elevated levels of this oxygen radical could be involved in membrane breakdown in older

ISSN:0730-2312    

DOI:10.1002/jcb.240480310

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractIn the rat liver epithelial cell line, WB, the ability of TGF‐β1to inhibit DNA synthesis was shown to correlate with its ability inhibit phosphorylation of the protein product of the retinoblastoma susceptibility gene, pRb. When WB cells were serum‐starved, then refed with serum‐containing medium, a peak of DNA synthesis occurred at about 18 h. Autoradiographs showed that 43.6% of cell nuclei could be labeled with3H‐thymidine at this time. When TGF‐β1was added simultaneously with serum, it blocked DNA synthesis and reduced the number of labeled nucleii to 6.3%. Cells treated with serum alone for 18 h also showed a pronounced increase in the highly phosphorylated form of pRb, as shown by mobility shifts in immunoblots, and in active phosphorylation of pRb, as shown by32P incorporation. Simultaneous addition of TGF‐β1with serum abolished both32P incorporation into pRb and its mobility shift on immunoblots. The effect of TGF‐β1on DNA synthesis measured at 18 h was sharply reduced if the cells were incubated with serum for 8 h (and thus allowed to enter S) before the addition of TGF‐β1. If TGF‐β1was added after 8 h of serum treatment, its ability to inhibit pRb phosphorylation at 18 h was unchanged. If TGF‐β1was added after 13 h of serum treatment, its effects on pRb phosphorylation were reduced. Thus, as the cell population moved into S, the ability of TGF‐β1to inhibit both pRb phosphorylation and DNA synthesis was lost. In higher passages of WB cells the dose‐response for inhibition of DNA synthesis by TGF‐β1was shifted to the right. Inhibition of pRb phosphorylation by TGF‐β1was also lost in higher passage WB cells. Thus, the passage‐dependent loss of sensitivity to inhibition of DNA synthesis accompained the loss of sensitivity to inhibition of pRb phosphorylation. Since the phosphorylation of pRb is believed to be required for the progression of cells from G1to S, inhibition of pRb phosphorylation may be either a cause or a consequenc

ISSN:0730-2312    

DOI:10.1002/jcb.240480311

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY