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Z Ao

Publications and source records attributed to Z Ao.

At least 37 records · Page 2Linked to original sources
PubMed1996

A mitochondrial membrane protein defined by a novel monoclonal antibody is preferentially detected in apoptotic cells.

Studies toward the biologic and molecular understanding of programmed cell death have been stimulated by the recent identification of genes and their products that regulate apoptosis. A panel of mAbs has been raised against dying cells in the present study by immunizing mice with apoptotic Jurkat cells. One of these Abs, anti-7A6, was found to react with apoptotic cells. Using ELISA or flow cytometry, little reactivity of anti-7A6 was observed in normal or digitonin-permeabilized human peripheral blood lymphocytes and a number of hemopoietic cell lines tested. The Ab, however, strongly reacted with these cells when they were induced to undergo apoptosis by irradiation or treatment with apoptosis-inducing agents. Cell sorting and DNA fragmentation experiments revealed that 7A6-positive cells, but not 7A6-negative cells, had apparent DNA fragments characteristic of cells undergoing apoptosis. By immunoblot, under reducing conditions, anti-7A6 detected a 38-kDa protein band in the cell lysates prepared from apoptotic cells. Immunoelectron microscopy showed the 7A6 Ag to be localized to the membrane of mitochondria in apoptotic Jurkat cells. These results indicate that anti-7A6 defines a novel epitope on the mitochondrial membrane protein that appears to be exposed on cells undergoing apoptosis, suggesting that the 7A6 molecule may be involved in the molecular cascade of apoptotic cell death.

Animals↗
PubMed1996

Requirement of Fas(CD95), CD45, and CD11a/CD18 in monocyte-dependent apoptosis of human T cells.

Our previous studies demonstrated that upon activation, monocytes (Mo) were able to sensitize peripheral blood T (PBT) cells to apoptosis induced by treatment with PMA. However, it is unknown what gene products provide the death signal to the sensitized PBT cells and how activated Mo enable PBT cells to become susceptible to apoptosis. Here, we show that PBT cells, but not Mo, express functional Fas ligand upon treatment with PMA. Moreover, this Mo-dependent T cell apoptosis could be blocked by a Fas-Ig fusion protein, as well as by a nonlytic mAb against Fas molecule. These results strongly suggest involvement of Fas-Fas ligand interaction in the death of PBT cells. Unlike Fas-induced apoptosis, however, Mo-dependent T cell death was completely inhibited by overexpression of the Bcl-2 protein, and PMA alone was sufficient to trigger apoptosis in T cells when Mo were included in culture. Furthermore, anti-CD11a, anti-CD18, or anti-CD45/CD45RA mAbs; could prevent PBT cells from death triggered by PMA plus Mo, suggesting that these Ags participate in the apoptotic process. The participation of CD45RA in the death of PBT cells was further demonstrated by the observation that the J45.01 cell line, a CD45-deficient variant of Jurkat cells, did not undergo apoptosis by this Mo-dependent mechanism. When transfected with cDNA encoding CD45RA, J45.01 cells acquired apoptotic response to PMA stimulation in the presence of Mo to a similar, but lesser, degree than normal Jurkat cells.

Antigens, CD↗
PubMed1996

Large scale isolation, growth, and function of porcine neonatal islet cells.

Based upon existing methods of isolating fetal porcine islet tissue, a simple, reliable procedure was developed for the preparation of porcine neonatal islet cell aggregates with a reproducible and defined cellular composition. After 9 d of in vitro culture, tissue from one neonatal pig pancreas yielded approximately 50,000 islet cell aggregates, consisting of primarily epithelial cells (57%) and pancreatic endocrine cells (35%). During the culture period, the total beta cell mass decreased initially, but subsequently increased 1.5-fold between days 3 and 9. Transplantation of grafts consisting of 3 x 10(5) beta cells (1,000 aggregated) under the kidney capsule of alloxan-diabetic nude mice corrected hyperglycemia in 75% (10/13) of the animals, whereas, 100% (20/20) of recipients implanted with 6 x 10(5) beta cells (2,000 aggregates) achieved euglycemia within 8 wk posttransplantation. Nephrectomy of the graft bearing kidney at 14 wk posttransplantation resulted in hyperglycemia in all recipients, and examination of the grafts revealed the presence of numerous well-granulated insulin- and glucagon-containing cells. The cellular insulin content of these grafts was 20 to 30-fold higher than at the time of transplantation. These results indicate that the neonatal porcine pancrease can be used as a source of large numbers of viable islet cells, which have the potential for growth both in vitro and in vivo, and exhibit the metabolic capacity to correct diabetes in nude mice.

Animals↗
PubMed1996

Variables in organ donors that affect the recovery of human islets of Langerhans.

In an attempt to reduce the variability in the yields of human islets isolations and to identify donor factors that were potentially deleterious, we retrospectively reviewed 153 human islets isolations in our center over a 3-year period. Isolations were performed using controlled collagenase perfusion via the duct, automated dissociation, and Ficoll purification. Factors leading to successful isolations (recovery of >100,000 islet equivalents at a purity >50%) were analyzed retrospectively using univariate and multivariate analysis. Critical factors in the multiorgan cadaveric donors that were identified using univariate analysis included donor age (P<0.01), body mass index (BMI)(P<0.01), cause of death (P<0.01), and prolonged hypotensive episodes (systolic blood pressure <90 mmHg or mean arterial pressure <60 mmHg for > 15 min) requiring high vasopressors (>15 microgram/kg/min dopamine or >5 microgram/kg/min Levophed) (P>0.01). Independent analysis of 19 donor variables using multivariate logistic stepwise regression showed six factors were statistically significant. Odds ratio (OR) showed that donor age (OR 1.1, P<0.01), local procurement team (OR 10.9, P<0.01), and high BMI (OR 1.4, P<0.01) had a positive correlation with islet recovery. In contrast, hyperglycemia (all blood glucose >10 mmol/L) (OR 0.63, P<0.01), frequency and duration of cardiac arrest (OR 0.7, P<0.01), and increased duration of cold storage before islet isolation (OR 0.83, P<0.01) had negative correlation. Using these combinations of factors, the prediction of success was 85% accurate. By donor age, success was 13% for 2.5- to 18-year-old donors (n=23), 37% for 19- to 28-year-old donors (n=30), 65% for 29- to 50-year-old donors (n=70), and 83% for 51- to 65-year-old (n=29) donors. However, when vitro function was assessed by perifusion, the insulin secretory capabilities of islets isolated from the >50-year-old donor group was significantly reduced as compared with the 2.5- to 18-year-old group (P<0.02). Multiple regression analysis using postdigestion and postpurification islet recovery as outcome variables identified BMI, procurement team, pancreas weight, and collagenase digestion time factors tht can affect the recovery of human islets. Locally procured pancreases and donors with elevated minimum blood glucose levels were identified as factors that affect the insulin secretory capabilities of the isolated islets. This review of parameters suggests an improved approach to the prediction of successful islet isolation from human pancreases. Selection of suitable pancreases for processing may improve consistency in human islet isolation and thereby decrease costs.

Adult↗
PubMed1994

Cadaver pancreas recovery technique. Impact on islet recovery and in vitro function.

Previous studies in rodent and canine animal models have suggested a detrimental impact on islet recovery and function when pancreas excision is preceded by in situ vascular flushing with cold preservation solutions. We studied the efficacy of islet isolation from 19 consecutive cadaver pancreases procured alternately by initial pancreatectomy before in situ flush (group 1, our standard procurement technique, n = 9) or pancreas removal following in situ vascular flushing with cold University of Wisconsin solution (group 2, n = 10). Once procured, pancreases were weighed, the main pancreatic duct was cannulated, and 150 ml of collagenase solution was injected. The pancreases were transported to the isolation laboratory and processed within 2 hr. Islets were counted and sized after dithizone staining, and the islet equivalents were calculated. Aliquots of isolated islets were cryopreserved using established techniques. Islet function of both freshly isolated and frozen-thawed islets was assessed using a glucose-stimulated perifusion system. Significantly more pancreas was harvested after University of Wisconsin flush (90.6 +/- 6.9 g for group 1 versus 66.7 +/- 4 for group 2, P < 0.05). The quantity of islets per gram of processed pancreas released during enzymatic digestion from each of the experimental groups did not differ significantly (4.5 +/- 0.6 x 10(3) islet equivalents per gram for primary pancreatectomy versus 4.0 +/- 0.4 x 10(3) University of Wisconsin flush). Similarly, following Ficoll purification, the overall yields of islets did not differ significantly. Total islet yield in the primary pancreatectomy group was 181 +/- 25 x 10(3) islet equivalents (2.7 +/- 0.3 x 10(3) IE/g) versus 217 +/- 41 x 10(3) for the University of Wisconsin flush group (2.9 +/- 0.8 x 10(3) islet equivalents/g; P not significant). No differences were observed in in vitro viability. Perifusion stimulation indexes (peak/basal insulin release) were 5.9 +/- 1.3 for group 1 and 7.1 +/- 1.5 for group 2. These results conflict with published results in animal models and indicate that large numbers of viable islets can be recovered from cadaver pancreas utilizing either procurement technique. The decreased operating time, simplicity, and safety favor the use of total pancreatectomy after limited in situ vascular flushing as the method of choice for pancreas procurement for subsequent islet isolation.

Cadaver↗
PubMed1994

Influence of vascular and luminal hexoses on rat intestinal basolateral glucose transport.

The influence of luminal and vascular hexoses in rats on glucose transport across the jejunal basolateral membrane (BLM) was measured using isolated membrane vesicles prepared from infused animals. In vivo vascular infusions of glucose produced an increase in glucose transport across BLM vesicles. Sucrose, mannose, galactose, and fructose had no significant effect. Plasma glucose concentrations were unaffected by galactose and sucrose vascular infusions, while mannose and fructose produced a modest rise, and glucose increased plasma glucose to 20 mM. Insulin release was significantly increased by vascular infusion of glucose and fructose, while mannose produced only a small sustained rise. Sucrose and galactose had no effect. Perfusion through the lumen of the rat jejunum in vivo, for up to 4 h, with glucose, fructose, sucrose, or lactate (100 or 25 mM) produced a significant increase in the maximal rate of glucose transport (up to 4- to 5-fold) across BLMs. Galactose and mannose had no effect. Luminal glucose perfusion produced a small nonsignificant increase in glucose inhibitable cytochalasin B binding to BLM vesicles, and no change was seen in the microsomal pool of binding sites. The abundance of GLUT2 in the jejunal BLM, as determined by Western blotting, was unaffected by luminal perfusion of 100 mM glucose for 4 h. Fructose almost completely inhibited the carrier-mediated uptake of glucose in control and upregulated jejunal BLM vesicles. These results are discussed in relation to the physiological role of the upregulation of GLUT2 activity by luminal and vascular hexoses.

Animals↗