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L W Powell

Publications and source records attributed to L W Powell.

At least 91 records · Page 5Linked to original sources
PubMed1994

The endocytic pathway for H-ferritin established in live MOLT-4 cells by laser scanning confocal microscopy.

We have established the intracellular destination of the putative immunoregulatory protein, human recombinant H (heavy)-ferritin, in the transformed T-cell line MOLT-4, by laser scanning confocal microscopy of live cells. A series of confocal images was collected over a 60 min time course using indirect immunofluorescence of H-ferritin and transferrin, their respective monoclonal antibodies, and fluorescein isothiocyanate (FITC)-labelled IgG. A marked drop in FITC fluorescence after 40 min of H-ferritin internalization, indicative of an acidic environment, and co-localization with tetramethylrhodamine isothiocyanate-labelled-dextran strongly suggests that H-ferritin is transferred to the lysosome. In contrast, transferrin was observed to return to the cell surface. Electron microscopy confirmed that H-ferritin was transferred to the lysosome. The receptor-mediated endocytosis and lysosomal delivery of H-ferritin may thus potentiate its putative immunoregulatory activity.

Endocytosis↗
PubMed1994

Isolation of CA dinucleotide repeats close to D6S105; linkage disequilibrium with haemochromatosis.

The gene for hereditary haemochromatosis (HC) is linked to HLA-A and D6S105 on chromosome 6p. Both markers have also been reported to display linkage disequilibrium with the disease. However, their physical localization relative to one another has not been established. We demonstrate by fluorescent in situ hybridisation that D6S105 lies at least 1-2 Mb telomeric of HLA-A. The haemochromatosis critical region extending from proximal of HLA-A to distal of D6S105 is therefore large. To improve the genetic resolution in this region more highly polymorphic markers are required. We have therefore isolated three novel CA dinucleotide repeats close to D6S105. A linkage disequilibrium study, with two of these microsatellites, in HC patients and controls lends support to the conclusion that D6S105 is a close marker to the haemochromatosis gene.

Alleles↗
PubMed1994

Pathways of intracellular trafficking and release of ferritin by the liver in vivo: the effect of chloroquine and cytochalasin D.

We have previously shown that the clearance of exogenous ferritin and the release of endogenous ferritin into both serum and bile are altered by the microtubular inhibitor colchicine. In this study we further examined the role of the lysosome-endosome pathway in ferritin metabolism. We examined the effect of the lysosomotropic agent chloroquine and the microtubular inhibitor cytochalasin D on the uptake and release of ferritin by normal and iron-loaded rats under basal conditions and in the presence of an exogenous tissue ferritin load. Either chloroquine (50 mg/kg body wt) or cytochalasin D (0.9 microgram/100 gm body wt/min) was administered to normal and iron-loaded rats at zero time. Rats were also infused with either saline solution or rat liver ferritin containing a trace amount of 125I-ferritin. The clearance of 125I-ferritin from the circulation was not affected by chloroquine or cytochalasin D either in normal or in iron-loaded rats; however, both chloroquine and cytochalasin D decreased the serum ferritin concentration in normal rats to 39% +/- 9% and 22% +/- 7% of the baseline serum ferritin levels, respectively, implying that both drugs inhibited the release of endogenous ferritin in normal rats. In iron-loaded rats both chloroquine and cytochalasin D decreased the biliary ferritin concentration to 11% +/- 1% and 37% +/- 4% of the baseline ferritin levels, respectively, and the 125I protein-bound counts per minute in the bile to 50% of the control result. This finding is consistent with an inhibitory effect of both drugs on the biliary excretion of endogenous ferritin and the intracellular transport of exogenous ferritin, respectively. In the presence of an exogenous tissue ferritin load, there was no detectable inhibitory effect of either drug on the biliary excretion of either endogenous or exogenous ferritin. These results provide the following evidence: (a) the receptor-mediated endocytosis of ferritin is not dependent on functioning lysosomes or microfilaments; (b) the release of endogenous ferritin into the serum of normal rats and the bile of iron-loaded rats is a chloroquine-sensitive, microfilament-dependent process; (c) the biliary excretion of trace amounts of exogenous ferritin is dependent on both chloroquine-sensitive vesicles and microfilaments; and (d) increased levels of exogenous ferritin are excreted directly into the bile by way of a second microfilament-independent, chloroquine-insensitive pathway. This study provides support for a physiological mechanism for the release of ferritin from the liver.

Alanine Transaminase↗
PubMed1993

Preneoplastic significance of hepatic iron-free foci in genetic hemochromatosis: a study of 185 patients.

Sublobular nodules of hepatocytes free of iron or exhibiting much less iron than the surrounding parenchyma, referred to in this study as iron-free-foci, are frequently found in the livers of patients with genetic hemochromatosis complicated by hepatocellular carcinoma. To test the hypothesis that such nodules are preneoplastic lesions, iron-free foci were sought in the initial liver biopsy specimens of 185 patients with untreated and uncomplicated genetic hemochromatosis. Iron-free foci were found in 14 (7.6%) patients, all men, aged from 38 to 76 yr, with heavy iron overload and with fibrosis or cirrhosis. Twelve patients with iron-free foci were followed for 0.9 to 15 yr (7 +/- 6 yr). In six (50%), HCC developed, compared with 2 (8%) from a control group consisting of 24 patients without IFF matched according to age, sex, degree of fibrosis, liver iron amount and follow-up duration. The mean number of iron-free foci per iron-free foci-positive specimen was 3.2 +/- 2.1. Ten patients had dysplastic aspects in their iron-free foci, and four had intrahepatocytic iron-positive inclusions at the periphery of iron-free foci. Proliferative cell nuclear antigen was positive in 75% of iron-free foci and in 24% +/- 21% of hepatocyte nuclei in iron-free foci. This study clearly demonstrates that iron-free foci are proliferative lesions and strongly suggests that such nodules are preneoplastic foci. Therefore the finding of IFF in the initial liver biopsy specimen from a patient with genetic hemochromatosis should lead to regular screening for hepatocellular carcinoma.

Adult↗
PubMed1993

Localization of the hemochromatosis gene close to D6S105.

The hemochromatosis (HC) gene is known to be linked to HLA-A (6p21.3); however, its precise location has been difficult to determine because of a lack of additional highly polymorphic markers for this region. The recent identification of short tandem repeat sequences (microsatellites) has now provided this area with a number of markers with similar polymorphic index to the HLA serological polymorphisms. Using four microsatellites--D6S105, D6S109, D6S89, and F13A--together with the HLA class I loci HLA-A and HLA-B in 13 large pedigrees clearly segregating for HC, we have been able to refine the location of the HC gene. We identified no recombination between HC and HLA-A or D6S105, and two-point analyses placed the HC gene within one centimorgan (cM) of HLA-A and D6S105 (HLA-A maximum of the lod score [Zmax] of 9.90 at recombination fraction [theta] of 0.0, and D6S105 Zmax of 8.26 at theta of 0.0). The markers HLA-B, D6S109, D6S89, and F13A were separated from the HC locus by recombination, defining the centromeric and telomeric limits for the HC gene as HLA-B and D6S109, respectively. A multipoint map constructed using HLA-B, HLA-A, and D6S109 indicates that the HC gene is located in a region less than 1 cM proximal to HLA-A and less than 1 cM telomeric of HLA-A. These pedigree data indicate an association between HC and specific alleles at HLA-A and D6S105 (i.e., HLA-A3 and D6S105 allele 8).(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1993

Effect of chronic iron overload on procollagen gene expression.

The pathogenesis of hepatic fibrosis and cirrhosis in genetic hemochromatosis may involve a direct effect of excess iron on collagen synthesis in the liver. To investigate this theory, we measured procollagen messenger RNA levels (types I, III and IV) in the livers of rats in which we produced chronic parenchymal iron overload by feeding them dietary carbonyl iron (2.5%, wt/wt) for up to 18 mo. This feeding resulted in predominantly parenchymal iron deposition in a periportal distribution similar to that seen in genetic hemochromatosis. Increased amounts of collagen fibrils were observed in iron-loaded livers on electron microscopy; all iron-loaded livers showed some periportal fibrosis. Although very high hepatic iron concentrations (range = 340 to 1,100 mumol/gm dry wt) were achieved in the carbonyl iron-loaded rats, we saw no consistent difference between steady-state messenger RNA levels for procollagens types I, III and IV in control and iron-loaded livers examined at five different time points up to 18 mo. Messenger RNA levels of the cytokine transforming growth factor-beta 1, which has been implicated as having a role in the production of extracellular matrix proteins, were also measured. No significant differences were observed between iron-loaded and control livers. These results suggest that excess parenchymal iron does not have a direct effect on the expression of the procollagens or transforming growth factor-beta 1 genes in iron-loaded livers and that factors other than, or in addition to, iron are necessary for fibrosis to occur.

Animals↗
PubMed1993

Differentiation between heterozygotes and homozygotes in genetic hemochromatosis by means of a histological hepatic iron index: a study of 192 cases.

The biochemical hepatic iron index, defined as the ratio of hepatic iron concentration (expressed as micromoles per gram dry weight) to age permits accurate prediction of genetic status in patients with genetic hemochromatosis. However, the hepatic iron concentration is not always available. Therefore a histological hepatic iron index, defined as the ratio of total histological iron score (range = 0 to 60) to age, was evaluated in a total of 192 Australian and French patients with genetic hemochromatosis. These subjects had been classified previously as heterozygotes (n = 18) or homozygotes (n = 174) according to clinical and familial data only. Biochemical hepatic iron index and histological hepatic iron index were well correlated (Spearman's test: rho = 0.75, p < 0.0001). Both were significantly (p < 0.0001) increased in homozygotes (respectively, 6.7 +/- 3.8 [range = 1.2 to 22.6] and 0.62 +/- 0.28 [range = 0.14 to 1.5]) compared with heterozygotes (respectively, 1 +/- 0.4 [range = 0.45 to 1.6] and 0.08 +/- 0.05 [range = 0 to 0.14]). The histological hepatic iron index was less than 0.15 in all heterozygotes and greater than 0.15 in all but two homozygotes. These data show that the age-dependent nature of iron accumulation can also be accommodated by calculating the histological hepatic iron index and that histological study is an accurate means of predicting the genetic status of hemochromatosis patients when hepatic iron concentration is not available.

Adult↗
PubMed1993

Concordance of iron storage in siblings with genetic hemochromatosis: evidence for a predominantly genetic effect on iron storage.

Phenotypic concordance between siblings has been demonstrated in some inherited conditions, and such data provide strong evidence that the severity of disease is affected by genetic factors. We assessed the concordance of liver iron stores between siblings in 22 sibling pairs (15 same-sex pairs and 7 opposite-sex pairs) with genetic hemochromatosis. In this study population a wide range was found in the hepatic iron concentration and the hepatic iron index (32 to 833 mumol/gm dry wt and 1.65 to 14.4, respectively), which could not be accounted for by differing exposure to the environmental factors that influence iron stores. Despite the large variation of hepatic iron concentration within the group, siblings of identical sex had accumulated similar amounts of liver iron. A highly significant correlation for hepatic iron concentration (r = 0.81) and hepatic iron index (r = 0.70) was found between siblings of the same sex. The hepatic iron concentration and the hepatic iron index of one sibling was less than 50% of the other in only three same-sex sibling pairs. In each instance, reasons existed (blood donation, age at diagnosis and human leukocyte antigen nonidentity) for this discordance. In six of the seven opposite-sex pairs, the woman had lower hepatic iron stores than her male sibling, consistent with previous studies on the extent of iron overload in women and its modification by physiological blood loss.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1993

Distribution of body water in patients with cirrhosis: the effect of liver transplantation.

We compared total body water and intracellular-extracellular distribution of body water between male patients with mild liver disease without ascites (n = 9), male patients with severe liver disease and gross ascites (n = 6) and a group of age-, sex-, height- and weight- matched controls (n = 6). In addition, we documented the effects of liver transplantation on intracellular, extracellular and total body water in 12 patients (6 men and 6 women) by means of deuterium oxide dilution and whole-body potassium counting. We saw no significant difference in total body water between the healthy controls, patients without ascites and patients with ascites (46.5 +/- 9.2 kg, 45.4 +/- 6.6 kg and 50.4 +/- 5.1 kg, respectively), although, as expected, extracellular water was increased in patients with ascites compared with healthy controls and cirrhotic patients without ascites (36.9 +/- 6.5 kg vs. 25.4 +/- 4.4 kg, p = 0.005; and 36.9 +/- 6.5 kg vs. 27.0 +/- 5.3 kg, p = 0.002, respectively). We found no difference between non-ascitic patients and healthy controls (25.4 +/- 4.4 kg vs. 27.0 +/- 5.3 kg). However, intracellular water was significantly reduced in patients with severe liver disease compared with that in controls (13.6 +/- 3.3 kg vs. 21.5 +/- 4.2 kg, p = 0.005) or patients without ascites (13.6 +/- 3.3 kg vs. 18.3 +/- 2.9 kg, p = 0.01). The reduction of intracellular water appears to be due to loss of body cell mass.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1992

Universal occurrence of glomerular abnormalities in patients receiving liver transplants.

We conducted a prospective study of renal histology and function in 18 consecutive nonalcoholic patients who underwent orthotopic liver transplantation (OLT). Despite well-preserved renal function, all patients had abnormal renal biopsies. Four patterns of glomerular injury were identified: minor glomerular abnormalities (eight patients), hepatic glomerulosclerosis (seven), membranoproliferative glomerulonephritis (one), and IgA nephropathy (one). In one patient there was insufficient tissue to allow classification. There was a trend toward lower plasma bilirubin and higher plasma albumin in patients with minor glomerular abnormalities than in the group of patients with more severe forms of glomerular injury (29 v 82 mumol/L, 35.5 v 30 g/L; P = 0.1, 0.1 greater than P greater than 0.05, respectively). Glomerular changes persisted in the three patients who died within 7 weeks post-OLT. IgM immunofluorescence was present in all biopsies and IgA in 11. IgM-containing circulating immune complexes occurred in five patients, suggesting a pathogenic role for IgM immune complex deposition. The significance of cirrhosis-associated glomerular abnormalities is not yet known. They may contribute to the hepatorenal syndrome and the renal dysfunction that occurs in up to 94% of patients post-OLT.

Adolescent↗
PubMed1992

Characterization of the ferritin receptors of human T lymphoid (MOLT-4) cells.

We have previously demonstrated that distinct binding sites exist for human recombinant H ferritin (HrHF) and human liver ferritin (HLF) on human T lymphoid cells (MOLT-4). This study demonstrates that these binding sites have the characteristics of receptors specific for HrHF, and the binding characteristics and internalization of HrHF to MOLT-4 cells have now been examined. Iodinated HrHF was displaced by an excess of unlabeled HrHF. Heavy ferritin was the major subunit bound with only a small amount of light-ferritin binding, consistent with our immunofluorescence studies. Scatchard plot analysis of the competitive binding data for HrHF revealed an association constant of 6.3 to 6.7 x 10(7) L/mol with approximately 6000 to 15,000 receptor sites per MOLT-4 cell. Internalization of HrHF was demonstrated with pronase. Chloroquine substantially reduced the uptake of HrHF. Release of internalized HrHF was not observed when cells were rewarmed to 37 degrees C. These results indicate that HrHF is internalized by a mechanism consistent with receptor-mediated endocytosis, with possible involvement of the lysosome. The internalized HrHF remains associated with the cell. Although lymphoid cell growth and differentiation were not examined in this study, the presence of the demonstrated receptors may indicate a regulatory role for heavy ferritin in such cells.

Binding, Competitive↗
PubMed1992

Effect of cell proliferation on H-ferritin receptor expression in human T lymphoid (MOLT-4) cells.

We have previously demonstrated the presence of receptors specific for human recombinant H ferritin (HrHF) on human T lymphoid cells (MOLT-4), and changes in receptor number and binding affinity with growth and cell cycling have now been examined. Specific binding of HrHF was maximal in MOLT-4 cells harvested during exponential growth with the cells in the DNA synthesis phase of the cell cycle. Specific binding decreased progressively to the plateau phase of growth with the cells in the resting phase of the cell cycle. Scatchard analysis of the competitive binding data for HrHF demonstrated that this decrease in binding was associated with a reduction in receptor number, from 42,140 per cell to 10,306 per cell. Receptor binding affinity increased only minimally over this period, from 7.1 x 10(7) L/mol to 14.9 x 10(7) L/mol. These results indicate that growth- and cell cycle-induced changes in H-ferritin receptor expression are primarily associated with changes in receptor number rather than receptor binding affinity. The present study demonstrates that the expression of this receptor is associated with the proliferative status of the cell and suggests that the H-ferritin receptor may mediate the putative regulatory role of H-ferritin.

Cell Cycle↗
PubMed1991

Fine mapping of a human chromosome 6 ferritin heavy chain pseudogene: relevance to haemochromatosis.

We have used a somatic cell hybrid regional mapping panel for the short arm of chromosome 6, linkage analysis and a population study to map in detail a previously described ferritin heavy chain pseudogene sequence on chromosome 6. Our results show that this sequence maps to the short arm of chromosome 6 centromeric to the glyoxylase locus. The ferritin pseudogene locus is thus distant from the locus for the iron storage disease haemochromatosis, confirming previous evidence that this sequence is not a candidate for the haemochromatosis gene.

Chromosome Mapping↗
PubMed1991

The effect of iron and ethanol on rat hepatocyte collagen synthesis.

1. Carbonyl iron (2.5% w/w) in rat chow was used to induce iron loading in rat hepatocytes. 2. Acute exposure of cultured hepatocytes from control and iron-loaded rats to ethanol (25-100 mM) resulted in a significant inhibition of protein synthesis. 3. Inhibition of protein synthesis in hepatocytes from iron-loaded rats was primarily due to impaired amino acid uptake by these cells. 4. High concentrations of ethanol stimulated the rate of protein degradation by hepatocytes from iron-loaded rats. 5. Acute administration of ethanol to hepatocytes from control animals did not stimulate the absolute rates of collagen biosynthesis nor induce Type I procollagen mRNA. 6. Acute administration of ethanol did not inhibit procollagen synthesis. 7. Iron overload induced Type I procollagen mRNA and increased the absolute rates of collagen synthesis in hepatocytes. 8. These findings may be relevant for the development of hepatic fibrosis in patients with genetic hemochromatosis who consume excess ethanol.

Amino Acids↗
PubMed1991

Intestinal transferrin receptors and iron absorption in the neonatal rat.

The transferrin receptor is a major protein found on the basolateral membranes of intestinal epithelial cells, yet its possible role in intestinal iron metabolism and also in iron absorption is unclear. We have studied intestinal transferrin receptor expression during the peri- and postnatal development of the small intestine of the rat using immunohistochemistry with a monoclonal antibody to the rat receptor. Two major changes in transferrin receptor expression in the developing small intestine were found, a decrease in receptor expression associated with birth, and an increase at the time of weaning. Around the time of weaning there was a large decrease in iron absorption, but there was no direct correlation between absorption and transferrin receptor expression. However, at both birth and weaning there were major changes in intestinal cell kinetics, and the distribution of receptor correlated well with the distribution of proliferating cell populations. In addition, as the intestinal epithelial cells differentiated and stopped dividing, there was a redistribution of transferrin receptors from the cell surface to intracellular sites. These data suggest that the most likely role of the transferrin receptor in the neonatal intestine is in the supply of iron to the developing epithelial cells in the crypts, and that the receptor does not play a direct role in iron transit across the intestinal epithelium.

Aging↗