Twenty-four novel mutations in Wilson disease patients of predominantly European ancestry.

Wilson disease (WND), an autosomal recessive disorder of copper transport, shows wide genotypic and phenotypic variability, with hepatic and/or neurological symptoms. The WND gene, ATP7B, encodes a copper transporting ATPase that is involved in the transport of copper into the plasma protein ceruloplasmin, and in the excretion of copper from the liver. ATP7B mutations result in copper storage in liver and brain. From 247 WND patients worldwide whose DNA has been sequenced in our laboratory, we have identified 24 new mutations. The origins of the patients were European white (one deletion, one nonsense, one splice site, and 18 missense), Chinese (one deletion, one missense) and Bangladeshi (one missense). Most of these had strong support as disease causing mutations, based on conservation between species, structural changes, and absence in controls. One missense mutation in a Chinese patient was considered uncertain because of its conservative nature and position in the protein. We also identified 15 nucleotide substitutions (11 of them new) causing silent or intronic changes, none of which produce an additional splice site that could lead to disease. Characterization of mutations, both disease-causing and normal variants, is essential for accurate molecular diagnosis of this condition.

Genetic variation in the promoter and 5' UTR of the copper transporter, ATP7B, in patients with Wilson disease.

ATP7B is a copper-transporting P-type ATPase defective in the copper transport disorder, Wilson disease (WND). We have sequenced the 5' UTR and promoter region of ATP7B in 37 unrelated WND patients in whom partial sequencing of the coding region and intron/exon boundaries of the gene had failed to identify one or both disease-causing mutations. Three patients were found to be heterozygous for a 15 bp deletion between nucleotides -424 and -441. This deletion had been previously identified as the most common mutation in Sardinian WND patients. Two novel single-nucleotide changes were also identified within the 5' UTR and promoter of ATP7B; however, these were found at a similar frequency in control chromosomes and are apparently normal variants. These results suggest that mutations in regulatory elements of ATP7B are uncommon in patients of European ancestry, except in Sardinia.

Noninvasive localization of human atherosclerotic lesions with indium 111-labeled monoclonal Z2D3 antibody specific for proliferating smooth muscle cells.

BACKGROUND: Targeting exclusive antigens in atherosclerotic plaques with antibodies may provide a noninvasive means to detect rapidly proliferative atherosclerotic lesions. 111In-labeled negative charge-modified Z2D3 F(ab')2 (Z2D3) specific for an antigen expressed exclusively by proliferating smooth muscle cells has been shown to accumulate in rabbit atherosclerotic plaques. METHODS: The safety, biodistribution, accumulation, and elimination of Z2D3 were assessed in 11 patients who were candidates for carotid endarterectomy. The presence of atheromas in these patients was confirmed by angiography and Doppler ultrasound. Z2D3 (250 microg) labeled with 5 mCi of 111In was administered by slow intravenous injection. Planar and single photon emission computed tomography (SPECT) images were obtained 4, 24, 48, and 72 hours later. Carotid endarterectomy was performed and the surgical specimens were imaged, weighed, gamma-counted, and analyzed by immunostaining. RESULTS: Uptake of Z2D3 at the site of the carotid plaques was observed in the planar and SPECT views at 4 hours in all subjects. In addition, antibody uptake was noted in the contralateral vessel in 5 subjects. SPECT images identified the atherosclerotic plaques with focal uptake. The antibody uptake corresponded with the angiographic location of the disease. Immunohistochemical studies of the endarterectomy specimens confirmed the localization of Z2D3 into the plaque areas containing smooth muscle cells. Adverse drug reactions were not observed. CONCLUSION: This study demonstrates the feasibility of targeting atherosclerotic lesions with negative charge-modified antibody. It also proposes the possibility of selective identification of various components of atherosclerotic plaque, which may contribute to determining strategies of intervention in future.

99Tcm-MAG3 renogram deconvolution in normal subjects and in normal functioning kidney grafts.

This study provides values of transit times obtained by 99Tcm- mercaptoacetyl triglycine (99Tcm-MAG3) renogram deconvolution for both normal subjects and kidney graft recipients. The analysis included 50 healthy kidney units from 25 volunteers and 28 normal functioning kidney grafts. The parameters calculated for the whole kidney (WK) and for the renal parenchyma (P) were: mean transit time (MTT) and times at 20% (T20) and 80% (T80) of renal retention function initial height. For healthy kidneys the WK MTT was 174 +/- 27 s and P MTT 148 +/- 22 s. The WK T20 values were 230 +/- 33 s and P T20 231 +/- 34 s. The WK T80 was 108 +/- 19 s and P T80 106 +/- 12 s. Whole kidney and parenchymal values of transit times for normal functioning kidney grafts do not present significant differences with respect to healthy kidneys.

Polyclonal 111In-IgG, 125I-LDL and 125I-endothelin-1 accumulation in experimental arterial wall injury.

To test iodine-125 labelled low-density lipoprotein (125I-LDL), polyclonal indium-111 labelled immunoglobulin G (111In-IgG) and iodine-125 labelled endothelin-1 uptake in metabolically active atheromatous plaques after arterial wall injury, we performed balloon de-endothelialization of carotid arteries or abdominal aortas in 24 New Zealand male rabbits which were fed with a normal diet (n = 14) or a hypercholesterolaemic diet (n = 10) after surgery. Six weeks later the animals were injected with 200 microCi of 125I-LDL and/or with 100 microCi of 111In-IgG or with 9 microCi of 125I-endothelin-1. Forty-eight hours later the animals were sacrificed. Carotid arteries and aortas were removed, counted and fixed for autoradiography and light microscopy examination. Contralateral carotid arteries and thoracic aortas served as controls. Significant 111In-IgG uptake was observed in the injured arteries at autoradiography, with localization mainly in the healing edges, and at well counting. The percentage of the injected dose per gram (%D.inj/g) was 0.0188 +/- 0.06 versus 0.0059 +/- 0.003 in controls (P < 0.05). There was no difference in 111In-IgG uptake between arteries with injury alone and those with active atheroma formation at the site of the injury. Significant 125I-LDL uptake was observed only when lipid deposition was present at light microscopy (%D.inj/g of 0.0024 +/- 0.0005 vs. 0.0010 +/- 0.0003 in controls, P < 0.05). 125I-endothelin-1 accumulation was observed in four of five injured aortas both at autoradiography, with diffuse localization, and at well counting (%D.inj/g of 0.0012 +/- 0.0004 in the abdominal aortas vs 0.0008 +/- 0.0003 in the thoracic aortas).(ABSTRACT TRUNCATED AT 250 WORDS)