Routine germline BRCA1 and BRCA2 testing in patients with ovarian carcinoma: analysis of the Scottish real-life experience.

OBJECTIVE: To determine the rates of germline BRCA1 and BRCA2 mutations in Scottish patients with ovarian cancer, before and after a change in testing policy. DESIGN: Retrospective cohort study. SETTING: Four cancer/genetics centres in Scotland. POPULATION: Patients with ovarian cancer undergoing germline BRCA1 and BRCA2 (gBRCA1/2) sequencing before 2013 (under the 'old criteria', with selection based solely on family history), after 2013 (under the 'new criteria', with sequencing offered to newly presenting patients with non-mucinous ovarian cancer), and in the 'prevalent population' (who presented before 2013, but were not eligible for sequencing under the old criteria but were sequenced under the new criteria). METHODS: Clinicopathological and sequence data were collected before and for 18 months after this change in selection criteria. MAIN OUTCOME MEASURES: Frequency of germline BRCA1, BRCA2, RAD51C, and RAD51D mutations. RESULTS: Of 599 patients sequenced, 205, 236, and 158 were in the 'old criteria', 'new criteria', and 'prevalent' populations, respectively. The frequency of gBRCA1/2 mutations was 30.7, 13.1, and 12.7%, respectively. The annual rate of gBRCA1/2 mutation detection was 4.2 before and 20.7 after the policy change. A total of 48% (15/31) 'new criteria' patients with gBRCA1/2 mutations had a Manchester score of <15 and would not have been offered sequencing based on family history criteria. In addition, 20 patients with gBRCA1/2 were identified in the prevalent population. The prevalence of gBRCA1/2 mutations in patients aged >70 years was 8.2%. CONCLUSIONS: Sequencing all patients with non-mucinous ovarian cancer gives a much higher annual gBRCA1/2 mutation detection rate, with the frequency of positive tests still exceeding the 10% threshold upon which many family history-based models operate. TWEETABLE ABSTRACT: BRCA sequencing all non-mucinous cancer patients increases mutation detection five fold.

Refining the primary open-angle glaucoma GLC1C region on chromosome 3 by haplotype analysis.

The GLC1C locus for primary open-angle glaucoma (POAG) is inherited as an autosomal dominant trait. This region on chromosome 3 is 11 cM long. DNA samples from members of a Greek and an American GLC1C family were obtained to determine whether additional typing of microsatellite markers in family members might narrow the region. GLC1C family members were evaluated clinically for POAG on the basis of open angles, intraocular pressures, cupping of discs, and visual fields. DNA samples from the Greek and Oregon GLC1C families were used to further refine the GLC1C region using microsatellite markers. A total of 22 affected members were identified in the two families. Common alleles for D3S3637 and D3S3612 were present in the disease haplotype from both families, suggesting that they may have a common founder. A newly diagnosed patient in the American family had a recombination in the distal portion of the GLC1C haplotype. This recombination narrows the GLC1C region from 11 to 4 cM.

Proximal promoter of the surfactant protein D gene: regulatory roles of AP-1, forkhead box, and GT box binding proteins.

Surfactant protein D (SP-D) plays roles in pulmonary host defense and surfactant homeostasis and is increased following lung injury. Because AP-1 proteins regulate cellular responses to diverse environmental stimuli, we hypothesized that the conserved AP-1 motif (at -109) and flanking sequences in the human SP-D promoter contribute to the regulation of SP-D expression. The AP-1 sequence specifically bound to fra-1, junD, and junB in H441 lung adenocarcinoma nuclear extracts. Mutagenesis of the AP-1 motif in a chloramphenicol acetyltransferase reporter construct containing 285 base pairs of upstream sequence nearly abolished promoter activity, and co-transfection of junD significantly increased wild type but not mutant promoter activity. The sequence immediately downstream of the AP-1 element contained a binding site for HNF-3 (FOXA), and simultaneous mutation of this site (fox-d) and an upstream FoxA binding site (-277, fox-u) caused a 4-fold reduction in chloramphenicol acetyltransferase activity. Immediately upstream of the AP-1-binding site, we identified a GT box-containing positive regulatory element. Despite finding regions of limited homology to the thyroid transcription factor 1-binding site, SP-D promoter activity did not require thyroid transcription factor 1. Thus, transcriptional regulation of SP-D gene expression involves complex interactions with ubiquitous and lineage-dependent factors consistent with more generalized roles in innate immunity.

The role of computed tomography scans in evaluating sinus disease in pediatric patients.

OBJECTIVES: To determine the incidence and severity of sinus abnormalities in children undergoing computed tomography (CT) of the sinuses for suspected chronic sinusitis. To compare these findings with abnormalities noted on random CT scans. METHODS: Sixty CT scans, performed for evaluation of sinus disease in symptomatic children aged 2-12, were compared with 50 CT scans of children aged 2-12 of the orbits or sinuses obtained for indications other than sinusitis. A staging system was applied to assess the severity of abnormalities. RESULTS: Mucoperiosteal thickening was present in 60% of symptomatic and 46%, of random CT scans (logistic regression, P = 0.144). Children aged 2-4 and 9-12 had an increased prevalence of abnormalities in both groups, although these findings were not statistically significant (logistic regression, P = 0.817). Early stage sinus disease was present in the majority of random (96%) and symptomatic (85%) children. CONCLUSIONS: There is a high incidence of mucoperiosteal thickening in the paranasal sinuses of children. CT scans of the sinuses should be obtained from children who are being considered for sinus surgery after failing the appropriate medical therapy. Decisions regarding the need for sinus surgery should not be solely based on imaging abnormalities.

GLC1F, a new primary open-angle glaucoma locus, maps to 7q35-q36.

BACKGROUND: A large family with adult-onset primary open-angle glaucoma (POAG) was identified. OBJECTIVE: To initiate a genome-wide scan to map the POAG locus in this family. METHODS: Blood samples or buccal swabs were obtained from 25 members of a large family with POAG after informed consent was obtained. Members and their spouses were evaluated clinically for POAG on the basis of intraocular pressures, cupping of discs, and visual fields. DNA samples were used for a genome-wide screen using microsatellite markers. RESULTS: Ten affected family members in 4 generations showed evidence of POAG including intraocular pressures of 22 mm Hg or more, and/or optic cup-disc ratios of 0.6 or more, and/or visual field defects consistent with glaucomatous damage. Primary open-angle glaucoma segregated as an autosomal dominant trait, with the disease locus mapping to 7q35-q36 between markers D7S2442 and D7S483 with a multipoint lod score of 4.06. CONCLUSION: A sixth gene for POAG (GLC1F) has been mapped to 7q35-q36 in a family with at least 4 generations affected. CLINICAL RELEVANCE: The mapping of this locus further confirms that primary open-angle glaucoma is a heterogeneous group of diseases with at least 6 different loci resulting in a similar phenotype. The eventual ability to classify which major POAG gene an affected person carries could have ramifications for selecting the most effective treatment regimen for that person.

Malattia leventinese: refinement of the genetic locus and phenotypic variability in autosomal dominant macular drusen.

PURPOSE: To study the phenotypic variability in patients inheriting the disease gene for malattia leventinese (dominant macular drusen) and refine the localization of the gene. METHODS: A family with dominant radial drusen was ascertained and studied with clinical examination and DNA linkage analysis. Inheritance of the disease gene was determined by DNA analysis and used to document the variability in phenotypic expression. RESULTS: Fifty family members were studied with fundus photography and genotyping. Linkage analysis showed that the disease in this family was linked to chromosome 2p16-21 with a maximum lod score of 3.72 at D2S2153. An affected patient with obligate recombinations allowed refinement of the disease interval to a 6.2-cM region between D2S2227 and D2S378. The phenotype of older affected patients varied from severe geographic atrophy or subretinal fibrosis to a single druse adjacent to the optic disk. Small and medium-sized, nonradial, and soft macular drusen seen in four older individuals in the family were not specifically associated with the disease haplotype. CONCLUSIONS: Refinement of the localization of the gene for malattia leventinese will facilitate its positional cloning. Genotypic documentation of the variable expression of the disease shows that a single, large, subretinal druse adjacent to the optic disk is consistent with inheritance of the disease gene. Soft macular drusen in low abundance were not specifically associated with inheritance of the disease gene. These results will facilitate the genetic counseling of patients with malattia leventinese. It is unknown what proportion of age-related macular degeneration arises from mutations in disease genes for dominant drusen.

Age-related macular degeneration. Clinical features in a large family and linkage to chromosome 1q.

OBJECTIVES: To identify the chromosomal location of a disease-causing gene and to describe the clinical characteristics of a large family with age-related macular degeneration (ARMD). METHODS: An ARMD pedigree was identified, and the disease state of family members was documented by stereoscopic fundus photography and was classified using a modified version of the Wisconsin Age-Related Maculopathy Grading System. A genome-wide screen at approximately 6-centimorgan spacing using a DNA-pooling strategy combined with shared-segment analysis was used to identify likely chromosomal regions. The entire family was then screened at each likely locus, and 1 positive locus was refined by screening with markers at an average density of 0.5 centimorgan and subjected to parametric linkage analysis. RESULTS: In the 10 affected family members, ARMD was manifest by the presence of large, soft, confluent drusen accompanied by varying degrees of retinal pigment epithelial degeneration and/or geographic atrophy. Age-related macular degeneration segregated as an autosomal-dominant trait, with the disease locus mapping to chromosome 1q25-q31 between markers D1S466 and D1S413, with a multipoint lod score of 3.00. CONCLUSION: Age-related macular degeneration localized to chromosome 1q25-q31 (gene symbol, ARMD1) as a dominant trait in a large family with a predominantly dry phenotype. CLINICAL RELEVANCE: Identification of ARMD genes will facilitate early diagnosis and aid in understanding the molecular pathophysiological mechanisms of ARMD. This knowledge will contribute to the development of preventive and improved treatment strategies.

Insulin-like growth factor binding protein-5 expression by human trabecular meshwork.

PURPOSE: The authors found transcript expression for insulin-like growth factor binding protein-5 (IGFBP-5) while screening for uniquely expressed trabecular meshwork (TM) mRNAs. Because the insulin-like growth factor (IGF) autocrine-paracrine system may provide an important signaling mechanism between TM cells and the outflow pathway, the expression of IGFBP-5 and IGF-I receptor in the TM was characterized. METHODS: Poly(A+) RNA was isolated from cell cultures of human TM, ciliary body, retinal pigment epithelium, and skin fibroblasts and subjected to reverse transcription-polymerase chain reaction (RT-PCR) differential display analysis. A unique 980-bp band present in the TM was cloned and sequenced. Additional PCR and Northern analyses were used to define trabecular IGFBP-5 expression. Western immunoblots and confocal immunohistochemistry were used to evaluate the protein expression patterns of IGFBP-5 and the IGF-I receptor. IGF-I and IGF-II were added to trabecular cells in culture, and matrix metalloproteinase production was evaluated. RESULTS: A unique differential display band was identified in the TM. Sequencing of this band identified it as the 3'-untranslated region of IGFBP-5. RT-PCR, using a variety of specific primers for IGFBP-5, Northern analysis, Western immunoblots, and immunohistochemical analysis, confirmed that IGFBP-5 was expressed in the TM. However, IGFBP-5 was also present at low levels in the ciliary body and skin fibroblasts by Northern and Western analysis, in contrast with the differential display findings. In addition, the IGF-I receptor was expressed by the TM and showed cell-surface staining by immunohistochemistry. Trabecular IGFBP-5 was distributed throughout the meshwork in the extracellular matrix and the cells with more staining in the juxtacanalicular region than in the uveal meshwork. IGF-I, but not IGF-II, modestly increased trabecular stromelysin and gelatinase B but not collagenase, gelatinase A, or tissue inhibitor of metalloproteinases 1 or 2. CONCLUSIONS: IGFBP-5 and IGF-I receptor were expressed at significant levels by TM cells and may serve an important role in trabecular function.

Mapping a gene for adult-onset primary open-angle glaucoma to chromosome 3q.

Glaucoma is the third-leading cause of blindness in the world, affecting >13.5 million people. Adult-onset primary open-angle glaucoma (POAG) is the most common form of glaucoma in the United States. We present a family in which adult-onset POAG is inherited as an autosomal dominant trait. Twelve affected family members were identified from 44 at-risk individuals. The disease-causing gene was mapped to chromosome 3q21-24, with analysis of recombinant haplotypes suggesting a total inclusion region of 11.1 cM between markers D3S3637 and D3S1744. This is the first report of mapping of an adult-onset POAG gene to chromosome 3q, gene symbol GLC1C.

The effect of absorbable stenting on postoperative stenosis of the surgically enlarged maxillary sinus ostia in a rabbit animal model.

OBJECTIVE: To determine the effect of absorbable stenting on postoperative stenosis of the surgically enlarged maxillary sinus in a rabbit model. DESIGN: A randomized controlled animal study with each specimen serving as its own control. Animals had their maxillary ostia surgically enlarged bilaterally. Video images were made of each enlarged ostia. Unilateral stenting was performed using rolled absorbable stenting (Gelfilm, Upjohn Co, Kalamazoo, Mich). The animals were killed at 3 months and repeated images were made of each ostia. An image processing system (IBAS-AT, Kontron Instruments, Milan, Italy) was used to individually calibrate each image and then measure areas. Surgical and 3-month areas were compared with the examiner blinded to the stented side. SUBJECTS: Twelve specific pathogen-free New Zealand white rabbits. RESULTS: Stented ostia had an increased stenosis compared with unstented ostia, but this increase was not statistically significant (P = .08, Student t test). There was no within-animal change in nonstented ostia compared with an 18% decrease in the stented ostia. CONCLUSIONS: Stenting did not statistically change the amount of stenosis of the surgically enlarged ostia in a rabbit model, although a trend toward more scarring when compared with the nonstented side was observed.

Interactions of recombinant human pulmonary surfactant protein D and SP-D multimers with influenza A.

To further study the structure and function of surfactant protein D (SP-D), recombinant human SP-D (rhSP-D) was isolated from the culture medium of Chinese hamster ovary (CHO)-K1 cells stably transfected with a full-length hSP-D cDNA. Although a significant fraction of the secreted rhSP-D was recovered as dodecamers similar to recombinant rat SP-D (rrSP-D), a major fraction accumulated as multimers of dodecamers indistinguishable from human proteinosis SP-D. As previously shown for the rat protein, rhSP-D agglutinated specific strains of influenza A virus (IAV), inhibited viral hemagglutinin activity, and protected neutrophils (PMN) from deactivation by IAV. However, the potency of rhSP-D multimers was severalfold greater than for purified dodecamers, comparable to natural, proteinosis hSP-D. Although rhSP-D multimers were also more potent than the serum collectins in mediating viral aggregation and protection of PMN, they were less potent than conglutinin in inhibiting infectivity in vitro. These studies establish that the propensity of hSP-D to form multimers of dodecamers is determined by its primary structure and demonstrate carbohydrate recognition domain valency-dependent interactions of SP-D with IAV.

Racial disparities in outcomes of military and civilian births in California.

OBJECTIVE: To examine racial disparities in prenatal care utilization, birth weight, and fetal and neonatal mortality in a population for whom financial barriers to health care services are minimal. STUDY-DESIGN: Using linked birth, fetal death, and infant death certificate files, we examined prenatal care utilization, birth weight distribution, and fetal and neonatal mortality rates for all white and black births occurring in military hospitals in California from January 1, 1981, to December 31, 1985. These patterns were compared with the experience of their civilian counterparts during the same time period. RESULTS: Black mothers had higher percentages of births occurring in teenaged and unmarried mothers than did white mothers in military and civilian populations. First-trimester prenatal care initiation was lower for blacks in the military (relative risk, 0.79; 95% confidence interval, 0.75-0.82) and civilian (relative risk, 0.51; 95% confidence interval, 0.50-0.52) populations. However, the scale of the disparity in prenatal care utilization was significantly smaller (P < .001) in the military group. Rates of low birth weight and fetal and neonatal mortality among blacks were elevated in the military and civilian groups. However, the racial disparity in low birth weight was significantly smaller in the military group (P < .01 and P < .001, respectively). CONCLUSIONS: In populations with decreased financial barriers to health care, racial disparities in prenatal care use and low birth weight were reduced. However, the persistence of significant disparities suggests that more comprehensive strategies will be required to ensure equity in birth and neonatal outcome.

Weill-Marchesani syndrome--possible linkage of the autosomal dominant form to 15q21.1.

Weill-Marchesani syndrome comprises short stature, brachydactyly, microspherophakia, glaucoma, and ectopia lentis is regarded as an autosomal recessive trait (McKusick 277600). We present two families each with affected individuals in 3 generations demonstrating autosomal dominant inheritance of Weill-Marchesani syndrome. Linkage analysis in these 2 families suggests a gene for Weill-Marchesani syndrome maps to 15q21.1. The dislocated lenses and connective tissue disorder in these families suggests that fibrillin-1 and microfibril-associated protein 1, which both map to 15q21.1, are candidate genes for Weill-Marchesani syndrome. Immunohistochemistry staining of skin sections from family 1 showed an apparent decrease in fibrillin staining compared to control individuals.

Variance estimation for complex surveys using replication techniques.

The analysis of survey data requires the application of special methods to deal appropriately with the effects of the sample design on the properties of estimators and test statistics. The class of replication techniques represents one approach to handling this problem. This paper discusses the use of these techniques for estimating sampling variances, and the use of such variance estimates in drawing inferences from survey data. The techniques of the jackknife, balanced repeated replication (balanced half-samples), and the bootstrap are described, and the properties of these methods are summarized. Several examples from the literature of the use of replication in analysing large complex surveys are outlined.

Biosynthesis of surfactant protein D. Contributions of conserved NH2-terminal cysteine residues and collagen helix formation to assembly and secretion.

Surfactant protein D (SP-D) is preferentially secreted as dodecamers consisting of four collagenous trimers cross-linked by disulfide bonds. In these studies, we examined the biosynthesis of wild-type rat SP-D (RrSP-D) and selected mutants by stably transfected CHO-K1 cells to determine the roles of a conserved N-linked oligosaccharide, the collagen helix, and interchain disulfide bonds in SP-D assembly and secretion. The major intracellular form of RrSP-D accumulated in the RER as complexes containing up to four trimeric subunits. Disulfide cross-link formation and RrSP-D secretion were selectively inhibited by 2,2'-dipyridyl, an inhibitor of prolyl and lysyl hydroxylase, and by 2 mM dithiothreitol, but unaffected by tunicamycin or elimination of the consensus sequence for glycosylation at Asn70. Although mutants with serine substituted for Cys15 and Cys20 (RrSP-Dser15/20) are secreted as trimeric subunits, proteins with single cysteine substitutions were retained in the cell. Surprisingly, the secretion of RrSP-Dser15/20 was unaffected by 2,2'-dipyridyl. These studies strongly suggest that the most important and rate-limiting step for the secretion of SP-D involves the association of cross-linked trimeric subunits to form dodecamers stabilized by specific inter-subunit disulfide cross-links. Interference with collagen helix formation prevents secretion by interfering with efficient disulfide cross-linking of the NH2-terminal domain.

Site-directed mutagenesis of Cys-15 and Cys-20 of pulmonary surfactant protein D. Expression of a trimeric protein with altered anti-viral properties.

Surfactant protein D (SP-D) molecules are preferentially assembled as dodecamers consisting of trimeric subunits associated at their amino termini. The NH2-terminal sequence of each monomer contains two conserved cysteine residues, which participate in interchain disulfide bonds. In order to study the roles of these residues in SP-D assembly and function, we employed site-directed mutagenesis to substitute serine for cysteine 15 and 20 in recombinant rat SP-D (RrSP-D), and have expressed the mutant (RrSP-Dser15/20) in Chinese hamster ovary (CHO-K1) cells. The mutant, which was efficiently secreted, bound to maltosyl-agarose, but unlike RrSP-D, was assembled exclusively as trimers. The constituent monomers showed a decreased mobility on SDS-polyacrylamide gel electrophoresis resulting from an increase in the size and sialylation of the N-linked oligosaccharide at Asn-70. Although RrSP-Dser15/20 contained a pepsin-resistant triple helical domain, it showed a decreased Tm, and acquired susceptibility to proteolytic degradation. Like RrSP-D, RrSP-Dser15/20 bound to the hemagglutinin of influenza A. However, it showed no viral aggregation and did not enhance the binding of influenza A to neutrophils (PMN), augment PMN respiratory burst, or protect PMNs from deactivation. These studies indicate that amino-terminal disulfides are required to stabilize dodecamers, and support our hypothesis that the oligomerization of trimeric subunits contributes to the anti-microbial properties of SP-D.

Bioglass middle ear prosthesis: long-term results.

The purpose of this study was to review the University of Florida's long-term results with Bioglass middle ear prostheses. Between April 1984 and November 1987, 37 patients were implanted with Bioglass prostheses (25 total and 12 partial ossicular replacements). Twenty-one patients had postoperative data of at least 24 months (range, 24 to 126 months; mean, 86 months; median, 100 months), and five patients had > 10 years' follow-up. In three cases, portions of fractured prostheses extruded, leaving an intact tympanic membrane. One patient with a total ossicular prosthesis was reexplored at 38 months for conductive hearing loss and found to have a prosthesis fracture (n = 1). There were no extrusions of intact prostheses, even in patients in whom the prosthesis was placed directly under the tympanic membrane or graft (n = 12). After 24 months, the mean pure-tone average air-bone gap was 24 dB (24% had ABG < or = 10 dB; 53% had ABG < or = 20 dB). Air-bone gap closures were stable over time. Our results demonstrated that Bioglass middle ear prostheses have excellent long-term tissue compatibility. The four failures are attributed to fractures in early experimental prototypes.

Characterization of the human surfactant protein D promoter: transcriptional regulation of SP-D gene expression by glucocorticoids.

We have previously described the characterization of genomic clones encoding the entire translated sequence of human pulmonary surfactant protein D (SP-D). We now describe the characterization of a genomic fragment (H5E7) that encodes the entirety of the first translated exon (Exon 2), Intron 1, a short transcribed untranslated sequence (Exon 1; 39 bp), and approximately 4 kb of sequence upstream from the transcription initiation site. The start site was identified by 5'-RACE-PCR cloning and primer extension. A putative TATA box (CATAAATA) was identified approximately 30 bp upstream of the start site. Complete sequencing of a HindIII/SacI fragment (HS-1674) encoding approximately 1.7 kb of sequence 5' to the TATA demonstrated multiple potential cis-regulatory elements including half-site glucocorticoid response elements (GRE), a canonical AP-1 consensus, several AP-1 like sequences, E-box sequences, NF-IL-6 and PEA3 motifs, and putative interferon response elements. H441 lung adenocarcinoma cells, which express low levels of SP-D mRNA, and liver HepG2 cells, were transiently co-transfected with chloramphenicol acetyl transferase (CAT) reporter constructs containing up to 3,000 base pairs of upstream sequence, and with constructs encoding beta-gal. H441 cells transfected with constructs containing at least 161 bp of upstream sequence gave normalized levels of CAT activity greater than or equal to that obtained for parallel positive control transfections using pTK-CAT. Treatment of the cells for 48 h with 50 nM dexamethasone (Dex) gave a 2- to 5-fold increase in CAT activity. Interestingly, a 5'-deletion construct containing 161 bp of upstream sequence (pFS161-CAT) conferred both cell type-restricted and dexamethasone-responsive expression. These studies emphasize the potential complexity of SP-D gene regulation, and further support the hypothesis that the effects of glucocorticoids on SP-D production in vivo are regulated at the level of transcription.

Differences in survival between black and white patients with diabetic end-stage renal disease.

OBJECTIVE: To evaluate whether the longer survival of blacks with diabetic end-stage renal disease (ESRD) relative to whites is due to racial differences in type of diabetes, comorbidity at ESRD onset, and ESRD treatment modality and to examine whether survival differences between blacks and whites occur only in certain population subgroups. RESEARCH DESIGN AND METHODS: The Michigan Kidney Registry was used to ascertain all blacks and whites (n = 594) with diabetic ESRD in southeastern Michigan, with ESRD onset at age < 65 years during 1974-1983. Patients were followed through 1988. Medical records were abstracted for type of diabetes, comorbidity at ESRD onset, and other factors. RESULTS: Median survival among insulin-dependent diabetes mellitus patients was 27 months in blacks and 17 months in whites, and among non-insulin-dependent diabetes mellitus patients was 30 months in blacks and 16 months in whites. After adjustment for confounding factors by Cox proportional hazards analysis, the death rate was 45% lower in blacks than in whites on dialysis (relative death rate [RDR] = 0.55, 95% confidence interval [CI] = 0.44-0.69), but was similar in blacks and whites with a renal transplant (RDR = 0.99, 95% CI = 0.64-1.52). Compared with dialysis, transplantation was associated with lower mortality in both races (white, RDR = 0.50, 95% CI = 0.36-0.70; blacks, RDR = 0.89, 95% CI = 0.60-1.34), although the effect was not statistically significant in blacks. Racial differences in survival did not vary by type of diabetes or any additional factor. CONCLUSIONS: Survival after ESRD onset is longer in blacks than in whites treated with dialysis, even after adjusting for comorbidity and other factors that affect survival. Survival does not differ by race among transplant patients.

Recombinant pulmonary surfactant protein D. Post-translational modification and molecular assembly.

Pulmonary surfactant protein D (SP-D) is a member of a family of collagenous C-type lectins that includes the serum mannose binding proteins and surfactant protein A. Recent studies have shown that rat SP-D (rSP-D) molecules are assembled as tetramers of trimeric subunits (12 mers) and that dodecamers can participate in higher orders of molecular assembly involving interactions of the amino-terminal peptide domains. In order to further study the assembly of SP-D in vitro, Chinese hamster ovary K1 cells were transfected with a full-length rat SP-D cDNA, and stable transfectants with high levels of SP-D production (approximately 6 x 10(6) dodecamers/cell/24 h) were obtained using a glutamine synthetase selection system. The secreted molecules (RrSP-D), which were purified by affinity chromatography on maltosyl-agarose, comigrated with rSP-D on SDS-polyacrylamide gel electrophoresis in the presence and absence of reduction, and coeluted with rSP-D dodecamers from 4% agarose. The major bacterial collagenase-resistant peptide showed a decreased mobility on reduction consistent with the formation of intrachain disulfide bonds. A 17-kDa pepsin-resistant fragment was isolated following overnight digestion with pepsin at 27 degrees C, confirming the formation of a triple helical domain comparable in size and thermal stability to that of natural SP-D. The expressed protein contained sialylated endoglycosidase F-sensitive carbohydrate; amino acid analysis of acid and alkaline hydrolysates demonstrated essentially normal levels of hydroxyproline, hydroxylysine, and hydroxylysine-glycosides. Electron microscopic studies showed a molecular structure indistinguishable from lung SP-D, with a similar small subpopulation of molecules showing higher orders of multimerization. Solid-phase neoglycoprotein binding assays gave the same saccharide inhibition profile as natural rat SP-D, and both proteins showed efficient saccharide-dependent agglutination of Escherichia coli. These studies demonstrate that a single genetically distinct chain type can account for the various and complex molecular assemblies of SP-D, and further verify the potential physiologic significance of the disulfide-bonded multimers and higher aggregates isolated from rat, bovine, and human lung lavage.