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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.

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.

The Marfan syndrome is a serious heritable connective-tissue disorder characterized primarily by ocular, cardiovascular, and musculoskeletal abnormalities but also involving multiple other tissues and organs of the body. Inherited as an autosomal dominant disorder, the etiology and pathogenesis of the Marfan syndrome are presently unknown. We have documented consistent apparent deficient content of elastin-associated microfibrillar fibers by indirect immunofluorescent (IF) studies of Marfan skin, as well as deficient accumulation of related fibrous materials in cultures of Marfan fibroblasts as compared with normal controls and patients with other heritable disorders of connective tissue. These data have suggested that abnormalities in the microfibrillar component of elastic-fiber systems may have a role in the etiology and pathogenesis of the Marfan syndrome. In the present study, we have analyzed the IF staining patterns of skin and fibroblast cultures from Marfan syndrome patients and normal first-degree relatives in nine Marfan kindreds. Three of these families had at least one affected individual in each of 2 generations, permitting intergenerational comparison of IF patterns. Six kindreds had one or more affected individuals in a single generation, making comparisons between siblings and/or parent-child possible. In all cases, IF abnormalities cosegregated with the Marfan phenotype and all nonaffected family members were normal. Within family groups containing more than one affected individual, the IF staining patterns were similar between affected patients. These data provide further confirmation of consistent and relatively specific deficiency of microfibrillar fibers in Marfan syndrome.

Eight patients developed the preleukemic syndrome after having been exposed to cytotoxic drugs for other primary diseases. All eight subsequently developed acute nonlymphocytic leukemia (ANLL). Survival following the onset of the preleukemic syndrome ranged from 5-34 months with a median of 11 months. Once overt leukemia developed median survival was three months. No patient responded to conventional antileukemic therapy. A higher incidence of chromosomal abnormalities was noted in bone marrow cells of this group (five of six) when compared with a larger group of preleukemic patients not known to have been exposed to mutagens (four of 21). In addition, two patients had second chromosome studies after overt leukemia developed and there was no evidence of cytogenetic clonal evolution. The clinical course and cytogenetic data in these patients attest to the relevance of our criteria in identifying the preleukemic syndrome and suggest that the leukemic clone was fully established during the preleukemic phase. Thus, patients previously exposed to cytotoxic therapy who develop the preleukemic syndrome may be viewed as having early leukemia.

Each of three families of northern European origin contains a mentally retarded son with hemoglobin H (Hb H) disease. One parent is a carrier of mild alpha-thalassemia and the other is normal, suggesting that this form of Hb H disease results from the interaction between an inherited defect of alpha-chain production and one member of the pair in chromosome 16 and a new mutation on the other. Restriction-enzyme analysis indicated that the new mutation was not the same in the other three patients, and demonstrated at least two hitherto undescribed lesions involving the alpha-globin gene cluster. Unless the association between the Hb H disease and mental retardation is fortuitous, the new mutations may also be related to the development changes in these children. Since the mutations only came to light because there was concurrent inheritance of an additional alpha-thalassemia determinant, this type of mutation of chromosome 16 may have been overlooked in other mentally retarded patients.

A hemoglobin variant with the same electrophoretic mobility as hemoglobin S was found in three generations of a black family. No clinical symptoms or findings were present in subjects heterozygous for this mutant. Except for target forms of mature erythrocytes, they have no abnormal hematologic findings. Structural studies demonstrated a previously undescribed substitution, beta51 Pro replaced by Arg, in the abnormal fraction which accounts for about one-third of the total hemoglobin. This fraction is more unstable in vitro at 65 degrees than normal A hemoglobin. Both whole blood and purified abnormal hemoglobin have increased oxygen affinity and a slightly decreased Bohr effect.

Hemoglobin Chico was discovered in an asymptomatic 3-year-old boy when a mild anemia was detected by a routine blood count. Affected individuals in three generations are also mildly anemic. The abnormal hemoglobin amounts to about 45% of the total. It separates from Hb A by cellulose acetate electrophoresis at pH 8.5 with a mobility similar to Hb J but does not separate in citrate agar at pH 6.2. Stability in isopropanol is slightly decreased. Its structure differs from the normal by the substitution of a threonyl residue for lysyl residue at position 66(E10) of the beta chain. The P50 of the oxygen equilibrium curve of whole blood at 37 degrees C was 38 torr compared with controls of 27 +/- 2 torr. The P50 binding studies of the isolated Hb Chico revealed a unique right shift of the equilibrium curve with an oxygen binding constant (1/P50) about half of normal. The remaining allosteric properties were essentially normal. This significant decrease in oxygen affinity appears to be due to changes in the heme region which result from the substitution of the normal beta 66 lysyl by the threonyl residue.

A child, trisomic for the distal short arm of chromosome 2 due to a familial 2/18 translocation, has elevated levels of activity of erythrocyte acid phosphatase [orthophosphoric-monoester phosphohydrolase (acid optimum), 3.1.3.2] Ferguson-Smith et al. [(1973) Nature New Biol. 243, 271-274] previously had found decreased levels of activity and loss of expression of an erythrocyte acid phosphatase allele in a subject who lacked one of the two homologous regions containing the distal three bands of chromosome 2. They suggested that the locus for erythrocyte acid phosphatase is located on that segment. Our findings provide further evidence for this assignment and also suggest an in vivo gene dosage effect of this autosomal locus, which depends on both the type and number of alleles present.

Blood hemoglobin concentration decreases during the first postnatal month of canine life. Red cell production, as indicated by reticulocyte levels, decreases following birth and does not increase until the second postnatal month. Bleeding and transfusion studies were performed to determine if the canine erythropoietic system is defective during this early neonatal period or is operating at a less than maximal rate due to adequate oxygen delivery to those tissues which regulate erythropoiesis. There is no fundamental defect in the erythropoietic system since bleeding stimulated reticulocyte formation during the first postnatal month. Conversely, the reticulocytosis that normally occurs during the second postnatal month was suppressed when oxygen delivery was increased by a red cell transfusion. We conclude that adequate oxygen delivery following birth causes a reduction in red cell production and results in postnatal anemia. Due to increasing metabolic oxygen requirements of continued growth, the blood oxygen delivery becomes inadequate by the second postnatal month and red cell production is stimulated.

The specific activity of argininosuccinate synthetase (micromoles of 14CO2 per milligram of protein per hour) was 0.00104 and 0.00087 in fibroblasts derived from two patients with citrullinemia, and was undetectable in both fibroblasts and cultured lymphocytes from a third patient. In five obligate heterozygotes the specific activity in fibroblasts was 0.012-0.029 and in nine control subjects was 0.058 +/- 0.014 (0.030-0.076). In both control and patient cells, the maximum activity was obtained at pH 8.5 and there was no inhibition of normal argininosuccinate synthetase by any of the mutant cells.

We have studied the red cell pyruvate kinase (PK) variants from eight patients representing five families with pyruvate kinase deficiency-associated hemolytic anemia. The kinetic properties, electrophoretic mobilities, and immunological reactivity with anti-normal red cell pyruvate kinase were determined. The patients differ in the severity of their clinical condition and in the molecular properties of their red cell pyruvate kinase variants. The most seriously affected patient (PK Beaverton) has no electrophoretically demonstrable red cell isozymes. The activity present is due to the M2 isozyme, however red cell isozyme can be detected immunologically. PK Molalla and PK Lake Oswego are thermolabile variants with normal kinetic parameters. PK Molalla, in addition, has altered electrophoretic mobility. PK Multnomah and PK Milwaukie have decreased affinity for the substrate phosphoenolpyruvate, and PK Multnomah also has altered electrophoretic mobility. PK Coos Bay shows electrophoretic variation and a slightly decreased affinity for phosphoenolpyruvate consistent with an increased modulating effect of fructose-1,6-diphosphate.

Hemoglobin Casper (beta106Leu replaced by Pro) can be separated from hemoglobin (Hb) A by isoelectric focusing on polyacrylamide gel. This abnormal hemoglobin was estimated to be 30% of teh total by both isoelectric focusing and heat lability kinetics. Its oxygen equilibrium curves indicate a high oxygen affinity, low degree of subunit interaction, and a decreased Bohr effect. Mixtures of Hb Casper and Hb A appear to bind oxygen as if no hybrid molecules exist.

We have studied synthesis of specific proteins in two permanent lines of Friend virus-induced erythroleukemia cells (Friend line 745 and Ostertag line FSD-1, both derived from DBA/2 mice). By 96 hr following treatment with 1-2% dimethyl sulfoxide (Me2SO), up to 25% of the protein being synthesized by both these cultures is hemoglobin. At that time, hemoglobin constitutes up to 10% of the cellular soluble protein. Both lines synthesize heme and globin coordinately, and alpha and beta globin chains in a nearly balanced 1:1 ratio. However, the ratio of betaMajor:betaMinor chains synthesized by these induced Friend leukemia (FL) cells is approximately 9 in the FSD-1 line and 1.3 in the Friend Clone 745 line, whereas it is 4 in normal adult DBA/2 mouse erythrocytes. Evidence for the latter conclusion was obtained by electrophoresis of FL hemoglobins on cellulose acetate membranes, and also by chromatographic separation of alpha, betaMajor, and betaMinor globins on carboxymethylcellulose in 8 M urea at 20 degrees C. Carbonic anhydrase activity per mg protein is 3 times higher in induced than in control cultures. 2,3-diphosphoglyceric acid is not found in induced FL cells. Induced and control FL cells agglutinate strongly and equally with Phaseolus vulgaris phytohemagglutinin. The developmental process in these cultured leukemia cells appears to be an aberrant erythropoiesis.

Five patients with juvenile chronic granulocytic leukemia are described. Two patients had multiple cafe-au-lait spots compatible with von Recklinghausen's neurofibromatosis, and three had cutaneous xanthomata. Recurrent cutaneous leukemic infiltrates were noted in two patients. The clinical course of all five patients was characterized by recurrent respiratory symptoms and pulmonary infiltrates that responded to antileukemic therapy in three. Chemotherapy controlled the symptoms but did not influence the eventually fatal outcome.

Eleven members of a large Finnish family from Astoria, Oregon were studied because of an erythrocytosis. No abnormality was detected by the usual hemoglobin electrophoretic tests, but an abnormal variant was separated by reverse phase HPLC. All of the affected individuals have an increased oxygen affinity with a P50 for whole blood at 37 degrees C averaging 18 torr. Fifty percent of their hemoglobin was found to have a threonyl residue in place of the normal prolyl residue at position 36 (C2) of the beta globin chain. This abnormality is identical to Hb Linkoping which was recently reported in a Finnish man living in Sweden.

Erythrocyte 2,3-diphosphoglycerate levels are determined principally by a single molecule which has both diphosphoglycerate mutase (DPGM) and diphosphoglycerate phosphatase activities. An antiserum containing precipitating antibodies against this molecule was used to measure its amount in red cells from normal controls, from subjects with hypothyroidism or hyperthyroidism, and from a Japanese proband who has hemolytic anemia attributed to heterozygosity for a genetic variant. The amount of DPGM in normal adult erythrocytes is 0.98 +/- 0.014 mg/gm Hb. Slightly increased amounts are found in normal cord blood and in subjects with hyperthyroidism. Hypothyroid subjects have a significantly decreased concentration of DPGM, 0.82 +/- 0.06 mg/gm Hb. Restoration of euthyroid status during treatment results in a return of DPGM amounts to normal. The red cells of the Japanese subject contained 0.67 mg/gm Hb of DPGM as measured by immunoprecipitation.

Hb Burke [beta 107 (G9) Gly replaced by Arg] was discovered in a young woman with hemolytic anemia. A substitution in this position has not been previously reported either in the human beta-chain or in any of the animal beta-chains so far sequenced. The abnormal hemoglobin shows heat instability and a lowered oxygen affinity. The substitution of a large charged arginine residue for the small glycine residue in the G helix next to a heme contact (Leu-106) may be responsible for these effects. Hb Burke is compared with five other hemoglobins having Gly-Arg substitutions in other parts of the molecule.

The minor hemoglobin components, hemoglobin AIa+b and hemoglobin AIc, were measured in the 10% youngest and 10% oldest erythrocytes of 15 normal and 14 diabetic subjects. Erythrocyte fractions were obtained by centrifugation in isopyknic concentrations of dextran: 28.5% of 40,000-mol wt dextran yeilded the 10% lightest of young cells, and 30.5% dextran provided the 10% heaviest or old erythrocytes. Both normal and diabetic erythrocytes contain increased amounts of Hb AIa+b and Hb AIc in old as compared to young cells. In normal subjects, young cells contained 1.2+/-0.2%, and old cells contained 1.8+/-0.4% Hb AIa+b. Corresponding values for diabetic cells were 1.7+/-0.6 and 2.6+/-0.9%. Hb AIc increased from 3.1+/-0.8 to 6.0+/-1.1% in normals and from 5.1+/-2.1 to 10.1+/-3.7% in diabetics. The results indicate that both cell age and diabetes are significant determinants of the amounts of Hb AIa+b and Hb AIc.

A rightward shift in the blood oxygen dissociation curve occurs during the 1st mo of canine life. A detailed peptide analysis indicated that dogs do not have a separate fetal hemoglobin. Other erythrocyte components such as ATP, K+, Na+, and H+ were excluded as significant mediators of the postnatal oxygen affinity change. Erythrocyte 2,3-DPG levels essentially zero in fetal dogs, increased rapidly during the 1st mo of canine life. There was a significant correlation between this postnatal 2,3-DPG increase and the postnatal decrease in blood oxygen affinity. Dialyzed hemolysates of fetal or adult canine blood have the same intrinsic oxygen affinity and the same response to normal adult levels of 2,3-DPG. Furthermore, the magnitude and direction of this 2,3-DPG-induced decrease in oxygen affinity in vitro are comparable to the in vivo postnatal change in oxygen affinity.

C-myc proto-oncogene transcripts from serially harvested, colony-stimulating activity (CSA)-stimulated, normal progenitor-enriched human bone marrow cells were compared to those of the promyelocytic leukemia cell line HL-60 and to those of freshly obtained human myeloid leukemic cells. During the early culture period both normal and leukemic cells expressed the c-myc oncogene. In normal cells maximal expression occurred after 24 h of culture and did not occur in the absence of CSA. At this time, progranulocytes predominated in the cultured cells. Although cellular proliferation occurred for 96 h in vitro, c-myc expression ceased after 24-36 h. Terminally differentiated cells predominated in these cultures by 120 h. In contrast, although leukemic cells also expressed c-myc in vitro, transcription persisted throughout the culture period and, in the case of HL-60 cells, occurred in the absence of exogenous CSA. We also noted that normal cells with only one diploid gene copy exhibited, after 24 h of culture, only twofold fewer transcripts than did HL-60 cells in which there were 16 myc copies. Furthermore, c-myc mRNA degradation rates were similar in normal cells and in HL-60 cells. We conclude that c-myc transcription is a normal event in granulopoiesis linked to proliferative activity as well as to primitive developmental stage. Furthermore, the most consistent abnormality in leukemic cells in vitro is their failure to suppress transcriptional activity of this gene. We suggest that c-myc transcription in HL-60 cells may be appropriate for cells arrested at that developmental stage and that the amplified genes in HL-60 cells are quiescent relative to c-myc in normal cells at the same differentiation stage. The techniques described herein may be of value in identifying mechanisms by which normal hematopoietic cells suppress c-myc expression and aberrancies of these mechanisms in leukemic cells.

Hematologic parameters influencing tissue oxygen delivery in dogs during the first 4 mo of life have been investigated. The rapid growth and increase in body temperature during this period imply an increased metabolic rate and increased tissue oxygen demand. Hemoglobin concentration and hematocrit decrease during the 1st mo following birth. The total red cell mass does not decrease during this period. The observed hemodilution can be attributed to an increasing plasma volume in the growing animal. The blood oxygen affinity decreases during this same period, resulting in a more effective tissue oxygen delivery. Erythropoiesis, as estimated from the percent circulating reticulocytes, decreases following birth and does not increase until 1 mo of postnatal life. The increase of erythropoietic activity during the 2nd mo of postnatal life coincides with an increase in red cell mass, hematocrit, and hemoglobin concentration.

The oxygen affinities of blood from eight Basenji dogs homozygous for classical erythrocyte pyruvate kinase deficiency and four dogs heterozygous for the defect were compared with blood from 14 Labrador retrievers and two normal Basenji dogs. The homozygous dogs showed significant anemia compared to heterozygous and normal dogs (P is less than 0.01). The average blood P50 value (at 38C and plasma pH of 7.40) for both homozygous and heterozygous dogs was significantly higher (P is less than 0.01) than for normal dogs (33.6+/-0.4 and 31.8+/-0.7 vs 30.8+/-0.6 mm Hg). The concentrations of 2,3-DPG in the blood of both heterozygous and homozygous dogs were significantly higher than normal values. Four months after splenectomy P50 values declined to normal in four homozygous Basenji dogs without any change in the degree of anemia or blood 2,3-DPG concentrations. Iron kinetic studies showed a shorter plasma clearance time in a homozygous than in a normal dog with the heterozygote falling midway between. The red cell life span in the normal and heterozygous dogs was approximately 120 days. The 59Fe studies on the homozygous dog indicate markedly different survival characteristics which can be attributed to the existence of three populations of red cells differing in their life spans.

Three members of a family who have erythrocytosis and a new hemoglobin, designated hemoglobin Yakima, are described. The abnormal hemoglobin is characterized by the substitution of histidine for aspartic acid at residue 99 in the beta-chain. Of three possible structure-function relations which would account for the increased oxygen affinity of hemoglobin Yakima, only two seem likely. These are: (a) an intrachain shift in the normal relations between the F and G helices and the heme group, or (b) an effect of the substituted side chain at a region of contact between nonpolar residues of the alpha- and beta-chains which favors the oxyhemoglobin quarternary structure.