Background The aim of this study was to identify characteristics of non-alcoholic fatty liver dis... more Background The aim of this study was to identify characteristics of non-alcoholic fatty liver disease (NAFLD) in adults with HFE p.C282Y/p.C282Y. Methods We retrospectively studied non-Hispanic white hemochromatosis probands with iron overload (serum ferritin (SF) > 300 µg/L (M), > 200 µg/L (F)) and p.C282Y/p.C282Y at non-screening diagnosis who did not report alcohol consumption > 14 g/d, have cirrhosis or other non-NAFLD liver disorders, use steatogenic medication, or have diagnoses of heritable disorders that increase NAFLD risk. We identified NAFLD-associated characteristics using univariate and multivariable analyses. Results There were 66 probands (31 men, 35 women), mean age 49 ± 14 (SD) y, of whom 16 (24.2%) had NAFLD. The following characteristics were higher in probands with NAFLD: median SF (1118 µg/L (range 259, 2663) vs. 567 µg/L (247, 2385); p = 0.0192); prevalence of elevated ALT/AST (alanine/aspartate aminotransferase) (43.8% vs. 10.0%; p = 0.0056); and prevalence of type 2 diabetes (T2DM) (31.3% vs. 10.0%; p = 0.0427). Mean age, sex, and prevalences of human leukocyte antigen-A*03 positivity, body mass index ≥ 30.0 kg/m 2 , hyperlipidemia, hypertension, and metabolic syndrome in probands with/without NAFLD did not differ significantly. Logistic regression on NAFLD using variables SF, elevated ALT/AST, and T2DM revealed: SF (p = 0.0318; odds ratio 1.0-1.0) and T2DM (p = 0.0342; 1.1-22.3). Median iron removed to achieve iron depletion (QFe) in probands with/without NAFLD did not differ significantly (3.6 g (1.4-7.2 g) vs. 2.8 g (0.7-11.0 g), respectively; p = 0.6862). Conclusions NAFLD in hemochromatosis probands with p.C282Y/p.C282Y is associated with higher median SF and greater T2DM prevalence, after adjustment for other factors. NAFLD does not influence QFe significantly.
Brackish water forms might be transported in bilge water or ballast tanks of ships. Thc American ... more Brackish water forms might be transported in bilge water or ballast tanks of ships. Thc American blue crab Oallinectes sapidus was probably transported to I srael in ballast tanks•. Because ships passing through the Panama Canal must, maintain a minimum draft, empty 8hips required to take on additional salt water ballast probably transport microscopic marine organisms from ocean to ocean'O. Several ports in the Chesapeake Bay-• for example, Baltimore, Norfolk and Newport Newsare either large commercial ports, sites of naval bases 0 1' both. West Point, Virginia, adjacent to station P-30, is visited by ocean.going freighters loading paper products. The controlling depth of t h e river channel to this point is less than 6•5 m, so bilges and ballast tanks are pumped to enable more cargo to be loaded. This may be the way in w hich brackish water organisms from similar regions elsewhere are being introduced. Three specimens of M. inexpectata from the Pamunkey River, Virginia, have been deposited in the Nationa l Museum of Natural Seicnces, Ottawa, Canada. One of us (D. R. C.) thanks the Nationa l Research Council of Canada for sllpport through a postdoctorate fellowship.
The immune macroglobulin from the paddlefish, Polyodon spathula, was purified and characterized p... more The immune macroglobulin from the paddlefish, Polyodon spathula, was purified and characterized physicochemically. The macroglobulin had a sedimentation coefficient of 14.2 S and a molecular weight of 660,000. Total reduction and alkylation in 7 M guanidine hydrochloride resulted in dissociation into heavy (H) and light (L) chains. The H and L chains were separated by gel filtration and found to have molecular weights of 58,100 and 21,000, respectively, employing sedimentation equilibrium in 5.0 M guanidine hydrochloride. Based on absorbance, equimolar quantities of the chains were recovered from the macroglobulin. These data are compatible with previously reported electron microscopic studies
To investigate the specificity of an inducible hemolymph bactericidin, West Indian spiny lobsters... more To investigate the specificity of an inducible hemolymph bactericidin, West Indian spiny lobsters were immunized with a variety of antigens. Bactericidal assays were conducted with strain EMB-1, a gram-negative bacillus isolated from normal lobster intestine. Animals given saline solution with or without 0.3% Formalin exhibited only a slight response similar to sham controls. Injections of type 2 pneumococci or bovine albumin produced slightly higher responses. These were much less pronounced than the homologous response to EMB-1. Formalin exhibited an adjuvant-like effect with EMB-1.
Hagfish were shown to be capable of synthesizing bactericidins after injection of gram-negative b... more Hagfish were shown to be capable of synthesizing bactericidins after injection of gram-negative bacteria. The bactericidins could be detected as early as 2 days after injection. The degree of specificity was not as impressive as in mammalian systems.
Since relatively few quantitative immunological studies had been conducted with invertebrates, th... more Since relatively few quantitative immunological studies had been conducted with invertebrates, this study was designed to learn more about the immune potential of the oyster, a molluscan representative. Experiments measured the primary and secondary clearance rates of T2 coliphage in the oyster. The fate of phage injected intracardially or intramuscularly was traced by measuring serum and tissue fluid plaque-forming unit levels at various times postinjection. Phage-neutralizing activity was also measured. Although circulating T2-neutralizing antibody was not induced in the oyster, secondary injections of T2 were cleared more rapidly than primary injections. The difference in clearance rates between primary and secondary groups was not as pronounced in the oyster as in the lemon shark, an animal capable of producing high levels of serum antibody to T2.
West Indian spiny lobsters, Panulirus argus , synthesized a hemolymph bactericidin after being in... more West Indian spiny lobsters, Panulirus argus , synthesized a hemolymph bactericidin after being injected with killed suspensions of gram-negative bacillus EMB-1 isolated from the normal gut of this lobster. To study differences between the primary response and secondary response, animals were given a primary antigen injection of EMB-1 followed by a second injection of the same antigen 22 to 51 days later. As a rule, secondary bactericidal responses were enhanced over the primary in a manner reminiscent of specific anamnesis in mammalian immunoglobulin synthesis. Immunological memory was also suggested when tertiary responses were compared to secondary and by the persistence of residual titers for many days or weeks without additional antigenic stimulation.
V\ 7e read with interest the article by Wilke et al 1 and VV would like to comment on Table I, wh... more V\ 7e read with interest the article by Wilke et al 1 and VV would like to comment on Table I, which summarizes the use of low-dose methotrexate for the treatment of rheumatoid arthritis. Under "Unique findings," the authors, referring to the article by Weinblatt et al, 2 state that HLA-DR2 was found with increased frequency in patients who had a marked response to methotrexate. We have conducted and published two studies demonstrating that HLA-DR2 is not a marker for disease response in patients with rheumatoid arthritis receiving methotrexate. The first study involved a group of 32 patients treated at our institution. Since they were not part of a controlled study, response was evaluated based on available data from the different treating rheumatologists. We found no association between response and DR2 or any other HLA specificity. 3 We then conducted a second and more rigorous study that involved 40 of 49 patients from the Cooperative Systematic Studies on the Rheumatic Diseases (Methotrexate-clinical trial). 4 No association was apparent. Moreover, no significant difference in the frequency distribution of any HLA-DR specificity was found between the eight responders and the 32 non-responders, and no responder was HLA-DR2 positive. 4 Thus we believe that neither HLA-DR2, nor any other HLA-DR specificity, can be considered a genetic marker for response to methotrexate.
Proceedings of the National Academy of Sciences, 1971
The immune IgM-like macroglobulins were isolated from three species of bony fishes ( Ictalurus pu... more The immune IgM-like macroglobulins were isolated from three species of bony fishes ( Ictalurus punctatus, Lepisosteus osseus , and Polyodon spathula ) representing the three orders of the subclass Actinopterygii. These macroglobulins were found to have sedimentation coefficients of 14 S and molecular weights of 600,000-630,000. The carbohydrate compositions were determined and found to be different from that of human macroglobulins. After reduction and alkylation, the heavy and light chains could be separated by gel filtration and were found to have molecular weights of approximately 70,000 and 23,000 respectively. The amino acid compositions of these chains were similar to those of mammalian IgM heavy and light chains. The macroglobulins were found by electron microscopy to have a tetrameric structure, in contrast to the pentameric structure found in mammalian, chicken, and shark IgM, i.e. in species later on the evolutionary scale.
Brackish water forms might be transported in bilge water or ballast tanks of ships. Thc American ... more Brackish water forms might be transported in bilge water or ballast tanks of ships. Thc American blue crab Oallinectes sapidus was probably transported to I srael in ballast tanks•. Because ships passing through the Panama Canal must, maintain a minimum draft, empty 8hips required to take on additional salt water ballast probably transport microscopic marine organisms from ocean to ocean'O. Several ports in the Chesapeake Bay-• for example, Baltimore, Norfolk and Newport Newsare either large commercial ports, sites of naval bases 0 1' both. West Point, Virginia, adjacent to station P-30, is visited by ocean.going freighters loading paper products. The controlling depth of t h e river channel to this point is less than 6•5 m, so bilges and ballast tanks are pumped to enable more cargo to be loaded. This may be the way in w hich brackish water organisms from similar regions elsewhere are being introduced. Three specimens of M. inexpectata from the Pamunkey River, Virginia, have been deposited in the Nationa l Museum of Natural Seicnces, Ottawa, Canada. One of us (D. R. C.) thanks the Nationa l Research Council of Canada for sllpport through a postdoctorate fellowship.
The purpose of these experiments was to test the hypothesis that impaired glucose-stimulated insu... more The purpose of these experiments was to test the hypothesis that impaired glucose-stimulated insulin secretion in NIDDM is due to mutations in the islet beta cell/liver glucose transporter (GLUT 2) gene. Using oligonucleotide primers flanking each of the 11 exons, the structural portion of the gene was studied by PCR-SSCP analysis. DNA from African-American females (n = 48), who had gestational diabetes but developed overt NIDDM after delivery, was studied. Each SSCP variant was sequenced directly from genomic DNA. Two amino acid substitutions from the previously reported sequence were found, one in exon 3 and the other in exon 4B. Four additional silent mutations in the coding region, and six intron mutations outside the splice junction consensus sequences, were also identified. The mutation GTC x ATC in exon 4B substituted Va1197 to Ile ~97. This amino acid substitution was found in only one NIDDM patient in a single allele, and was not found in 52 control subjects. This residue exists in the fifth membrane spanning domain, and Val at this position is conserved in mouse and rat GLUT 2, and human GLUT 1 to GLUT 4. The other codon change in exon3, ACT xATT, substituted Thr H~ to Ile la~ in the second membrane spanning domain. To determine the frequency of this non-conservative amino acid substitution, a PCR-LCR assay was developed. This assay was simple and highly specific for detection of this single nucleotide substitution. The allelic frequency of the ATT (Ile 1~~ in NIDDM patients (39.6%, n = 48) and that in controls (47.1%, n = 52) did not differ (p-0.32, Fisher's exact test). In conclusion, we identified two variant GLUT 2 glucose transporters in a subset of NIDDM patients. The rare variant in exon 4 B may contribute to the diabetic susceptibility and awaits further investigation. However, structural abnormalities of the GLUT 2 transporter associated with NIDDM appeared to be rare and were not likely to be a major determinant of genetic susceptibility to this type of diabetes in the population studied. [Diabetologia (1994) 37: 420-427]
We postulated that the severity of iron overload in homozygous hemochromatosis probands is relate... more We postulated that the severity of iron overload in homozygous hemochromatosis probands is related to the expression of HLA-A3 or D6S105 allele 8. Therefore, we used these markers to characterize Alabama hemochromatosis probands and normal control subjects. We then quantified the blood removed by phlebotomy to exhaust body iron stores and maintain normal serum ferritin concentrations in our hemochromatosis probands. Induction and maintenance phlebotomy requirements were significantly greater in presumed HLA-A3 homozygotes or in D6S105 allele 8 homozygotes than in homozygous probands lacking these markers. Intermediate values were observed in probands who were HLA-A3 or allele 8 heterozygotes, respectively. We also analyzed data from males and females separately. Among subjects of the same sex, the induction and maintenance phlebotomy requirements in subjects presumed to be HLA-A3 homozygotes or in allele 8 homozygotes were greater than those of other groups. Our results support the hypothesis that the severity of iron overload in hemochromatosis is determined predominantly by genetic factors, and provide evidence that two or more mutations for hemochromatosis exist. However, the design of our study does not permit a distinction to be made between allelic and locus heterogeneity for the hemochromatosis gene(s).
We evaluated Alabama hemochromatosis probands (n = 74) and normal control subjects (n = 142) for ... more We evaluated Alabama hemochromatosis probands (n = 74) and normal control subjects (n = 142) for expression of the hemochromatosis-associated mutations nt 845G-->A (845A; Cys282Tyr) and nt 187C-->G (His63Asp) in a gene linked to the major histocompatibility complex (MHC). We also tabulated parameters of iron metabolism and iron overload in probands and in obligate heterozygote family members of homozygous Cys282Tyr probands. Among probands, 59.4% were Cys282Tyr homozygotes and 20.3% were heterozygotes; 20.3% did not express this mutation. In normal control subjects, 14.7% were heterozygous for the Cys282Tyr mutation; one normal control subject was homozygous for the Cys282Tyr mutation. None (0 of 44) of our Cys282Tyr-homozygous hemochromatosis probands had the His63Asp mutation. Of the Cys282Tyr-heterozygous and -negative probands, the His63Asp mutation occurred in 26.7% (4/15) and 53.3% (8/15), respectively. In normal control subjects, 23.2% were heterozygous for the His63Asp mutation; 2.8% were homozygous. Induction phlebotomy requirements and other manifestations of iron overload were significantly greater in Cys282Tyr homozygotes than among other probands. Cys282Tyr-heterozygous probands had significantly higher values of serum iron parameters than did obligate Cys282Tyr heterozygotes whose values were, on the average, normal. Co-expression of HLA-A3, HLA-B7, and D6S105(8) was significantly more frequent in all subgroups of probands stratified by Cys282Tyr expression than in normal control subjects. These results demonstrate that the severity of iron overload in hemochromatosis is affected significantly by genetic factors. Further, our findings support the hypothesis that one or more MHC-linked genes other than that corresponding to the Cys282Tyr and His63Asp mutations contributes to increased iron absorption and iron overload in hemochromatosis probands.
Primary iron overload may be relatively common in African Americans, but its cause is incompletel... more Primary iron overload may be relatively common in African Americans, but its cause is incompletely understood. Thus, we evaluated genotype and phenotype characteristics of unselected African American index patients with primary iron overload who reside in central Alabama. All had hepatic iron concentration > or =30 micromol/g dry wt or > or =2.0 g of iron mobilized by phlebotomy to achieve iron depletion. Genotype analyses were performed in African American control subjects from the same region. There were 23 patients (19 men, 4 women); mean age at diagnosis was 52 +/- 12 years (1 SD) (range 32-69 years). Nine (39.1%) reported that they consumed > or =45 g of ethanol daily; five had chronic hepatitis C. Eight had some form of hemoglobinopathy or thalassemia. Mean serum transferrin saturation was 56 +/- 28% (range 15-100%). The geometric mean serum ferritin at diagnosis was 1076 ng/mL [95% confidence interval 297-3473 ng/mL]. Increased stainable liver iron was observed in hepatocytes only in 4 patients, in macrophages only in 8 patients, and in hepatocytes and macrophages in 8 patients. The mean quantity of iron mobilized by phlebotomy (corrected for iron absorbed during treatment) was 5.3 +/- 2.0 g (range 4.0-8.4 g). Iron removed by phlebotomy was greater in patients with hemoglobinopathy or thalassemia than in those without these forms of anemia (6.6 +/- 1.3 g vs 3.9 +/- 1.6 g, respectively; P = 0.0144). Daily consumption of > or =45 g of ethanol or chronic hepatitis C was not associated with an increased or decreased amount of phlebotomy-mobilized iron, on the average. The percentage of index patients positive for HFE C282Y was greater than that of controls (P = 0.0058). The respective percentages of phenotype positivity for HFE H63D, D6S105(8), and HLA-A*03 were similar in patients and controls. HFE S65C, I105T, and G93R were not detected in index or control subjects. Two of 13 patients were heterozygous for the ferroportin allele nt 744 G-->T (Q248H), although the phenotype frequency of this allele was similar in patients and 39 controls. Synonymous ferroportin alleles were also detected in some patients. The ceruloplasmin mutation nt 1099C-->T (exon 6; Arg367Cys) was detected in 1 of 2 patients tested. Abnormal alleles of beta-2 microglobulin, Nramp2, TFR2, hepcidin, or IRP2 alleles were not detected in either of the 2 patients so tested. We conclude that primary iron overload in African Americans is not the result of the mutation of a single gene. HFE C282Y, ferroportin 744 G-->T, and common forms of heritable anemia appear to account for increased iron absorption or retention in some patients.
Most cases of hemochromatosis are associated with mutations of the HFE gene on Ch6p. In southern ... more Most cases of hemochromatosis are associated with mutations of the HFE gene on Ch6p. In southern Italy and central Alabama, the percentages of patients with hemochromatosis who have "atypical" HFE genotypes (defined as lack of C282Y homozygosity, C282Y/H63D compound heterozygosity, or H63D homozygosity) are relatively great. A mutation of the transferrin receptor-2 gene (TFR2; exon 6, nt 750 C --> G, replaces TAC with stop signal TAG; Y250X) on Ch7q22 was recently identified in two Sicilian families with HFE mutation-negative hemochromatosis. We wanted to estimate the frequency of this mutation in persons from central Alabama. We evaluated Caucasian hemochromatosis probands with atypical HFE genotypes and African Americans with primary iron overload. We also studied control Caucasians, including persons of southern Italian/Sicilian heritage, and control African Americans. Analysis of genomic DNA was performed using a PCR-sequence-specific priming assay and positive control specimens from Sicilian hemochromatosis subjects heterozygous and homozygous for Y250X. Among Alabama subjects, this allele was not detected in 113 Caucasians, including 21 hemochromatosis probands with atypical HFE genotypes and 92 normal control subjects (including 27 of southern Italian/Sicilian descent). In African Americans, Y250X was not detected in 20 index cases with primary iron overload or in 274 unrelated control subjects. We conclude that Y250X is uncommon in Caucasians with hemochromatosis associated with atypical HFE genotypes, in African Americans with primary iron overload, and in the general Caucasian and African American population subgroups in central Alabama.
We studied DNA polymorphisms of HLA-DR and DQ alleles in 63 American black patients with systemic... more We studied DNA polymorphisms of HLA-DR and DQ alleles in 63 American black patients with systemic lupus erythematosus (SLE). We found no HLA-DR beta, DQ alpha, or DQ beta restriction fragment length polymorphism (RFLP) or RFLP-determined DR or DQ specificity associated with SLE in either the patients or in 57 control subjects. DRw52b was positively associated with renal involvement and negatively associated with anti-nuclear RNP antibodies. Antibodies to Ro (SS-A) and La (SS-B) were associated with DR3(DRw17), DQw2.3. Early-onset SLE (less than or equal to 20 years of age) was associated with DRw8, and the frequency of neuropsychiatric involvement correlated negatively with a 3.7-kb Taq I DQ alpha RFLP. This suggests a role for DR and DQ genes in the clinical and serologic expression of SLE in American blacks.
Proceedings of the National Academy of Sciences, 1978
Conventional fractionation procedures were used in an effort to define the subcellular distributi... more Conventional fractionation procedures were used in an effort to define the subcellular distribution of the Thy-1 alloantigen in whole mouse brain. After discontinuous sucrose density gradient centrifugation of isotonic postnuclear particulate fractions, the bulk of Thy-1 was recovered in regions of the gradients containing synaptosomes. The synaptosome fraction that banded at 1.2 M sucrose yielded a specific activity for Thy-1 significantly greater than the synaptosomes separating at 1.4 M sucrose. Osmotic lysis of both synaptosome fractions resulted in further enrichment in Thy-1 activity, with no concomitant decrease in yields. The synaptosomal membranes obtained in this way were subsequently treated with Triton X-100 and subjected to further density gradient centrifugation. Although the detergent treatment resulted in some loss of antigenic activity, the gradient fractions that contained Thy-1 also were found by electron microscopy to be richest in synaptic junctional complexes. ...
Metabolic Syndrome and Related Disorders, Aug 1, 2018
Background: We sought to determine associations with insulin resistance (IR) and metabolic syndro... more Background: We sought to determine associations with insulin resistance (IR) and metabolic syndrome (MetS) in African Americans. Methods: We studied African American adults without diabetes in a postscreening examination. Participants included Cases: transferrin saturation (TS) >50% and serum ferritin (SF) >300 mg/L (M), and TS >45% and SF >200 mg/L (F), regardless of HFE genotype; and Controls: TS/SF 25th to 75th percentiles and HFE wt/wt (wild type). We excluded participants with fasting <8 h; fasting glucose >126 mg/dL; hepatitis B or C; cirrhosis; pregnancy; or incomplete datasets. We analyzed age; sex; Case/Control; body mass index (BMI); systolic and diastolic blood pressures; neutrophils; lymphocytes; alanine aminotransferase; aspartate aminotransferase; elevated C-reactive protein (CRP >0.5 mg/L); TS; and SF. We computed homeostasis model assessment of insulin resistance (HOMA-IR) using fasting serum glucose and insulin, and defined IR as HOMA-IR fourth quartile (‡2.42). Results: There were 312 Cases and 86 Controls (56.3% men). Ninety-one percent had HFE wt/wt. None had HFE p.C282Y. A significant increasing trend across HOMA-IR quartiles was observed for BMI only. Multivariable regression on HOMA-IR revealed significant positive associations: age; BMI; lymphocytes; SF; and CRP >0.5 mg/L; and significant negative associations: neutrophils and TS. Logistic regression on IR revealed BMI [odds ratio (OR) 1.3 (95% confidence interval 1.2-1.4)] and CRP >0.5 mg/L [OR 2.7 (1.2-6.3)]. Fourteen participants (3.5%) had MetS. Logistic regression on MetS revealed one association: IR [OR 7.4 (2.1-25.2)]. Conclusions: In African Americans without diabetes, IR was associated with BMI and CRP >0.5 mg/L, after adjustment for other variables. MetS was associated with IR alone.
Background The aim of this study was to identify characteristics of non-alcoholic fatty liver dis... more Background The aim of this study was to identify characteristics of non-alcoholic fatty liver disease (NAFLD) in adults with HFE p.C282Y/p.C282Y. Methods We retrospectively studied non-Hispanic white hemochromatosis probands with iron overload (serum ferritin (SF) > 300 µg/L (M), > 200 µg/L (F)) and p.C282Y/p.C282Y at non-screening diagnosis who did not report alcohol consumption > 14 g/d, have cirrhosis or other non-NAFLD liver disorders, use steatogenic medication, or have diagnoses of heritable disorders that increase NAFLD risk. We identified NAFLD-associated characteristics using univariate and multivariable analyses. Results There were 66 probands (31 men, 35 women), mean age 49 ± 14 (SD) y, of whom 16 (24.2%) had NAFLD. The following characteristics were higher in probands with NAFLD: median SF (1118 µg/L (range 259, 2663) vs. 567 µg/L (247, 2385); p = 0.0192); prevalence of elevated ALT/AST (alanine/aspartate aminotransferase) (43.8% vs. 10.0%; p = 0.0056); and prevalence of type 2 diabetes (T2DM) (31.3% vs. 10.0%; p = 0.0427). Mean age, sex, and prevalences of human leukocyte antigen-A*03 positivity, body mass index ≥ 30.0 kg/m 2 , hyperlipidemia, hypertension, and metabolic syndrome in probands with/without NAFLD did not differ significantly. Logistic regression on NAFLD using variables SF, elevated ALT/AST, and T2DM revealed: SF (p = 0.0318; odds ratio 1.0-1.0) and T2DM (p = 0.0342; 1.1-22.3). Median iron removed to achieve iron depletion (QFe) in probands with/without NAFLD did not differ significantly (3.6 g (1.4-7.2 g) vs. 2.8 g (0.7-11.0 g), respectively; p = 0.6862). Conclusions NAFLD in hemochromatosis probands with p.C282Y/p.C282Y is associated with higher median SF and greater T2DM prevalence, after adjustment for other factors. NAFLD does not influence QFe significantly.
Brackish water forms might be transported in bilge water or ballast tanks of ships. Thc American ... more Brackish water forms might be transported in bilge water or ballast tanks of ships. Thc American blue crab Oallinectes sapidus was probably transported to I srael in ballast tanks•. Because ships passing through the Panama Canal must, maintain a minimum draft, empty 8hips required to take on additional salt water ballast probably transport microscopic marine organisms from ocean to ocean'O. Several ports in the Chesapeake Bay-• for example, Baltimore, Norfolk and Newport Newsare either large commercial ports, sites of naval bases 0 1' both. West Point, Virginia, adjacent to station P-30, is visited by ocean.going freighters loading paper products. The controlling depth of t h e river channel to this point is less than 6•5 m, so bilges and ballast tanks are pumped to enable more cargo to be loaded. This may be the way in w hich brackish water organisms from similar regions elsewhere are being introduced. Three specimens of M. inexpectata from the Pamunkey River, Virginia, have been deposited in the Nationa l Museum of Natural Seicnces, Ottawa, Canada. One of us (D. R. C.) thanks the Nationa l Research Council of Canada for sllpport through a postdoctorate fellowship.
The immune macroglobulin from the paddlefish, Polyodon spathula, was purified and characterized p... more The immune macroglobulin from the paddlefish, Polyodon spathula, was purified and characterized physicochemically. The macroglobulin had a sedimentation coefficient of 14.2 S and a molecular weight of 660,000. Total reduction and alkylation in 7 M guanidine hydrochloride resulted in dissociation into heavy (H) and light (L) chains. The H and L chains were separated by gel filtration and found to have molecular weights of 58,100 and 21,000, respectively, employing sedimentation equilibrium in 5.0 M guanidine hydrochloride. Based on absorbance, equimolar quantities of the chains were recovered from the macroglobulin. These data are compatible with previously reported electron microscopic studies
To investigate the specificity of an inducible hemolymph bactericidin, West Indian spiny lobsters... more To investigate the specificity of an inducible hemolymph bactericidin, West Indian spiny lobsters were immunized with a variety of antigens. Bactericidal assays were conducted with strain EMB-1, a gram-negative bacillus isolated from normal lobster intestine. Animals given saline solution with or without 0.3% Formalin exhibited only a slight response similar to sham controls. Injections of type 2 pneumococci or bovine albumin produced slightly higher responses. These were much less pronounced than the homologous response to EMB-1. Formalin exhibited an adjuvant-like effect with EMB-1.
Hagfish were shown to be capable of synthesizing bactericidins after injection of gram-negative b... more Hagfish were shown to be capable of synthesizing bactericidins after injection of gram-negative bacteria. The bactericidins could be detected as early as 2 days after injection. The degree of specificity was not as impressive as in mammalian systems.
Since relatively few quantitative immunological studies had been conducted with invertebrates, th... more Since relatively few quantitative immunological studies had been conducted with invertebrates, this study was designed to learn more about the immune potential of the oyster, a molluscan representative. Experiments measured the primary and secondary clearance rates of T2 coliphage in the oyster. The fate of phage injected intracardially or intramuscularly was traced by measuring serum and tissue fluid plaque-forming unit levels at various times postinjection. Phage-neutralizing activity was also measured. Although circulating T2-neutralizing antibody was not induced in the oyster, secondary injections of T2 were cleared more rapidly than primary injections. The difference in clearance rates between primary and secondary groups was not as pronounced in the oyster as in the lemon shark, an animal capable of producing high levels of serum antibody to T2.
West Indian spiny lobsters, Panulirus argus , synthesized a hemolymph bactericidin after being in... more West Indian spiny lobsters, Panulirus argus , synthesized a hemolymph bactericidin after being injected with killed suspensions of gram-negative bacillus EMB-1 isolated from the normal gut of this lobster. To study differences between the primary response and secondary response, animals were given a primary antigen injection of EMB-1 followed by a second injection of the same antigen 22 to 51 days later. As a rule, secondary bactericidal responses were enhanced over the primary in a manner reminiscent of specific anamnesis in mammalian immunoglobulin synthesis. Immunological memory was also suggested when tertiary responses were compared to secondary and by the persistence of residual titers for many days or weeks without additional antigenic stimulation.
V\ 7e read with interest the article by Wilke et al 1 and VV would like to comment on Table I, wh... more V\ 7e read with interest the article by Wilke et al 1 and VV would like to comment on Table I, which summarizes the use of low-dose methotrexate for the treatment of rheumatoid arthritis. Under "Unique findings," the authors, referring to the article by Weinblatt et al, 2 state that HLA-DR2 was found with increased frequency in patients who had a marked response to methotrexate. We have conducted and published two studies demonstrating that HLA-DR2 is not a marker for disease response in patients with rheumatoid arthritis receiving methotrexate. The first study involved a group of 32 patients treated at our institution. Since they were not part of a controlled study, response was evaluated based on available data from the different treating rheumatologists. We found no association between response and DR2 or any other HLA specificity. 3 We then conducted a second and more rigorous study that involved 40 of 49 patients from the Cooperative Systematic Studies on the Rheumatic Diseases (Methotrexate-clinical trial). 4 No association was apparent. Moreover, no significant difference in the frequency distribution of any HLA-DR specificity was found between the eight responders and the 32 non-responders, and no responder was HLA-DR2 positive. 4 Thus we believe that neither HLA-DR2, nor any other HLA-DR specificity, can be considered a genetic marker for response to methotrexate.
Proceedings of the National Academy of Sciences, 1971
The immune IgM-like macroglobulins were isolated from three species of bony fishes ( Ictalurus pu... more The immune IgM-like macroglobulins were isolated from three species of bony fishes ( Ictalurus punctatus, Lepisosteus osseus , and Polyodon spathula ) representing the three orders of the subclass Actinopterygii. These macroglobulins were found to have sedimentation coefficients of 14 S and molecular weights of 600,000-630,000. The carbohydrate compositions were determined and found to be different from that of human macroglobulins. After reduction and alkylation, the heavy and light chains could be separated by gel filtration and were found to have molecular weights of approximately 70,000 and 23,000 respectively. The amino acid compositions of these chains were similar to those of mammalian IgM heavy and light chains. The macroglobulins were found by electron microscopy to have a tetrameric structure, in contrast to the pentameric structure found in mammalian, chicken, and shark IgM, i.e. in species later on the evolutionary scale.
Brackish water forms might be transported in bilge water or ballast tanks of ships. Thc American ... more Brackish water forms might be transported in bilge water or ballast tanks of ships. Thc American blue crab Oallinectes sapidus was probably transported to I srael in ballast tanks•. Because ships passing through the Panama Canal must, maintain a minimum draft, empty 8hips required to take on additional salt water ballast probably transport microscopic marine organisms from ocean to ocean'O. Several ports in the Chesapeake Bay-• for example, Baltimore, Norfolk and Newport Newsare either large commercial ports, sites of naval bases 0 1' both. West Point, Virginia, adjacent to station P-30, is visited by ocean.going freighters loading paper products. The controlling depth of t h e river channel to this point is less than 6•5 m, so bilges and ballast tanks are pumped to enable more cargo to be loaded. This may be the way in w hich brackish water organisms from similar regions elsewhere are being introduced. Three specimens of M. inexpectata from the Pamunkey River, Virginia, have been deposited in the Nationa l Museum of Natural Seicnces, Ottawa, Canada. One of us (D. R. C.) thanks the Nationa l Research Council of Canada for sllpport through a postdoctorate fellowship.
The purpose of these experiments was to test the hypothesis that impaired glucose-stimulated insu... more The purpose of these experiments was to test the hypothesis that impaired glucose-stimulated insulin secretion in NIDDM is due to mutations in the islet beta cell/liver glucose transporter (GLUT 2) gene. Using oligonucleotide primers flanking each of the 11 exons, the structural portion of the gene was studied by PCR-SSCP analysis. DNA from African-American females (n = 48), who had gestational diabetes but developed overt NIDDM after delivery, was studied. Each SSCP variant was sequenced directly from genomic DNA. Two amino acid substitutions from the previously reported sequence were found, one in exon 3 and the other in exon 4B. Four additional silent mutations in the coding region, and six intron mutations outside the splice junction consensus sequences, were also identified. The mutation GTC x ATC in exon 4B substituted Va1197 to Ile ~97. This amino acid substitution was found in only one NIDDM patient in a single allele, and was not found in 52 control subjects. This residue exists in the fifth membrane spanning domain, and Val at this position is conserved in mouse and rat GLUT 2, and human GLUT 1 to GLUT 4. The other codon change in exon3, ACT xATT, substituted Thr H~ to Ile la~ in the second membrane spanning domain. To determine the frequency of this non-conservative amino acid substitution, a PCR-LCR assay was developed. This assay was simple and highly specific for detection of this single nucleotide substitution. The allelic frequency of the ATT (Ile 1~~ in NIDDM patients (39.6%, n = 48) and that in controls (47.1%, n = 52) did not differ (p-0.32, Fisher's exact test). In conclusion, we identified two variant GLUT 2 glucose transporters in a subset of NIDDM patients. The rare variant in exon 4 B may contribute to the diabetic susceptibility and awaits further investigation. However, structural abnormalities of the GLUT 2 transporter associated with NIDDM appeared to be rare and were not likely to be a major determinant of genetic susceptibility to this type of diabetes in the population studied. [Diabetologia (1994) 37: 420-427]
We postulated that the severity of iron overload in homozygous hemochromatosis probands is relate... more We postulated that the severity of iron overload in homozygous hemochromatosis probands is related to the expression of HLA-A3 or D6S105 allele 8. Therefore, we used these markers to characterize Alabama hemochromatosis probands and normal control subjects. We then quantified the blood removed by phlebotomy to exhaust body iron stores and maintain normal serum ferritin concentrations in our hemochromatosis probands. Induction and maintenance phlebotomy requirements were significantly greater in presumed HLA-A3 homozygotes or in D6S105 allele 8 homozygotes than in homozygous probands lacking these markers. Intermediate values were observed in probands who were HLA-A3 or allele 8 heterozygotes, respectively. We also analyzed data from males and females separately. Among subjects of the same sex, the induction and maintenance phlebotomy requirements in subjects presumed to be HLA-A3 homozygotes or in allele 8 homozygotes were greater than those of other groups. Our results support the hypothesis that the severity of iron overload in hemochromatosis is determined predominantly by genetic factors, and provide evidence that two or more mutations for hemochromatosis exist. However, the design of our study does not permit a distinction to be made between allelic and locus heterogeneity for the hemochromatosis gene(s).
We evaluated Alabama hemochromatosis probands (n = 74) and normal control subjects (n = 142) for ... more We evaluated Alabama hemochromatosis probands (n = 74) and normal control subjects (n = 142) for expression of the hemochromatosis-associated mutations nt 845G--&gt;A (845A; Cys282Tyr) and nt 187C--&gt;G (His63Asp) in a gene linked to the major histocompatibility complex (MHC). We also tabulated parameters of iron metabolism and iron overload in probands and in obligate heterozygote family members of homozygous Cys282Tyr probands. Among probands, 59.4% were Cys282Tyr homozygotes and 20.3% were heterozygotes; 20.3% did not express this mutation. In normal control subjects, 14.7% were heterozygous for the Cys282Tyr mutation; one normal control subject was homozygous for the Cys282Tyr mutation. None (0 of 44) of our Cys282Tyr-homozygous hemochromatosis probands had the His63Asp mutation. Of the Cys282Tyr-heterozygous and -negative probands, the His63Asp mutation occurred in 26.7% (4/15) and 53.3% (8/15), respectively. In normal control subjects, 23.2% were heterozygous for the His63Asp mutation; 2.8% were homozygous. Induction phlebotomy requirements and other manifestations of iron overload were significantly greater in Cys282Tyr homozygotes than among other probands. Cys282Tyr-heterozygous probands had significantly higher values of serum iron parameters than did obligate Cys282Tyr heterozygotes whose values were, on the average, normal. Co-expression of HLA-A3, HLA-B7, and D6S105(8) was significantly more frequent in all subgroups of probands stratified by Cys282Tyr expression than in normal control subjects. These results demonstrate that the severity of iron overload in hemochromatosis is affected significantly by genetic factors. Further, our findings support the hypothesis that one or more MHC-linked genes other than that corresponding to the Cys282Tyr and His63Asp mutations contributes to increased iron absorption and iron overload in hemochromatosis probands.
Primary iron overload may be relatively common in African Americans, but its cause is incompletel... more Primary iron overload may be relatively common in African Americans, but its cause is incompletely understood. Thus, we evaluated genotype and phenotype characteristics of unselected African American index patients with primary iron overload who reside in central Alabama. All had hepatic iron concentration &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt; or =30 micromol/g dry wt or &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt; or =2.0 g of iron mobilized by phlebotomy to achieve iron depletion. Genotype analyses were performed in African American control subjects from the same region. There were 23 patients (19 men, 4 women); mean age at diagnosis was 52 +/- 12 years (1 SD) (range 32-69 years). Nine (39.1%) reported that they consumed &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt; or =45 g of ethanol daily; five had chronic hepatitis C. Eight had some form of hemoglobinopathy or thalassemia. Mean serum transferrin saturation was 56 +/- 28% (range 15-100%). The geometric mean serum ferritin at diagnosis was 1076 ng/mL [95% confidence interval 297-3473 ng/mL]. Increased stainable liver iron was observed in hepatocytes only in 4 patients, in macrophages only in 8 patients, and in hepatocytes and macrophages in 8 patients. The mean quantity of iron mobilized by phlebotomy (corrected for iron absorbed during treatment) was 5.3 +/- 2.0 g (range 4.0-8.4 g). Iron removed by phlebotomy was greater in patients with hemoglobinopathy or thalassemia than in those without these forms of anemia (6.6 +/- 1.3 g vs 3.9 +/- 1.6 g, respectively; P = 0.0144). Daily consumption of &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt; or =45 g of ethanol or chronic hepatitis C was not associated with an increased or decreased amount of phlebotomy-mobilized iron, on the average. The percentage of index patients positive for HFE C282Y was greater than that of controls (P = 0.0058). The respective percentages of phenotype positivity for HFE H63D, D6S105(8), and HLA-A*03 were similar in patients and controls. HFE S65C, I105T, and G93R were not detected in index or control subjects. Two of 13 patients were heterozygous for the ferroportin allele nt 744 G--&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt;T (Q248H), although the phenotype frequency of this allele was similar in patients and 39 controls. Synonymous ferroportin alleles were also detected in some patients. The ceruloplasmin mutation nt 1099C--&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt;T (exon 6; Arg367Cys) was detected in 1 of 2 patients tested. Abnormal alleles of beta-2 microglobulin, Nramp2, TFR2, hepcidin, or IRP2 alleles were not detected in either of the 2 patients so tested. We conclude that primary iron overload in African Americans is not the result of the mutation of a single gene. HFE C282Y, ferroportin 744 G--&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt;T, and common forms of heritable anemia appear to account for increased iron absorption or retention in some patients.
Most cases of hemochromatosis are associated with mutations of the HFE gene on Ch6p. In southern ... more Most cases of hemochromatosis are associated with mutations of the HFE gene on Ch6p. In southern Italy and central Alabama, the percentages of patients with hemochromatosis who have &amp;amp;quot;atypical&amp;amp;quot; HFE genotypes (defined as lack of C282Y homozygosity, C282Y/H63D compound heterozygosity, or H63D homozygosity) are relatively great. A mutation of the transferrin receptor-2 gene (TFR2; exon 6, nt 750 C --&amp;amp;gt; G, replaces TAC with stop signal TAG; Y250X) on Ch7q22 was recently identified in two Sicilian families with HFE mutation-negative hemochromatosis. We wanted to estimate the frequency of this mutation in persons from central Alabama. We evaluated Caucasian hemochromatosis probands with atypical HFE genotypes and African Americans with primary iron overload. We also studied control Caucasians, including persons of southern Italian/Sicilian heritage, and control African Americans. Analysis of genomic DNA was performed using a PCR-sequence-specific priming assay and positive control specimens from Sicilian hemochromatosis subjects heterozygous and homozygous for Y250X. Among Alabama subjects, this allele was not detected in 113 Caucasians, including 21 hemochromatosis probands with atypical HFE genotypes and 92 normal control subjects (including 27 of southern Italian/Sicilian descent). In African Americans, Y250X was not detected in 20 index cases with primary iron overload or in 274 unrelated control subjects. We conclude that Y250X is uncommon in Caucasians with hemochromatosis associated with atypical HFE genotypes, in African Americans with primary iron overload, and in the general Caucasian and African American population subgroups in central Alabama.
We studied DNA polymorphisms of HLA-DR and DQ alleles in 63 American black patients with systemic... more We studied DNA polymorphisms of HLA-DR and DQ alleles in 63 American black patients with systemic lupus erythematosus (SLE). We found no HLA-DR beta, DQ alpha, or DQ beta restriction fragment length polymorphism (RFLP) or RFLP-determined DR or DQ specificity associated with SLE in either the patients or in 57 control subjects. DRw52b was positively associated with renal involvement and negatively associated with anti-nuclear RNP antibodies. Antibodies to Ro (SS-A) and La (SS-B) were associated with DR3(DRw17), DQw2.3. Early-onset SLE (less than or equal to 20 years of age) was associated with DRw8, and the frequency of neuropsychiatric involvement correlated negatively with a 3.7-kb Taq I DQ alpha RFLP. This suggests a role for DR and DQ genes in the clinical and serologic expression of SLE in American blacks.
Proceedings of the National Academy of Sciences, 1978
Conventional fractionation procedures were used in an effort to define the subcellular distributi... more Conventional fractionation procedures were used in an effort to define the subcellular distribution of the Thy-1 alloantigen in whole mouse brain. After discontinuous sucrose density gradient centrifugation of isotonic postnuclear particulate fractions, the bulk of Thy-1 was recovered in regions of the gradients containing synaptosomes. The synaptosome fraction that banded at 1.2 M sucrose yielded a specific activity for Thy-1 significantly greater than the synaptosomes separating at 1.4 M sucrose. Osmotic lysis of both synaptosome fractions resulted in further enrichment in Thy-1 activity, with no concomitant decrease in yields. The synaptosomal membranes obtained in this way were subsequently treated with Triton X-100 and subjected to further density gradient centrifugation. Although the detergent treatment resulted in some loss of antigenic activity, the gradient fractions that contained Thy-1 also were found by electron microscopy to be richest in synaptic junctional complexes. ...
Metabolic Syndrome and Related Disorders, Aug 1, 2018
Background: We sought to determine associations with insulin resistance (IR) and metabolic syndro... more Background: We sought to determine associations with insulin resistance (IR) and metabolic syndrome (MetS) in African Americans. Methods: We studied African American adults without diabetes in a postscreening examination. Participants included Cases: transferrin saturation (TS) >50% and serum ferritin (SF) >300 mg/L (M), and TS >45% and SF >200 mg/L (F), regardless of HFE genotype; and Controls: TS/SF 25th to 75th percentiles and HFE wt/wt (wild type). We excluded participants with fasting <8 h; fasting glucose >126 mg/dL; hepatitis B or C; cirrhosis; pregnancy; or incomplete datasets. We analyzed age; sex; Case/Control; body mass index (BMI); systolic and diastolic blood pressures; neutrophils; lymphocytes; alanine aminotransferase; aspartate aminotransferase; elevated C-reactive protein (CRP >0.5 mg/L); TS; and SF. We computed homeostasis model assessment of insulin resistance (HOMA-IR) using fasting serum glucose and insulin, and defined IR as HOMA-IR fourth quartile (‡2.42). Results: There were 312 Cases and 86 Controls (56.3% men). Ninety-one percent had HFE wt/wt. None had HFE p.C282Y. A significant increasing trend across HOMA-IR quartiles was observed for BMI only. Multivariable regression on HOMA-IR revealed significant positive associations: age; BMI; lymphocytes; SF; and CRP >0.5 mg/L; and significant negative associations: neutrophils and TS. Logistic regression on IR revealed BMI [odds ratio (OR) 1.3 (95% confidence interval 1.2-1.4)] and CRP >0.5 mg/L [OR 2.7 (1.2-6.3)]. Fourteen participants (3.5%) had MetS. Logistic regression on MetS revealed one association: IR [OR 7.4 (2.1-25.2)]. Conclusions: In African Americans without diabetes, IR was associated with BMI and CRP >0.5 mg/L, after adjustment for other variables. MetS was associated with IR alone.
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Papers by Ronald Acton