Homozygous familial hypercholesterolemia in Japan

Fifty-one homozygous patients with familial hypercholesterolemia, including our six patients, are described in this paper. Twenty were men and 31 were women. Their ages ranged between two and 52 years, with a mean of 16.8 years. Six patients exceeded the third decade. The mean age at death in seven patients was 17 years. The serum cholesterol levels were between 508 and 1,108 mg/dl. The mean and standard deviation (SD) of serum cholesterol were 713 ± 142 mg/dl. The serum cholesterol levels in the 35 parents (obligate heterozygotes) were between 246 and 571 mg/dl, except in one patient in whom the serum cholesterol level (936 mg/dl) was suggestive of homozygous familial hypercholesterolemia, considering the serum cholesterol level (354 mg/dl) of her heterozygous husband. The mean and SD of serum cholesterol levels of the 34 heterozygote parents were 342 ± 79 mg/dl. The mean and SD of serum cholesterol in 119 normal subjects were 187 ± 30 mg/dl. Thus, trimodai distribution was evident in the serum cholesterol levels of normal subjects, the heterozygotes and the homozygotes in Japan. The frequency of parental consanguinity was at least 33 per cent. The frequencies of ischemic heart disease in the age groups 0–9, 10–19 and above 20 years were 25 per cent (four of 16 patients), 33 per cent (six of 18 patients) and 53 per cent (nine of 17 patients), respectively. The frequency of homozygotes in Japan was in close accordance with those of Western countries. Therefore, the treatment of hypercholesterolemia and prevention of premature coronary heart disease in familial hypercholesterolemic patients are very important problems in Japan as well as in the West.     

An Alu-mediated 31.5-kb deletion as the cause of factor XI deficiency in 2 unrelated patients

Factor XI deficiency (MIM 264900) is an autosomal bleeding disorder of variable severity. Inheritance is not completely recessive as heterozygotes may display a distinct, if mild, bleeding tendency. Recent studies have shown the causative mutations of factor XI deficiency, outside the Ashkenazi Jewish population, to be highly heterogeneous. We studied 39 consecutively referred patients with factor XI deficiency to identify the molecular defect. Conventional mutation screening failed to identify a causative mutation in 4 of the 39 patients. Epstein-Barr virus (EBV)-transformed cells from these 4 patients were converted from a diploid to haploid chromosome complement. Subsequent analysis showed that 2 of the patients had a large deletion, which was masked in the heterozygous state by the presence of a normal allele. We report here the first confirmed whole gene deletion as the causative mutation of factor XI deficiency, the result of unequal homologous recombination between flanking Alu repeat sequences.     

Assessment of Actin FS and Actin FSL sensitivity to specific clotting factor deficiencies

We present a two centre study designed to assess the sensitivity of Actin FS and Actin FSL to deficiencies of factor VIII, IX, XI or XII. The study was undertaken at two centres to avoid bias due to the investigations being undertaken on one analyser. Samples from patients with a factor VIII (n = 36, F VIII = < 1.0–50 iu/dl), factor IX (n = 22, F IX = 2–48 iu/dl), factor XI (n = 23, F XI = 5–50 u/dl) or a factor XII (n = 18, F XII = 1–50 u/dl) deficient state were studied. Activated partial thromboplastin times (APTT) were determined using two batches of Actin FS and of Actin FSL; comparison of APTT results between centres was facilitated by the conversion of clotting times to ratios (test ÷geometric mean normal clotting time). APTT ratios were considered to be elevated if greater than two standard deviations above the mean normal. The factor deficient status of each sample was verified by assaying all samples for factors VIII, IX, XI and XII. Clotting factor assays were performed on a Sysmex CA-1000^? fitted with research software, which permitted the auto-dilution and testing of three serial dilution of both a reference preparation and each patient's sample. Assay results were calculated using parallel-line Bioassay principles. This procedure allowed for variation in clotting times due to the effect of temporal drift of any of the reagents within the assay system. Actin FS and Actin FSL demonstrate acceptable sensitivity to factor VIII deficiency, however, both reagents failed to detect a large proportion of factor XI (17.4% and 30.4% of samples, respectively) and factor XII (66.7% and 72.2%, respectively) deficiencies. The detection rate with Actin FSL for factor IX deficiency was also poor (36.4% not detected). As factor IX and XI deficiencies are both associated with haemorrhagic disorders, the inability of these reagents to detect such abnormalities gave cause for concern.     

Detection of glucose-6-phosphate dehydrogenase deficient heterozygotes

Heterozygotes for the Mediterranean type of severe G-6-PD deficiencywere investigated by a variety of tests.

The methemoglobin reduction test was most successful in detecting heterozygotes (about 80 per cent). Enumeration of methemoglobin containingcells on blood films (Kleihauer-Betke technic) did not improve these results.Quantitation of enzyme level was less successful (65 per cent), and determination of decolorization time by the BCB technic was least sensitivein heterozygote detection.

Methemoglobin reduction technics reflect a more indirect effect of themutant gene than enzyme assay. The superiority of these technics in heterozygote detection is probably caused by the genetically determined presenceof both normal and enzyme-deficient cells in G-6-PD deficient heterozygotes.Since methemoglobin reduction is carried out by individual cells, the population of enzyme deficient cells does not reduce methemoglobin, and thereforeeven a minority of deficient cells leads to abnormal test results. In contrast,enzyme assay is less successful for heterozygote detection, since measurementof enzyme level is carried out on hemolyzed red cells, where cellular mosaicismno longer exists.

An additional source of variation of enzyme levels in heterozygotes iscaused by the existence of genetically determined control of normal enzymelevel. Possession of a high capacity allele for G-6-PD activity may placea heterozygote in the normal range of enzyme activity.

The various tests were also applied to subjects with the mild Greek typeof G-6-PD deficiency. Males with this mutation had enzyme levels varyingbetween 12-45 per cent of the mean of normal males. Methemoglobin reduction test results were considerably less abnormal in hemizygotes with themild type of Mediterranean deficiency than in heterozygotes with the severedeficiency. Fewer heterozygotes with the mild deficiency were detected.

Submitted on February 4, 1966 Accepted on May 31, 1966     

Factor XI deficiency in an Arab Moslem family in Israel

An arab moslem family with members affected by PTA deficiency is described. 3 children were found to have major deficiency, factor XI procoagulant activity being 3, 3 and 4 units/dl. 8 members, including parents, paternal grandparents and 4 siblings, were found to have minor deficiency of factor XI (40 to 68 units/dl). Assays of immunoreactive material in 4 members corresponded to the level of procoagulant activity. In this family, gene expression is autosomal recessive. The only bleeding episode reported was haematuria in the propositus. No other spontaneous, post-trauma or post-operative bleeding was noted. The PTA deficiency was reported until now, mainly in ashkenazi jews. This family is the first case of PTA deficiency ever reported in arab moslems.     

Bleeding manifestations in heterozygotes with congenital FVII deficiency: a comparison with unaffected family members during a long observation period*

Objectives: To determine whether heterozygotes with FVII deficiency have a bleeding tendency or not.

Patients and methods: Eighty-four patients (OK) heterozygous for FVII deficiency, at the onset of the study, were paired with unaffected family members and followed for a long period of time (mean 22.6 years) for the occurrence of bleeding. Diagnosis of heterozygosis had to be based on family studies, clotting, immunological assays and genetic analysis.

Results: The mean FVII activity level was 0.51?IU/dl (range 35–65) and 94?IU/dl (range 88–118) in the heterozygotes and in the normal counterparts, respectively. Documented bleeding manifestations occurred in eight heterozygotes and in seven normal subjects. Statistical analysis of the difference was not significant. Bleeding manifestations were easy bruising, bleeding after tooth extractions, menorrhagia, epistaxis with no difference among the two groups. There was no strict correlation between bleeding and FVII activity levels.

Conclusions: The study indicates that heterozygotes for FVII deficiency show rare bleeding manifestations which are also present in the unaffected family members with normal FVII levels. This indicates that Factor VII activity levels played no role in the occurrence of the bleeding symptoms. Furthermore, FVII levels of around 0.40?IU/dl are capable of assuring a normal hemostasis.     

Factor XI deficiency acquired by liver transplantation

Factor XI deficiency (the Rosenthal syndrome), an autosomal recessive genetic defect, was transmitted to a patient after orthotopic liver transplantation. The deficiency was manifested by an isolated prolonged activated partial thromboplastin time (aPTT) after surgery. Hematologic evaluation using specific factor analysis revealed an absolute deficiency of factor XI. Stored serum obtained from the organ recipient before transplantation showed normal factor XI levels. When the liver donor's family was questioned, it was discovered that he was of Ashkenazi Jewish descent and that he had a history of bleeding after dental procedures. Before his death from intracerebral bleeding, he was documented to have an isolated prolonged aPTT value. This case shows that potentially morbid genetic defects can be transmitted by organ transplantation. It also provides evidence confirming that the liver is the only site of factor XI production.     

A molecular genetic study of factor XI deficiency

Factor XI deficiency is a rare bleeding diathesis found predominantly in Ashkenazi Jewish kindreds. A recent study of six Jewish patients identified three distinct mutations (Types I, II, and III) in the factor XI gene that were sufficient to fully define the genotypes of the patients. We have investigated 63 patients with factor XI deficiency and find overall allele frequencies of 44% for the type II mutation, 31% for the type III mutation, and 0% for the type I mutation. Therefore, 25% of the mutant factor XI alleles in our sample remain undefined. However, the distribution of mutant alleles is significantly different between Jewish and non-Jewish populations with hitherto undefined mutations accounting for 84% of the disease alleles in non-Jewish patients. Plasma factor XI:C levels were found to differ significantly between different homozygous and compound heterozygous genotypes and the inheritance of the II/III genotype was found to carry an increased risk of the most severe bleeding tendency.     

A Japanese family with high density lipoprotein deficiency

Two siblings with marked reduction of plasma high density lipoprotein (HDL) were found in a Japanese family. Their plasma cholesterol levels were very low (30-60 mg/dl), especially in the HDL fraction (0-1 mg/dl). The concentration of apolipoprotein (Apo) A-I in their plasma was 2-3 mg/dl and that of Apo A-II was 1.5-2.0 mg/dl, determined by means of a single radial immunodiffusion technique. An ultracentrifugally separated HDL fraction contained two different populations of lipoprotein particles, as shown by electron microscopy; a small particle with a diameter of 50-70 A and a relatively large particle at 200 A. Plasma lecithin: cholesterol acyltransferase activity was substantially retained in both cases. Hepatosplenomegaly was present and liver biopsy revealed lipid deposition in reticuloendothelial cells, although the tonsils were apparently normal. No severe atherosclerotic lesions were noticed. The results from these two cases were consistent with the characteristic features of homozygotes of familial HDL deficiency (Tangier disease). HDL cholesterol levels were relatively low in the parents and two children from one patient, which is consistent with the heterozygote state. Two other cases in the kindred were also found to have relatively low HDL cholesterol levels, besides these 4 cases of obligate heterozygotes. Apo A-I and Apo A-II levels in the plasma of the obligate heterozygotes, however, were within the normal range. Plasma low density lipoprotein in the patients moved faster in polyacrylamide gel electrophoresis than those of normal subjects, as did those in the heterozygotes.     

Hereditary plasma thromboplastin antecedent (PTA,FXI)

A rare case of factor XI (PTA) deficiency was discovered in a Saudi family in the Riyadh area. Nine members of the family were studied. Two were found to have a severe PTA deficiency’ levels of factor XI clotting activity were 0.01 i.u./ml and 0.02 i.u./ml respectively. Both plasmas were markedly deficient in factor XI antigen and appeared to be negative for cross-reactive material (CRM-). The parents were first cousins and both were found to have a minor PTA deficiency. Factor XI levels were: mother 0.048 i.u./ml and father 0.33 i.u./ml. Another sibling was found to have a FXI level of 0.47 i.u./ml. Menorrhagia and bleeding for 1 day after tooth extraction were the main bleeding manifestations found in one member with severe PTA deficiency. Clinically this member presented with iron deficiency anaemia. Other family members had no significant history of bleeding tendency. This is the first report of a Saudi Arabian family with PTA deficiency.     

Dental surgery in patients with severe factor XI deficiency without plasma replacement.

Bleeding following dental extraction is frequently the first manifestation of severe factor XI deficiency. Safe oral surgery has previously been performed in such patients by using plasma replacement therapy with or without concomitant administration of antifibrinolytic agents. The aim of this study was to determine whether such patients can undergo safe dental extractions using only an antifibrinolytic agent. The study group consisted of 19 patients with severe factor XI deficiency (factor XI:C level less than 14 U/dl) who had previously bled following dental extractions (14 patients) or other trauma (five patients). Tranexamic acid, 1 g q.i.d., was given from 12 h before surgery, until 7 days afterwards. No excessive bleeding was observed following dental extractions. One patient had slight oozing after 3 days which ceased spontaneously. Thus, plasma replacement no longer appears necessary for patients with severe factor XI deficiency requiring dental extractions.     

Hereditary plasma thromboplastin antecedent (PTA, FXI) deficiency in a Saudi family

A rare case of factor XI (PTA) deficiency was discovered in a Saudi family in the Riyadh area. Nine members of the family were studied. Two were found to have a severe PTA deficiency; levels of factor XI clotting activity were 0.01 i.u./ml and 0.02 i.u./ml respectively. Both plasmas were markedly deficient in factor XI antigen and appeared to be negative for cross-reactive material (CRM-). The parents were first cousins and both were found to have a minor PTA deficiency. Factor XI levels were: mother 0.048 i.u./ml and father 0.33 i.u./ml. Another sibling was found to have a FXI level of 0.47 i.u./ml. Menorrhagia and bleeding for 1 day after tooth extraction were the main bleeding manifestations found in one member with severe PTA deficiency. Clinically this member presented with iron deficiency anaemia. Other family members had no significant history of bleeding tendency. This is the first report of a Saudi Arabian family with PTA deficiency.     

A family with hereditary factor X deficiency with a point mutation Gla32 to Gln in the Gla domain (factor X Tokyo)

We report a new family with hereditary factor X deficiency. The propositus had a markedly prolonged prothrombin time, a mild prolongation of activated partial thromboplastin time and a clotting time activated by Russell's viper venom. Factor X activity in plasma was 3 u/dl (normal range 56-138 u/dl). Factor X antigen level was 61 u/dl. Molecular analysis revealed a homozygous mutation, Glu (GAG) to Gln (CAG) at residue 32 which normally undergoes gamma-carboxylation within the gamma-carboxyglutamic acid rich domain. The genotypes of family members completely correlated with their factor X activities. It is suggested that the Glu32 to Gln mutation is the molecular basis for the abnormal factor X in this family.     

Factor XI activity and factor XI antigen in homozygous and heterozygous factor XI deficiency

A relatively potent antiserum against highly purified, unactivated human factor XI antigen was raised in a rabbit. This antiserum, after concentration, neutralized 50% of the factor XI clotting activity of a standard normal plasma at an antiserum dilution of 1/900. The antiserum was used in a neutralization-inhibition assay to study the relation between factor XI clotting activity and factor XI antigen in plasma from ten unrelated patients with homozygous factor XI deficiency and from 12 heterozygous family members of these patients. No evidence of factor XI antigen significantly in excess of factor XI activity was found in either group. All data to date have been consistent with the hypothesis that hereditary factor XI deficiency represents a genetic disorder resulting from the absence of factor XI molecule. Severity of bleeding in factor XI deficiency could not be correlated with the level of factor XI activity or factor XI antigen.     

Protein-bound plasma homocyst(e)ine and identification of heterozygotes for cystathionine-synthase deficiency

We measured protein-bound plasma homocyst(e)ine in 15 normal adult subjects and nine heterozygotes for homocystinuria due to cystathionine -synthase deficiency. The mean (±SD) concentrations obtained in the two groups of subjects were 4.35±1.50 and 9.16±3.40 µmoll^–1, respectively. The mean values were significantly different, although the levels of three heterozygotes overlapped those of the control range. This method allows preliminary screening of the heterozygotes for homocystinuria and can be carried out by laboratories that have only facilities for amino acid analysis.     

Erythrocyte Pyrimidine 5''-Nucleotidase

In this study 31 family members of a patient with erythrocyte pyrimidine 5'-nucleotidase deficiency were studied. The activity of this enzyme in their erythrocytes is compared with levels in normal subjects and the problems surrounding heterozygote detection are discussed. The mean erythrocyte pyrimidine 5'-nucleotidase activity in 158 normal fresh blood samples was 130 +/- SD 29.8 mU/gHb. There was no significant difference between males and females. The enzyme level in a patient with non-spherocytic haemolytic anaemia was 6 mU/gHb. Of 31 relatives of the enzyme deficient patient examined five were clearly heterozygous for the enzyme defect. Their enzyme levels were below 50 mU/gHb. The father, who is an obligatory heterozygote, had an enzyme level of 87 mU/gHb which falls within the low values of the normal range. The distribution of enzyme activity in 26 family members having enzyme activities greater than 50 mU/gHb suggests that six of these may be carriers of the defective gene. We were unable to identify carriers by enzyme kinetic studies, electrophoresis, chromatographic examination of acid extractable nucleotides or measurement of enzyme levels in young and old erythrocyte populations. The last-mentioned technique showed that erythrocyte 5'-nucleotidase activity in reticulocytes may be as high as 1785 mU/gHb and declines rapidly as the cell ages reaching about 50 mU/gHb in the oldest cells. Blood samples which had been stored frozen were examined to see whether such samples were satisfactory for population studies. The mean enzyme activity 151 blood samples stored frozen for more than 12 months was 153+/-SD 44.7 mU/gHb. The increase in enzyme levels in the frozen samples appears to be greatest in samples showing haemolysis. In spite of the increased enzyme level frozen samples could be used to detect subjects with enzyme deficiency and some heterozygotes.     

Plasma replacement therapy during labor is not mandatory for women with severe factor XI deficiency.

Severe factor XI deficiency is an injury-related bleeding disorder. The risk of excessive post-partum hemorrhage in affected women has so far been evaluated in a relatively small number of patients and it is uncertain whether prophylactic treatment with fresh frozen plasma or factor XI concentrate is needed during or after vaginal or cesarean delivery. We retrospectively analyzed bleeding manifestations related to vaginal and/or cesarean deliveries in a cohort of 62 women with factor XI activity < 17 U/dl and evaluated whether replacement therapy is essential. Fifty-one women had 139 vaginal deliveries, six women had 13 cesarean deliveries, and five women had seven vaginal as well as five cesarean deliveries. Forty-three of the 62 women (69.4%) never experienced post-partum hemorrhage during 93 deliveries (85 vaginal, eight cesarean). Hemorrhage occurred in 19 women, which in six women accompanied each one of their 17 vaginal deliveries. Post-partum hemorrhage had no relationship with the abnormal genotype that caused factor XI deficiency nor with factor XI level. These observations suggest that the use of fresh frozen plasma or factor XI concentrate during and/or after vaginal delivery is not mandatory in women with severe factor XI deficiency and can be reserved for patients who develop excessive hemorrhage. For women requiring cesarean section it appears that the same policy can be advocated but more observations are needed.     

Severe hypercholesterolemia in a double heterozygote for lipoprotein lipase deficiency (LPL(Arita)) and apolipoprotein epsilon4: a report of a family with LPL(Arita)

Although heterozygous lipoprotein lipase (LPL) deficiency is not rare, only part of the phenotypes may have been reported in Japan. Here we describe a Japanese family with LPL(Arita), the most common mutation linked to familial LPL deficiency in Japan, and show for the first time a heterozygote for the mutation who had marked hypercholesterolemia due to increased low-density lipoprotein (LDL) cholesterol. The proband's mother, one of the heterozygotes for LPL(Arita) in the family, had both severe hypercholesterolemia (total cholesterol 306 mg/dl) with an especially increase in LDL-cholesterol and mild hypertriglyceridemia (180 mg/dl). She had normal LDL receptor activity and did not show clear evidence of possible causes of secondary hyperlipidemia. In addition to being heterozygous for LPL deficiency, she was also heterozygous for apo epsilon4. Because the epsilon4 allele is known to be associated with higher LDL-cholesterol, heterozygous apo epsilon4 may be one of causes of her LDL-cholesterol elevation. The other three heterozygotes for LPL(Arita) were moderate drinkers, and all of them had both remarkable hypertriglyceridemia and mild hypercholesterolemia due to increased very-low-density lipoproteins (VLDL). The results suggest that heterozygotes for LPL(Arita) can exhibit various phenotypes of hyperlipidemia, that is, hypertrigliceridemia and/or hypercholesterolemia due to not only increased VLDL but also increased LDL. The phenotypes appear to depend on some other genetic and environmental factors.     

Response to desmopressin of factors XI, X and V in patients with factor VIII deficiency and von Willebrand disease

Desmopressin [1-deamino-8-d-arginine vasopressin (DDAVP)] has been successfully used in the treatment of type 1 von Willebrand disease (VWD) and mild haemophilia A (MHA). Data suggest that DDAVP can increase factor XI (FXI) plasma levels and may represent an effective treatment for mild FXI deficiency. We assessed the DDAVP response of FXI coagulant activity (FXI:C), FXI antigen (FXI:Ag), factor V coagulant activity (FV:C), and factor X coagulant activity (FX:C) in 33 individuals with VWD or MHA. DDAVP did not produce a clinically significant increase in FXI:C, FXI:Ag, FX:C or FV:C in any patient. The mean +/- SD FXI:C pre-DDAVP (time 0) and at 1 h post-DDAVP was 90.7 (+/-22.9) U/dl and 92.1 (+/-20.9) U/dl, respectively. The mean (+/-SD) FXI:Ag at time 0 and 1 h was 92.2 (+/-20.1) U/dl and 89.9 (+/-21.3) U/dl, respectively. There was a small reduction at 1 h post-DDAVP in both FV:C, from 101.8 (+/-20.9) U/dl to 97.2 (+/-21.4) U/dl (P < 0.001), and FX:C from 103 (+/-19.5) U/dl to 98.8 (+/-18.7) U/dl (P < 0.001). No significant increase in FXI:C, FXI:Ag, FV:C or FX:C levels was seen at 4 h post-DDAVP. This study failed to demonstrate a clinically significant increase in the levels of FXI, FX or FV following administration of DDAVP.     

Hereditary factor VII deficiency: report of a case of intracranial hemorrhage

A case of factor VII deficiency in a 52-year-old woman who developed central nervous system hemorrhage is here reported. Screening coagulation tests were all normal except for prothrombin time, normotest and thrombotest. Specific assays of vitamin K-dependent factors revealed that factor VII activity was reduced (11 U/dl). The studies of the family demonstrated that 2 sisters out of 4 were heterozygous for the defect. The activity of factor VII in the offspring, classified as obligatory carriers, ranged between 62 and 78 U/dl, the antigen between 55 and 75 U/dl. The wide variability of factor VII in normal people and the possible compensative effect of normal alleles in carriers do not allow to define the variant, namely if the patient is a CRMR homozygote or a CRMR/CRM-double heterozygote.