Activated STING in a vascular and pulmonary syndrome.

BACKGROUND: The study of autoinflammatory diseases has uncovered mechanisms underlying cytokine dysregulation and inflammation. METHODS: We analyzed the DNA of an index patient with early-onset systemic inflammation, cutaneous vasculopathy, and pulmonary inflammation. We sequenced a candidate gene, TMEM173, encoding the stimulator of interferon genes (STING), in this patient and in five unrelated children with similar clinical phenotypes. Four children were evaluated clinically and immunologically. With the STING ligand cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), we stimulated peripheral-blood mononuclear cells and fibroblasts from patients and controls, as well as commercially obtained endothelial cells, and then assayed transcription of IFNB1, the gene encoding interferon-ß, in the stimulated cells. We analyzed IFNB1 reporter levels in HEK293T cells cotransfected with mutant or nonmutant STING constructs. Mutant STING leads to increased phosphorylation of signal transducer and activator of transcription 1 (STAT1), so we tested the effect of Janus kinase (JAK) inhibitors on STAT1 phosphorylation in lymphocytes from the affected children and controls. RESULTS: We identified three mutations in exon 5 of TMEM173 in the six patients. Elevated transcription of IFNB1 and other gene targets of STING in peripheral-blood mononuclear cells from the patients indicated constitutive activation of the pathway that cannot be further up-regulated with stimulation. On stimulation with cGAMP, fibroblasts from the patients showed increased transcription of IFNB1 but not of the genes encoding interleukin-1 (IL1), interleukin-6 (IL6), or tumor necrosis factor (TNF). HEK293T cells transfected with mutant constructs show elevated IFNB1 reporter levels. STING is expressed in endothelial cells, and exposure of these cells to cGAMP resulted in endothelial activation and apoptosis. Constitutive up-regulation of phosphorylated STAT1 in patients' lymphocytes was reduced by JAK inhibitors. CONCLUSIONS: STING-associated vasculopathy with onset in infancy (SAVI) is an autoinflammatory disease caused by gain-of-function mutations in TMEM173. (Funded by the Intramural Research Program of the National Institute of Arthritis and Musculoskeletal and Skin Diseases; ClinicalTrials.gov number, NCT00059748.).

The E326K mutation and Gaucher disease: mutation or polymorphism?

Gaucher disease is caused by mutations in the gene for human glucocerebrosidase, a lysosomal enzyme involved in the intracellular hydrolysis of glucosylceramide. While over 150 different glucocerebrosidase mutations have been identified in patients with Gaucher disease, not all reported mutations have been fully characterized as being causative. One such mutation is the E326K mutation, which results from a G to A nucleotide substitution at genomic position 6195 and has been identified in patients with type 1, type 2 and type 3 Gaucher disease. However, in each instance, the E326K mutation was found on the same allele with another glucocerebrosidase mutation. Utilizing polymerase chain reaction (PCR) screening and restriction digestions of both patients with Gaucher disease and normal controls, we identified the E326K allele in both groups. Of the 310 alleles screened from patients with Gaucher disease, the E326K mutation was detected in four alleles (1.3%). In addition, screening for the E326K mutation among normal controls from a random population revealed that three alleles among 316 screened (0.9%) also carried the E326K mutation. In the normal controls with the E326K allele, the glucocerebrosidase gene was completely sequenced, but no additional mutations were found. Because the E326K mutation may be a polymorphism, we caution that a careful examination of any allele with this mutation should be performed to check for the presence of other glucocerebrosidase mutations.

Scanning for telomeric deletions and duplications and uniparental disomy using genetic markers in 120 children with malformations.

We screened 120 children with sporadic multiple congenital anomalies and either growth or mental retardation for uniparental disomy (UPD) or subtelomeric deletions. The screening used short tandem repeat polymorphisms (STRP) from the subtelomeric regions of 41 chromosome arms. Uninformative marker results were reanalyzed by using the next available marker on that chromosome arm. In total, approximately 25,000 genotypes were generated and analyzed for this study. Subtelomeric deletions of 1 Mb in size were excluded for 27 of 40 chromosome arms. Among the 120 subjects none was found to have UPD, but five subjects (4%, 95% confidence interval 1-9%) were found to have a deletion or duplication of one or more chromosome arms. We conclude that UPD is not a frequent cause of undiagnosed multiple congenital anomaly syndrome. In addition, we determined that 9p and 7q harbor chromosome length variations in the normal population. We conclude that subtelomeric marker analysis is effective for the detection of subtelomeric duplications and deletions, although it is labor intensive. Given a detection rate that is similar to prior studies and the large workload imposed by STRPs, we conclude that STRPs are an effective, but impractical, approach to the determination of segmental aneusomy given current technology.

Leucocyte copper, a marker of copper body status is low in coronary artery disease.

To elucidate the relationship between leucocyte copper as a reliable, sensitive index of copper body status and extent of atherosclerosis in patients with Coronary Artery Disease (CAD) the present case-control study was carried out. 80 subjects were studied (23 females and 57 males), aged between 30-70, due to have a angiography. Individual angiograms were scored by combining the individual scores in all the major coronary arteries into one score of a scale 1.00 for patency to 0.00 for severe CAD. Serum and leucocyte copper and zinc were determined by GFAAS. No significant difference between patients with advanced CAD and relatively normal arteries were observed in the lipid profile and levels of plasma copper. Leucocyte copper had a significant link with the severity of atherosclerosis which was independent of sex. There was a linear relationship between the degree of decreasing leucocyte copper concentration and angiogram score. These findings give support to the hypothesis that marginal copper status, assessed by decreased leucocyte copper level, is associated with developing CAD.

Glucocerebrosidase mutations among African-American patients with type 1 Gaucher disease.

While the inherited deficiency of the enzyme glucocerebrosidase (Gaucher disease) is panethnic in its distribution, there have not been studies of the mutations encountered in specific ethnic groups in the United States, other than those on Ashkenazi Jews. We present the clinical descriptions and genotypes of seven patients of African-American ancestry with type 1 Gaucher disease, and summarize the published literature regarding the genotypes encountered in this population. All seven of the patients had moderate-to-severe manifestations of the disease, and all developed symptoms by adolescence. Genotypic analyses revealed that no two probands shared the same genotype. The common mutations N370S, c.84-85insG, IVS2+1 G-->A, and R463C were not seen. Mutation L444P was present on one allele in each of the patients; but the same mutation was encountered as a single point mutation in three of the patients, and as part of a recombinant allele in four of the patients. Southern blot analyses revealed a glucocerebrosidase fusion allele in one patient, and a duplication resulting from recombination in the region downstream from the glucocerebrosidase gene in three of the patients. Five different point mutations (A90T, R48W, N117D, R170C, and V352L), one deletion mutation (c.222-224 delTAC), and one insertion mutation (c.153-154 insTACAGC) were encountered. Our results demonstrate that there is significant genotypic heterogeneity among African-American patients with type 1 Gaucher disease, and that recombinations in the glucocerebrosidase gene locus are particularly common in this patient group. Published 2001 Wiley-Liss, Inc.

Current gender stereotypes and their evaluative content.

A sample of 173 undergraduate students rated 300 attributes to assess current gender stereotypes, and a second independent sample (n = 57) evaluated the favorability of these attributes. Statistical results indicated that the college students ascribed different attributes to men and women. Although the content of gender stereotypes seems to have remained unchanged over the years, the value attached to stereotypic gender traits seems to be changing. More specifically, a greater number of unfavorable attributes were used to describe men than women, thereby creating a more negative masculine stereotype.

Analysis and classification of 304 mutant alleles in patients with type 1 and type 3 Gaucher disease.

Gaucher disease results from the inherited deficiency of the enzyme glucocerebrosidase (EC 3.2.1.45). Although >100 mutations in the gene for human glucocerebrosidase have been described, most genotype-phenotype studies have focused upon screening for a few common mutations. In this study, we used several approaches-including direct sequencing, Southern blotting, long-template PCR, restriction digestions, and the amplification refraction mutation system (ARMS)-to genotype 128 patients with type 1 Gaucher disease (64 of Ashkenazi Jewish ancestry and 64 of non-Jewish extraction) and 24 patients with type 3 Gaucher disease. More than 97% of the mutant alleles were identified. Fourteen novel mutations (A90T, N117D, T134I, Y135X, R170C, W184R, A190T, Y304X, A341T, D399Y, c.153-154insTACAGC, c.203-204insC, c.222-224delTAC, and c.1122-1123insTG) and many rare mutations were detected. Recombinant alleles were found in 19% of the patients. Although 93% of the mutant alleles in our Ashkenazi Jewish type 1 patients were N370S, c.84-85insG, IVS2+1G-->A or L444P, these four mutations accounted for only 49% of mutant alleles in the non-Jewish type 1 patients. Genotype-phenotype correlations were attempted. Homozygosity or heterozygosity for N370S resulted in type 1 Gaucher disease, whereas homozygosity for L444P was associated with type 3. Genotype L444P/recombinant allele resulted in type 2 Gaucher disease, and homozygosity for a recombinant allele was associated with perinatal lethal disease. The phenotypic consequences of other mutations, particularly R463C, were more inconsistent. Our results demonstrate a high rate of mutation detection, a large number of novel and rare mutations, and an accurate assessment of the prevalence of recombinant alleles. Although some genotype-phenotype correlations do exist, other genetic and environmental factors must also contribute to the phenotypes encountered, and we caution against relying solely upon genotype for prognostic or therapeutic judgements.

Pretreatment plasma HVA and haloperidol response in acute mania.

INTRODUCTION: Pretreatment plasma homovanillic acid (HVA) levels have been reported to be a correlate of clinical response to typical antipsychotics for schizophrenic, bipolar manic, and mixed groups of psychotic patients. Biological markers of clinical response to antipsychotics could be useful for optimizing drug treatment. METHOD: Thirty-one consenting acute inpatient subjects between ages 19 and 66 years with a DSM-III-R clinical diagnosis of bipolar disorder, manic with psychotic features were entered into this double-blind study and were randomly assigned to receive either haloperidol 25 mg/day or haloperidol 5 mg for the 3-week study. Subjects also received one of the following concomitant medications: standard lithium, lorazepam 4 mg/day, or placebo. RESULTS: The primary multiple regression analysis, including all subjects on both haloperidol doses, yielded a significant main effect for pretreatment plasma HVA (n=31, F=5.7, P=0.025), indicating that higher pretreatment plasma HVA was predictive of better clinical response. In addition, the interaction between haloperidol dose and pretreatment plasma HVA was also significantly associated with clinical response (F=12.59, P=0.0015). When the two haloperidol doses were analyzed separately, we found that pretreatment plasma HVA was only correlated with clinical response in the low haloperidol 5 mg/day group (n=18, F=11.73, P=0.0038) and was unrelated to clinical response to the high haloperidol 25 mg/day group. LIMITATIONS: The sample size was small. Results may have been confounded by prior antipsychotic treatment and concomitant use of lithium or lorazepam. DISCUSSION: These results suggest that pretreatment plasma HVA could be useful for dosing antipsychotics. Patients with high plasma HVA levels would be good candidates for low-dose treatment because they are more likely to improve on such a dose, while patients with low plasma HVA levels might warrant more rapid dosage escalation.

Life-threatening splenic hemorrhage in two patients with Gaucher disease.

Massive splenomegaly is a frequent finding in patients with Gaucher disease, the most common of the sphingolipidoses. Even so, the risk for splenic rupture and intracapsular hemorrhage has not been emphasized due to the rarity of this occurrence and the fibrotic, rubbery consistency of splenic tissue in these patients. We report two adult patients with type 1 Gaucher disease who suffered life-threatening splenic bleeds that were not acutely diagnosed. Both patients ultimately required emergent splenectomies. Factors complicating the diagnosis of splenic hemorrhage in patients with Gaucher disease are discussed. Published 2000 Wiley-Liss, Inc.

Mutation of a gene encoding a putative chaperonin causes McKusick-Kaufman syndrome.

McKusick-Kaufman syndrome (MKKS, MIM 236700) is a human developmental anomaly syndrome comprising hydrometrocolpos (HMC), postaxial polydactyly (PAP) and congenital heart disease (CHD). MKKS has been mapped in the Old Order Amish population to 20p12, between D20S162 and D20S894 (ref. 3). Here we describe the identification of a gene mutated in MKKS. We analysed the approximately 450-kb candidate region by sample sequencing, which revealed the presence of several known genes and EST clusters. We evaluated candidate transcripts by northern-blot analysis of adult and fetal tissues. We selected one transcript with widespread expression, MKKS, for analysis in a patient from the Amish pedigree and a sporadic, non-Amish case. The Old Order Amish patient was found to be homozygous for an allele that had two missense substitutions and the non-Amish patient was a compound heterozygote for a frameshift mutation predicting premature protein truncation and a distinct missense mutation. The MKKS predicted protein shows amino acid similarity to the chaperonin family of proteins, suggesting a role for protein processing in limb, cardiac and reproductive system development. We believe that this is the first description of a human disorder caused by mutations affecting a putative chaperonin molecule.

Type 2 Gaucher disease: the collodion baby phenotype revisited.

The association of Gaucher disease, the inherited deficiency of lysosomal glucocerebrosidase (EC 3.2.1.45), and congenital ichthyosis was first noted a decade ago. Subsequently, a null allele type 2 Gaucher mouse was generated that also exhibited ichthyotic skin, confirming that the skin disorder and enzyme deficiency were directly related. This paper details the clinical and molecular characterisation of 6 cases of type 2 Gaucher disease presenting with the collodion baby phenotype. The identified mutant glucocerebrosidase alleles include two novel mutations (S196P and R131L) and two rare point mutations (R120W and R257Q), as well as alleles resulting from recombination with the nearby glucocerebrosidase pseudogene. There is significant genotypic heterogeneity in this rare subset of patients with type 2 Gaucher disease. Gaucher disease should be considered in the differential diagnosis of congenital ichthyosis in the newborn period.

Coronary angiography in the presence of peripheral vascular disease: femoral or brachial/radial approach?

Peripheral vascular disease is considered a relative contraindication to the femoral approach for coronary angiography, but no data exist comparing the femoral and brachial/radial routes under these circumstances. We examined the influence of vascular approach on outcome. Two hundred and ninety-seven patients, mean age 67.1 +/- 8.4 years, with clinical or radiographic evidence of aortofemoral peripheral arterial disease underwent diagnostic coronary angiography during a 3-year period at this cardiothoracic center. The approach was successful in 121 of 154 femoral cases (79%) compared with 130 of 143 brachial/radial cases (91%; P < 0.01). Of the 33 failed femoral cases, 15 were then approached from the other femoral artery, with success in 6 (40%), while 18 were approached from the arm, with success in all (100%; P < 0.01). Brachial/radial cases took significantly longer than femoral cases (51 +/- 19 vs. 42 +/- 22 mins; P < 0.01). In cases where the femoral pulse was considered normal, the femoral approach nonetheless failed in 19 of 95 (20%). Major vascular complications (e.g., pulseless limb, arterial dissection, hemorrhage, or false aneurysm) occurred in nine femoral cases vs. zero brachial/radial cases (P < 0.01). Patients with peripheral vascular disease who undergo coronary angiography from the femoral artery have a 1-in-5 risk of procedural failure, necessitating use of an alternative vascular approach, and a 1-in-20 risk of a major vascular complication. Normality of femoral arterial pulsation is not a good predictor of femoral success. Brachial/radial approaches take longer, but succeed more frequently and have a negligible major vascular complication rate. We believe that patients with peripheral vascular disease should undergo coronary angiography via brachial or radial approach. Cathet. Cardiovasc. Intervent. 49:32-37, 2000.

Glucocerebrosidase gene mutations in patients with type 2 Gaucher disease.

Gaucher disease, the most common lysosomal storage disorder, results from the inherited deficiency of the enzyme glucocerebrosidase. Three clinical types are recognized: type 1, non-neuronopathic; type 2, acute neuronopathic; and type 3, subacute neuronopathic. Type 2 Gaucher disease, the rarest type, is progressive and fatal. We have performed molecular analyses of a cohort of 31 patients with type 2 Gaucher disease. The cases studied included fetuses presenting prenatally with hydrops fetalis, infants with the collodion baby phenotype, and infants diagnosed after several months of life. All 62 mutant glucocerebrosidase (GBA) alleles were identified. Thirty-three different mutant alleles were found, including point mutations, splice junction mutations, deletions, fusion alleles and recombinant alleles. Eleven novel mutations were identified in these patients: R131L, H255Q, R285H, S196P, H311R, c.330delA, V398F, F259L, c.533delC, Y304C and A190E. Mutation L444P was found on 25 patient alleles. Southern blots and direct sequencing demonstrated that mutation L444P occurred alone on 9 alleles, with E326K on one allele and as part of a recombinant allele on 15 alleles. There were no homozygotes for point mutation L444P. The recombinant alleles that included L444P resulted from either reciprocal recombination or gene conversion with the nearby glucocerebrosidase pseudogene, and seven different sites of recombination were identified. Homozygosity for a recombinant allele was associated with early lethality. We have also summarized the literature describing mutations associated with type 2 disease, and list 50 different mutations. This report constitutes the most comprehensive molecular study to date of type 2 Gaucher disease, and it demonstrates that there is significant phenotypic and genotypic heterogeneity among patients with type 2 Gaucher disease. Hum Mutat 15:181-188, 2000. Published 2000 Wiley-Liss, Inc.

Is the perinatal lethal form of Gaucher disease more common than classic type 2 Gaucher disease?

In recent years there has been increased recognition of a severe perinatal lethal form of Gaucher disease, the inherited deficiency of lysosomal glucocerebrosidase. We previously reported a case of severe type 2 Gaucher disease which was seen in a medical center in Rotterdam and now present three new cases from two other families seen at the same center. Mutational analyses of these cases revealed two novel mutations, H311R and V398F, located in exons 8 and 9, respectively. The identification of four cases of lethal type 2 Gaucher disease in a single center seems to be a function of increased awareness of this phenotype, rather than of geographic clustering. The actual incidence of lethal type 2 Gaucher disease may be underestimated, as many cases may have been misclassified as collodion babies or hydrops of unknown cause.

Two novel polymorphic sequences in the glucocerebrosidase gene region enhance mutational screening and founder effect studies of patients with Gaucher disease.

Gaucher disease, an inherited glycolipid storage disorder, is caused by a deficiency of the catabolic enzyme glucocerebrosidase (EC 3.2.1.45). The gene for human glucocerebrosidase is located on chromosome 1q21 and has a highly homologous pseudogene situated 16 kb downstream. We report two novel polymorphic sequences in the glucocerebrosidase gene region: the first consists of a variable number of dinucleotide (CT) repeats located 3.2 kb upstream from the glucocerebrosidase gene, and the second is a tetranucleotide (AAAT) repeat found between the glucocerebrosidase gene and its pseudogene, 9.8 kb downstream from the functional gene. These polymorphic sequences, along with a previously reported PvuII polymorphism in intron 6 of the glucocerebrosidase gene, were analyzed in patients with Gaucher disease (n=106) and in two normal control populations, one of Ashkenazi Jewish ancestry (n=72) and the second comprising non-Jewish individuals (n=46). In these samples, strong linkage disequilibrium was found between mutations N370S, c.84-85insG, and R463C and specific haplotypes; no significant linkage disequilibrium was found when examining haplotypes of patients with the L444P mutation. Studies of these polymorphic sites in several instances also led to the recognition of genotyping errors and the identification of unusual recombinant alleles. These new polymorphic sites provide additional tools for mutational screening and founder effect studies of Gaucher disease.

Hydrops fetalis: lysosomal storage disorders in extremis.

In recent years there has been an increased recognition that hydrops fetalis may be an extreme presentation of many of the lysosomal storage disorders. Hydrops fetalis, the excessive accumulation of serous fluid in the subcutaneous tissues and serous cavities of the fetus, has many possible etiologies, providing a diagnostic challenge for the physician. Ten different lysosomal storage disorders have now been diagnosed in infants with hydrops fetalis, including mucopolysaccharidosis (MPS) VII and IVA, type 2 Gaucher disease, sialidosis, GMI gangliosidosis, galactosialidosis, Niemann-Pick disease type C, disseminated lipogranulomatosis (Farber disease), infantile free sialic acid storage disease (ISSD), and mucolipidosis II (I-cell disease). Frequently, these inborn errors of metabolism are recognized only after the unfortunate recurrence of hydrops fetalis in several pregnancies of a family. Making the diagnosis relies on the physician having a high index of suspicion and ordering appropriate testing, which can often be performed prenatally. In several of these disorders, including MPS VII, infantile galactosialidosis, type 2 Gaucher disease, and ISSD, hydrops fetalis is a relatively common presentation. A greater physician awareness of hydrops fetalis as a presentation of lysosomal disease will facilitate establishing a diagnosis in cases that would have previously been considered idiopathic and will enable a better estimation of the incidence of this association. Lysosomal disorders are among the few causes of nonimmune hydrops fetalis in which an accurate recurrence risk can be ascertained. With an early and accurate diagnosis, genetic counseling and family planning can be offered in these difficult cases.