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Werner syndrome (WS), a type of progeria, is a hereditary condition caused by a mutation in the WRN gene. A 62-year-old Japanese woman was diagnosed with WS at the age of 32 and has been visiting the hospital for follow-up since the last 30 years. The patient developed diabetes at the age of 46, and at the age of 60, her body mass index increased from 20.1 to 22.7 kg/m 2 owing to her unhealthy eating habits; her visceral fat area at the age of 61 was 233 cm 2 . With dietary control, her body weight, including the visceral fat and subcutaneous fat, decreased at the age of 62, and her insulin secretion, obesity, and fatty liver improved. We conducted the oral glucose challenge test four times, including at the prediabetic stage, to evaluate the insulin-secretion ability. The patient's insulin resistance gradually increased for more than 14 years, and her insulin secretion ability began to decrease 14 years after her diabetes diagnosis. Despite a remarkable decrease in body weight and fat mass with dietary management, the psoas muscle index did not decrease significantly in proportion to the body weight or fat mass. However, muscle mass monitoring is important for preventing the progression of sarcopenia. Hence, gradual reduction of visceral fat and weight by dietary management may be useful in treating diabetes in patients with WS, particularly in those whose visceral fat is significantly increased.

Guanine-quadruplexes (G4) are stable tetra-stranded DNA structures that may cause DNA replication stress and inhibit gene expression. Defects in unwinding these structures by DNA helicases may result in telomere shortening and DNA damage. Furthermore, due to mutations in WRN helicase genes in Werner syndrome, G4 motifs are likely to be key elements in the expression of premature aging phenotypes. The methylation of DNA plays a significant role in the stability and occurrence of G4. Thus, G4 frequency and DNA methylation mechanisms may be affected by excesses or deficiencies in methyl donors such as folate. B-Lymphocytes from Werner patients (n = 5) and healthy individuals (n = 5) were cultured in RPMI medium under condition of folate deficiency (20 nM) or sufficiency (200 nM) for 14 days. Cells were fixed on microscope slides for immunofluorescent staining to measure G4 frequency and γH2AX (a marker of DNA strand breaks) intensity, using automated quantitative imaging fluorescent microscopy. There was a significant increase (p < 0.05) in G4 levels in Werner syndrome patients compared to healthy controls. Werner and control cells grown in 20 nM folate media also showed significant increases in G4 (p < 0.001) and γH2AX (p < 0.01) signals compared with the same cells grown in 200 nM folate. Control cells grown in 20 nM folate also showed a significant reduction in DNA methylation levels (P < 0.05). The results of this study suggest that the occurrence of DNA G4 structures can be modulated in vitro via nutrients with important roles in methylation., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Werner syndrome (WS) is a premature aging disorder caused by mutations in a RecQ-family DNA helicase, WRN. Mice lacking part of the helicase domain of the WRN orthologue exhibit many phenotypic features of WS, including metabolic abnormalities and a shorter mean life span. In contrast, mice lacking the entire Wrn protein (i.e. Wrn null mice) do not exhibit a premature aging phenotype. In this study, we used a targeted mass spectrometry-based metabolomic approach to identify serum metabolites that are differentially altered in young Wrn helicase mutant and Wrn null mice. An antibody-based quantification of 43 serum cytokines and markers of cardiovascular disease risk complemented this study. We found that Wrn helicase mutants exhibited elevated and decreased levels, respectively, of the anti-inflammatory cytokine IL-10 and the pro-inflammatory cytokine IL-18. Wrn helicase mutants also exhibited an increase in serum hydroxyproline and plasminogen activator inhibitor-1, markers of extracellular matrix remodeling of the vascular system and inflammation in aging. We also observed an abnormal increase in the ratio of very long chain to short chain lysophosphatidylcholines in the Wrn helicase mutants underlying a peroxisome perturbation in these mice. Remarkably, the Wrn mutant helicase protein was mislocalized to the endoplasmic reticulum and the peroxisomal fractions in liver tissues. Additional analyses with mouse embryonic fibroblasts indicated a severe defect of the autophagy flux in cells derived from Wrn helicase mutants compared to wild type and Wrn null animals. These results indicate that the deleterious effects of the helicase-deficient Wrn protein are mediated by the dysfunction of several cellular organelles.

We describe three patients with Werner's syndrome (WS), two of whom had been mistakenly diagnosed as having scleroderma. We would like to discuss briefly the importance of differentiation of these two disorders from each other.

Minor inflammation-driven aging (inflammaging) has been proposed to explain human aging mechanism. To study the inflammatory condition associated with normal human aging, highly sensitive CRP (hsCRP) was examined in the sera collected from 217 healthy Japanese individuals aged between 1 and 100years and 41 mutation-proven Japanese Werner syndrome (WS) patients. The serum hsCRP was assayed by ELISA. The serum hsCRP level increased significantly (p<0.001) with normal aging from both sexes. The serum hsCRP was significantly elevated in WS (mean±SE: 11.0±1.6μg/ml) compared with age-matched normal population (1.3±0.3μg/ml, p<0.001) and normal elderly population ages between 71 and 100years (4.2±0.7μg/ml, p<0.001). Both normal aging and WS were associated with minor inflammation that can be evaluated by serum hsCRP. WS may be a good candidate to study inflammaging., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Oxidative stress markers including pentosidine and homocysteine were examined comparing them with inflammation markers including highly sensitive C-reactive protein (hsCRP) and matrix metalloproteinase-9 (MMP-9) in serum from patients with Werner syndrome (WS) and healthy individuals. Elevation of serum pentosidine correlated significantly with normal aging in healthy individuals (p < 0.0004). Serum pentosidine in WS increased significantly compared with age-matched healthy individuals (p < 0.05). Serum homocysteine levels increased insignificantly with normal aging in healthy individuals and in WS compared with age-matched healthy individuals. As both pentosidine and homocysteine levels did not correlate with hsCRP or MMP-9, both oxidative stress markers may be differentially regulated by inflammation.

The pathophysiological process of natural human aging has not been studied adequately due to the lack of an appropriate human model. Since recent investigations have suggested that inflammation possibly contributes to the pathogenesis of age-related disorders including atherosclerosis, cancer, and diabetes mellitus, the term "inflammaging," a combination of "inflammation" and "aging," has been coined. Werner syndrome (WS), caused by the loss of function of RecQ3 DNA/RNA helicase, is a typical progeroid syndrome mimicking natural aging, although it is extremely rare outside of Japan. We sought to examine WS patients from an immunological/inflammatory perspective. Sera from 14 mutation-proven WS patients (ages: 33-70 years) and 21 healthy Japanese adults ages 15 to 95 years were examined with ELISA for soluble Fas ligand (sFasL) to compare conventional inflammation markers. With natural aging, a statistically significant correlation (p < 0.0001) was observed in the serum level of sFasL. The sFasL in WS, a level comparable to that in healthy elderly ages 83 to 95 years, had significantly increased (p < 0.05) compared to that in young healthy individuals ages 15 to 70 years. A significant correlation was noted between the serum levels of conventional inflammation markers such as CRP (p < 0.025), ESR (p < 0.024), and WBC count (p < 0.0085). In conclusion, an increased level of serum sFasL in natural aging and WS patients may suggest a common pathophysiological mechanism: inflammation. WS may be a good model for analyzing inflammaging.

N-glycan profiling of the human serum glycoproteins including immunoglobulin fraction on different age groups of healthy persons shows substantial changes with increasing age in three major N-glycan structures. In individuals more than 40-50 years of age, there is an increase in under-galactosylated glycans and a decrease in the core alpha-1,6-fucosylated bi-galactosylated biantennary structure. These three glycan structures are also substantially changed in a Werner syndrome patient, to a level comparable or even more pronounced than those observed in a healthy Italian centenarian population. These data show that the glycosylation machineries in both liver cells and B-cells are affected in a similar way by the aging process despite their highly different nature. The observed changes in the glycan structures are indicative that biosynthetic processes are at the basis of the changes, possibly together with changes in serum clearing of glycan-altered proteins. Our data suggest that measurement of the N-glycan level changes could provide a noninvasive surrogate marker for general health or for age-related disease progression, and for monitoring the improvement of health after therapy.

The age dynamics of stable chromosome aberration (SCA) frequency was analysed by fluorescent in situ hybridization (FISH) in human blood lymphocytes derived from donors, irradiated by low doses of ionizing radiation (Chernobyl clean-up workers, nuclear weapon testers, etc.) and patients with hereditary premature aging--Werner's syndrome and Hutchinson-Gilford's syndrome. It was found that the level of SCA was age-dependent and increased in irradiated persons. So, the SCA level may be really an index of a so-called "radiation senescence", and may show a real biological age of irradiated persons. The patients with Werner's syndrome demonstrate increased SCA level in blood lymphocytes, corresponding to the premature aging of the organisms. But in the case of another form of premature aging--Hutchinson--Gilford's syndrome-- no rise of SCA level was found. Some possible reasons of such results are discussed.

Elevations in urinary hyaluronan have been used as the principal laboratory indicator for diagnosis of Hutchinson-Gilford Progeria Syndrome (HGPS). Previous reports have provided evidence suggesting that children with HGPS have altered hyaluronan metabolism as indicated by a mean 17-fold increase in urinary hyaluronan over normal values. In addition, adults with Werner's syndrome have elevated urinary hyaluronan and even more prominent elevations in serum hyaluronan over age-matched controls. It is not known whether serum hyaluronan is elevated or whether serum hyaluronan levels correlate with urinary hyaluronan levels in children with HGPS. In a large cohort of 19 HGPS patients, we sought to confirm elevations in urinary hyaluronan concentration, to establish whether serum hyaluronan is elevated, to measure the size of urinary hyaluronan, and to determine whether serum or urine hyaluronidase levels are altered. We have analyzed urinary and serum hyaluronan levels in patients with HGPS and control patients (1) by using an enzyme-linked immunosorbent assay (ELISA)-like method in which sample hyaluronan in solution and hyaluronan in solid phase compete for a solution of biotinylated hyaluronan-binding protein, and (2) by fluorophore-assisted carbohydrate electrophoresis. The size of urinary hyaluronan was measured by using Sepharose CL-6B size exclusion chromatography. Serum and urinary hyaluronidases were evaluated quantitatively, by using ELISA, and qualitatively, by using a gel detection method. HGPS patients did not show a significant elevation in either urinary or serum hyaluronan. We detected no difference in the size of urinary hyaluronan between HGPS children and age-matched controls. Serum and urinary hyaluronidase levels were not significantly different in normal and HGPS patients. These studies indicate that neither serum nor urinary hyaluronan concentration is a reliable diagnostic or prognostic marker for HGPS and underscore a difference between adult and childhood progerias.

Background: There is a well-established association between Werner's syndrome (WS) and hyperhyaluronic aciduria; however, to date hyaluronan (HA) in the serum has not been statistically linked with WS. Recently, the gene that causes WS has been defined as a DNA helicase on chromosome 8, and 19 different mutations in WS patients have been identified. It is not known whether the mutation type of the Werner helicase gene affects the levels of serum and urine HA in WS patients., Objective: To evaluate the association of serum HA with WS, and the relationship of the serum and urine HA levels to the mutation type., Methods: HA both in the serum and the urine was measured in 40 patients with WS and 114 normal controls. The serum and urine HA were quantified by sandwich binding protein assay and competitive ELISA-like method, respectively. The muation on WS helicase gene was analyzed by mutant-allele-specific amplification and oligomer ligation assay., Results: WS patients showed significantly higher levels of HA in the serum (mean +/- SD: 115.7 +/- 119.8 ng/ml, p < 0.001) and urine (1,040.8 +/- 777.3 ng/mg creatinine, Cre, p < 0.001) than age-matched controls (serum HA: 15.8 +/- 14.2 ng/ml, urine HA: 379.7 +/- 124.2 ng/mg Cre). The serum and urine HA levels of WS patients are almost equal to those of normal controls over 80 years (serum HA: 118.5 +/- 108.4 ng/ml, urine HA: 914.5 +/- 712.1 ng/mg Cre). There was a significant correlation between serum and urine HA levels in WS patients (r = 0.42, p = 0.007). Analysis of the mutation on the helicase gene in 22 WS patients showed that among 44 chromosomes, 3 (6.8%) chromosomes had type 1 mutation, 22 (50.0%) had type 4 mutation, 14 (31.8%) had type 6 mutation, and the rest had other mutations. The serum and urine levels of HA did not show any significant association with the mutation type., Conclusion: The hyperhyaluronic aciduria in WS reflects the high level of serum HA. The serum and urine HA levels are useful biochemical markers for WS irrespective of the muation type of the Werner helicase gene.

Werner's syndrome (WS) is an autosomal recessive disorder displaying many features consistent with accelerated ageing. Fibroblasts from WS patients show a distinct mutator phenotype (characterised by the production of large chromosomal deletions) and a profound reduction in proliferative capacity. The disorder results from a mutation in a novel ReqQ helicase. Recently, we demonstrated that the proliferative defect was corrected by the ectopic expression of telomerase. From these data, we propose that mutations in the wrn gene lead to deletions at or near the telomere which reduce the cells replicative life-span. This hypothesis predicts that cell types which retain the ability to upregulate telomerase as part of their response to a proliferative stimulus would fail to show any significant effect of wrn gene mutations upon life-span. Human T lymphocytes represent a well-characterised example of such a cell type. To test the hypothesis, WS T lymphocytes were cultured until they reached replicative senescence. These cultures displayed life-spans which did not differ significantly from those of normal controls. These findings are consistent with the hypothesis that the effects of wrn mutations on replicative life-span are telomere-mediated.

Three human RecQ DNA helicases, WRN, BLM and RTS, are involved in the genetic disorders associated with genomic instability and a high incidence of cancer. RecQL1 and RecQL5 also belong to the human RecQ helicase family, but their correlation with genetic disorders, if any, is unknown. We report here that in human B cells transformed by Epstein-Barr virus (EBV), human fibroblasts and umbilical endothelial cells transformed by simian virus 40, the expression of WRN, BLM, RTS and RecQL1 was sharply up-regulated. In B cells this expression was stimulated within 5-40 h by the tumor promoting agent phorbol myristic acetate (PMA). Interestingly, RecQL5beta, an alternative splicing product of RecQL5 with a nuclear localization signal, is expressed in resting B cells without significant modulation of its synthesis by EBV or PMA, suggesting it has a role in resting cells. We also roughly determined the number of copies per cell for the five RecQ helicase in B cells. In addition, levels of the different RecQ helicases are modulated in different ways during the cell cycle of actively proliferating fibroblasts and umbilical endothelial cells. Our results support the view that the levels of WRN, BLM, RTS and RecQL1 are differentially up-regulated to guarantee genomic stability in cells that are transformed or actively proliferating.

The patient was a 39-year-old Japanese male with a body height of 160 cm and weight of 48 kg who was diagnosed as Werner syndrome of homozygote for mutation 4. His plasma total cholesterol (TC), triglycerides (TGs), high density lipoprotein-cholesterol (HDL-C) and apolipoprotein A-I (apo A-I) levels were 7.2, 2.1, 1 mmol/l and 128 mg/dl, respectively. During the clinical course of treatment of this patient, his plasma levels of HDL-C and apo A-I declined drastically to levels of as low as 0.2 mmol/l and 10 mg/dl, respectively, with concurrent reciprocal increase in plasma TG levels. Plasma HDL-C, apo A-I and TG levels gradually returned to original values. Lipoprotein lipase activity and mass in post-heparin plasma were markedly low when the apo A-I and HDL-C levels decreased to 10 mg/dl and 0.21 mmol/l, respectively, and these values improved when the apo A-I and HDL-C levels returned to more normal values of 106 mg/dl and 0.94 mmol/l, respectively. The result of direct sequence of the exon 3 and 4, and the promoter region of the apo A-I gene of the patient revealed no single nucleotide changes. These results suggest that in the present patient, impaired hydrolysis of TGs in TG-rich lipoproteins, is due at least in part to a decreased LPL enzyme level, reduced the formation of nascent HDL, resulting in unusually low plasma levels of HDL-C and apo A-I.

We review recent data on senescence and immortalization of human B-lymphoblastoid cell lines (LCLs) transformed by the Epstein-Barr virus (EBV). Although EBV-transformed LCLs are generally believed to be immortalized, a series of recent studies, including ours, provided strong evidence that they are mostly mortal and have non-malignant properties, except for a small proportion of LCLs that are immortalized by developing a strong telomerase activity. A large proportion of mortal LCLs have exceptionally long lifespans. Some of them have a lifespan over 150 population-doubling levels, keeping a relatively constant telomere length in spite of the absence of a detectable telomerase activity, suggesting that they maintain telomeres by a pathway other than that using telomerase. Here we propose a model of an alternative pathway to maintain telomeres of such long-lived mortal LCLs by exploiting extra-chromosomal telomere repeat DNA, which was recently found by us.

Werner syndrome is a rare premature aging syndrome accompanied by severe atherosclerosis. The etiology of atherosclerosis is suspected to be due to its complications, namely diabetes mellitus, hyperinsulinemia and hyperlipidemia. But from an autopsy case we found that some other risk factors may be involved in the mechanism of atherosclerosis in this syndrome. Previously we revealed that the plasminogen activator inhibitor-1 (PAI-1) gene was being overexpressed in skin fibroblasts from a patient with this syndrome. PAI-1 is a potent inhibitor of tissue plasminogen activator and a possible risk factor of atherosclerosis. This led us to assess the plasma concentration of PAI-1. Our working hypothesis was that the PAI-1 gene was upregulated or not fully suppressed in cells responsible for the production of PAI-1 in plasma as well as in fibroblasts. The results show a high concentration of plasma PAI-1. One of the well-known physiological substances that induce the PAI-1 gene is tumor necrosis factor-alpha, which also induces other possible risk factors of atherosclerosis, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1. We found the serum concentrations of ICAM-1 to be elevated in patients with this syndrome. We conclude that high concentrations of PAI-1 and ICAM-1 in blood may be one of the potent causes of severe atherosclerosis in Werner syndrome.

DNA topoisomerase II is involved in DNA topologic changes through the formation of a cleavable complex. This is stabilized by the antitumor drug VP16, which results in DNA breakage, aberrant recombination, and cell death. In this work, we compare the chromosomal damage induced by VP16 with that induced by bleomycin (BLM) in lymphoblasts from patients affected by the chromosome breakage syndromes ataxia telangiectasia (AT), xeroderma pigmentosum (XP), and Bloom syndrome (BS), and by the progeroid syndromes Werner (WS) and Cockayne (CS). Patients affected by AT, XP, BS, and WS have a greatly enhanced risk of developing cancer. The results show that AF and WS cells are hypersensitive to VP16, as revealed in the higher proportion of metaphases showing exchange figures and more than two breaks. All lines except AT and one CS line showed normal sensitivity to BLM. Our data on the sensitivity to VP16 of all these mutant cells underline the fact that VP16 damage is amplified only in cells that have abnormal illegitimate recombination (i.e., AT and WS).

To assess the risk factors for atherosclerosis in Werner's syndrome (WS), coagulation/fibrinolytic system parameters and lipid levels were investigated in 9 non-smoker patients with WS and compared with normal control values (N). The levels of thrombin antithrombin III complex (p < 0.05), D-dimer (p < 0.05), tissue plasminogen activator (p < 0.005) and PA inhibitor 1 (p < 0.01) were significantly increased, while the level of thrombomodulin (p < 0.005) in the fasting plasma was significantly decreased in the WS cases compared with N. Lipid profiles confirmed that 8 of the 9 patients were of hyperlipidemia type IIb, 7 had hyperinsulinemia and 5 fulfilled the criteria for clinical diabetes mellitus. The hypercoagulable condition suggested the existence of multiple risk factors for atherosclerosis in WS in addition to the previously reported hyperinsulinemia and hyperlipidemia.

Werner syndrome (WS) is a rare autosomal recessive disorder characterized by prematurely aged appearance. Genetic linkage analysis has placed the relevant gene in subchromosomal band 8p12. DNA polymerase beta gene has been mapped to chromosome 8p12-11 and thought to be involved in DNA repair and possibly in recombination. Somatic cells from WS patients exhibit chromosomal instability, a markedly reduced replicative life span and slow growth. The functions of DNA polymerase beta gene and its position prompted us to examine this gene in WS patients. We have found the novel DNA polymerase beta cDNA species in blood samples from WS patients, which contain 107 bp insertions or 87 bp deletions in the catalytic domain of DNA polymerase beta. These mutations change the structure of DNA polymerase beta and thus the capacity of the DNA repair system would be impaired, which may account for the high mutation rate observed in WS.

Objective: GH and growth hormone binding protein in Werner's syndrome were investigated to elucidate their relation to the short stature., Design: The levels of GH binding protein and GH response to insulin-induced hypoglycaemia were determined. GH binding protein levels and its Scatchard analysis in Werner's syndrome were compared with those in normal subjects., Patients: Three patients with Werner's syndrome (one man aged 45 years and two women aged 39 and 38 years) and 41 normal subjects (18 men and 23 women aged 39.3 +/- 5.5 years, mean +/- SD) were studied., Measurements: GH binding protein levels were determined using an Ultrogel AcA44 minicolumn and GH levels were measured by a highly sensitive enzyme immunoassay., Results: Two out of three patients with Werner's syndrome had GH binding protein levels above the mean +/- 2SD value in normal subjects. GH secretion was impaired in Werner's syndrome as judged by the low GH response to insulin-induced hypoglycaemia., Conclusions: Elevated GH binding protein levels may lead to an increase in the bound form of GH, which is probably less bioactive, resulting in growth failure in association with the impaired GH secretion in Werner's syndrome.

Insulin resistance in Werner's syndrome (WS) was studied using the glucose clamp technique, and compared with physiologically aged and young subjects. Fasting immuno-reactive insulin (IRI) was increased in patients with Werner's syndrome compared with aged and young subjects. Metabolic clearance rate (MCR) of glucose was decreased in the aged and WS. A rightward shift of the dose-response curves of insulin and MCR of glucose was observed in the aged and WS with a more pronounced shift in the latter. MCR of insulin was also decreased in WS. [125I]insulin binding to erythrocytes was similar in the three groups. These results suggest that insulin resistance associated with WS is due to a post-binding defect manifested by a rightward shift of the dose-response curve of insulin-induced glucose disposal and a decrease in insulin clearance rate.

Abnormal lipoprotein metabolism and its cause were studied in 10 patients with Werner's syndrome. Seven of the 10 patients had hypercholesterolemia (greater than 250 mg dl-1). A significant positive correlation (p less than 0.01) was found between serum total cholesterol levels and LDL receptor activity. In monocyte-derived macrophages of patients with this syndrome, the uptake, lysosomal hydrolysis and re-esterification of free cholesterol are enhanced. This abnormal accumulation of cholesterol ester may cause accelerated conversion of macrophages to foam cells in Werner's syndrome.

We present the study of a patient suffering insulin resistance with Werner's syndrome which had abnormal glucose tolerance determined by euglycemic fixing ("Clamp") using an artificial pancreas (Biostator GCIIS, Miles Martin) and the binding of insulin to its receptor in erythrocytes. The results obtained show a dose-response curve of insulin serum levels to dextrose infusion rate, shifted significantly to the right and bottom which indicates a diminished insulin sensitivity; similarly a moderate decrease in receptor is obtained with no decrease in their affinity and absence of abnormalities in contraregulatory hormones. These results are compatible with a postreceptor alteration.

Studies were made on the abnormality of lipoprotein metabolism and its cause in 10 patients with Werner's syndrome. Seven of the 10 patients had hypercholesterolaemia (above 250 mg dl-1). Six of the seven patients with hypercholesterolaemia had thickened Achilles' tendons (greater than 9 mm). A significant positive correlation (P less than 0.01) was found between the serum total cholesterol levels and the thickness of Achilles' tendons in these 10 patients, suggesting that the substance precipitated in the thickened tendons is derived from serum cholesterol. Some first-degree relatives of three patients with both hypercholesterolaemia and xanthoma-like thickening of Achilles' tendons also suffered from hypercholesterolaemia. Moreover, the low density lipoprotein (LDL) receptor activities in peripheral lymphocytes of five patients with both hypercholesterolaemia and xanthoma-like tendons were significantly (P less than 0.001) lower than those of controls, whereas the LDL receptor activities of two patients without hypercholesterolaemia were almost the same as those of controls. These findings suggest that, at least these five patients with lower lymphocyte LDL receptor activities, and probably another patient with both hypercholesterolaemia and xanthoma-like thickening of Achilles' tendons suffered from familial hypercholesterolaemia (FH). If this is the case, this high frequency of association of Werner's syndrome with FH (in six of 10 patients) suggests some relationship between these two diseases.

The tissue sensitivity to insulin and the metabolic clearance rate of insulin were evaluated in a new case of Werner's syndrome, by means of the euglycemic clamp technique using an artificial endocrine pancreas. The possible contribution of pancreatic insulin secretion and of reduced hepatic insulin breakdown to the peripheral hyperinsulinism was also studied by simultaneous determination of C-peptide and insulin levels and by analysis of the C-peptide/insulin molar ratio. The data obtained are consistent with the presence of marked insulin resistance and of reduced metabolic clearance rate of insulin in our patient, confirming what has been observed previously using other techniques. The finding of a reduced C-peptide to insulin ratio during the intravenous glucose tolerance test suggests that hepatic extraction of insulin could also be altered in Werner's syndrome, as found in several hyperinsulinism conditions, therefore contributing to peripheral hyperinsulinism.

Four cases of accelerated aging, known as Werner's syndrome (WS), are presented. Plasma lipid and lipoprotein findings and lipid compositions of red cell membranes are compared to those of old and young healthy people. Typical clinical findings, as reported in the literature, confirmed the diagnosis. Furthermore, an elevated osmotic fragility of red blood cells (RBC) was observed. RBC membranes of WS patients showed phospholipid concentrations similar to those found in people aged over 70 years, which have been reported to be lower than those of mature donors aged below 30 years. Fatty acid patterns of RBC membranes and plasma phospholipids were indicative of some disturbance in phospholipid subclass distributions in WS-patients.

Premature development of atherosclerosis is a fatal complication together with cancer in patients with Werner' syndrome, a known syndrome of premature aging. Proliferation of arterial smooth muscle cells (SMC) is a key event in the formation of atherosclerosis. This paper studied the serum growth promoting activity of Werner's syndrome in cultured rabbit aortic SMC. Serum of cases of Werner's syndrome showed significantly higher growth-promoting activities than those of age-matched controls. The levels of serum growth hormone, plasma somatomedin C, and urine epidermal growth factor were not increased in Werner's syndrome. The level of serum insulin was generally increased in Werner's syndrome. However, there was no correlation between serum insulin level and growth promoting activity, and the effective dose of insulin for the promotion of SMC growth was ten thousand times higher than that contained in the serum. The above results suggest that elevated serum growth-promoting activity in Werner's syndrome is in part responsible for the premature atherosclerosis and that this elevated activity is derived from either platelet derived growth factor or an unknown growth factor(s) contained in the serum.

A significant improvement of hyperinsulinemic diabetes was observed in a patient with Werner's syndrome after the removal of a large parasagittal meningioma. To our knowledge this is the second case reported in the literature. The possible relationship between meningioma and diabetes in Werner's syndrome is discussed.

3 cases of Werner's syndrome were studied on serum lipid metabolism in comparison with clinical manifestation. These 3 cases showed hyperlipemia all with hypertriglycemia and increased level of very low density lipoprotein. The lipoprotein lipase activity was shown after intravenous injection of heparin. These results indicate a possible pathogenic facet of Werner's syndrome from the lipid metabolic point of view.

X-ray irradiation induced sister chromatid exchanges (SCE) in blood lymphocytes from patient with Down's syndrome and adult progeria (in both the cases radioresistant DNA synthesis takes place). In these diseases, likely as upon form II of xeroderma pigmentosum (the replicative DNA synthesis is radioresistant), X-ray irradiation lowers the rate of SCE compared with that in the control, then the SCE rate rises with the increase in radiation dose, reaching the rate of SCE in non-irradiated cells. In normal lymphocytes (in which ionizing radiation inhibits the replicative synthesis of DNA) the rate of SCE rises with the rise of radiation dose. Thus, the rate of SCE in X-ray irradiated lymphocytes is in reverse dependence with radioresistance of replicative synthesis of DNA. The data obtained are explained in accordance with the replicative hypothesis of the SCE nature (Painter, 1980a): in cells of patients with Down's syndrome, xeroderma pigmentosum form II and progeria of adults the time of existence of partly replicated clusters of replicons is decreased due to radioresistant replicative synthesis of DNA, but the presence of partly replicated clusters of replicons is necessary for SCE formation. Therefore the rate of SCE in X-irradiated cells of these patients decreases.

Different cellular parameters used to detect genetic instability were analyzed in lymphocytes from a patient affected by Werner's syndrome (WS). Cytogenetic studies indicated the presence of structural and numerical chromosomal abnormalities and the occurrence of variegated translocation mosaicism. The baseline mutation frequency was similar to that observed in normal donor samples. DNA repair investigations showing a normal capability to perform UV-induced DNA repair synthesis and a normal sensitivity to various mutagens (UVC light, mono- and bi-functional alkylating agents) indicate that different DNA repair mechanisms act normally in WS. In this feature, WS appears to differ from the other genetically determined syndromes in which chromosomal instability is associated with a marked hypersensitivity to specific DNA-damaging agents.