SGMS2 in primary osteoporosis with facial nerve palsy.

Pathogenic heterozygous variants in SGMS2 cause a rare monogenic form of osteoporosis known as calvarial doughnut lesions with bone fragility (CDL). The clinical presentations of SGMS2-related bone pathology range from childhood-onset osteoporosis with low bone mineral density and sclerotic doughnut-shaped lesions in the skull to a severe spondylometaphyseal dysplasia with neonatal fractures, long-bone deformities, and short stature. In addition, neurological manifestations occur in some patients. SGMS2 encodes sphingomyelin synthase 2 (SMS2), an enzyme involved in the production of sphingomyelin (SM). This review describes the biochemical structure of SM, SM metabolism, and their molecular actions in skeletal and neural tissue. We postulate how disrupted SM gradient can influence bone formation and how animal models may facilitate a better understanding of SGMS2-related osteoporosis.

A multi-omics study to characterize the transdifferentiation of human dermal fibroblasts to osteoblast-like cells.

Background: Various skeletal disorders display defects in osteoblast development and function. An in vitro model can help to understand underlying disease mechanisms. Currently, access to appropriate starting material for in vitro osteoblastic studies is limited. Native osteoblasts and their progenitors, the bone marrow mesenchymal stem cells, (MSCs) are problematic to isolate from affected patients and challenging to expand in vitro. Human dermal fibroblasts in vitro are a promising substitute source of cells. Method: We developed an in vitro culturing technique to transdifferentiate fibroblasts into osteoblast-like cells. We obtained human fibroblasts from forearm skin biopsy and differentiated them into osteoblast-like cells with ß-glycerophosphate, ascorbic acid, and dexamethasone treatment. Osteoblastic phenotype was confirmed by staining for alkaline phosphatase (ALP), calcium and phosphate deposits (Alizarin Red, Von Kossa) and by a multi-omics approach (transcriptomic, proteomic, and phosphoproteomic analyses). Result: After 14 days of treatment, both fibroblasts and MSCs (reference cells) stained positive for ALP together with a significant increase in bone specific ALP (p = 0.04 and 0.004, respectively) compared to untreated cells. At a later time point, both cell types deposited minerals, indicating mineralization. In addition, fibroblasts and MSCs showed elevated expression of several osteogenic genes (e.g. ALPL, RUNX2, BMPs and SMADs), and decreased expression of SOX9. Ingenuity Pathways Analysis of RNA sequencing data from fibroblasts and MSCs showed that the osteoarthritis pathway was activated in both cell types (p_adj. = 0.003 and 0.004, respectively). Discussion: These data indicate that our in vitro treatment induces osteoblast-like differentiation in fibroblasts and MSCs, producing an in vitro osteoblastic cell system. This culturing system provides an alternative tool for bone biology research and skeletal tissue engineering.

Abnormal Bone Tissue Organization and Osteocyte Lacunocanalicular Network in Early-Onset Osteoporosis Due to SGMS2 Mutations.

Pathological variants in SGMS2, encoding sphingomyelin synthase 2 (SMS2), result in a rare autosomal dominant skeletal disorder with cranial doughnut lesions. The disease manifests as early-onset osteoporosis or a more severe skeletal dysplasia with low bone mineral density, frequent fractures, long-bone deformities, and multiple sclerotic cranial lesions. The exact underlying molecular features and skeletal consequences, however, remain elusive. This study investigated bone tissue characteristics in two adult males with a heterozygous SGMS2 mutation p.Arg50* and significant bone fragility. Transiliac bone biopsy samples from both (patient 1: 61 years; patient 2: 29 years) were analyzed by bone histomorphometry, confocal laser scanning microscopy, and quantitative backscattered electron imaging (qBEI). Bone histomorphometry portrayed largely normal values for structural and turnover parameters, but in both patient 1 and patient 2, respectively, osteoid thickness (-1.80 SD, -1.37 SD) and mineralizing surface (-1.03 SD, -2.73 SD) were reduced and osteoid surface increased (+9.03 SD, +0.98 SD), leading to elevated mineralization lag time (+8.16 SD, +4.10 SD). qBEI showed low and heterogeneous matrix mineralization (CaPeak -2.41 SD, -3.72 SD; CaWidth +7.47 SD, +4.41 SD) with a chaotic arrangement of collagenous fibrils under polarized light. Last, osteocyte lacunae appeared abnormally large and round in shape and the canalicular network severely disturbed with short-spanned canaliculi lacking any orderliness or continuity. Taken together, these data underline a central role for functional SMS2 in bone matrix organization and mineralization, lacunocanalicular network, and in maintaining skeletal strength and integrity. These data bring new knowledge on changes in bone histology resulting from abnormal sphingomyelin metabolism and aid en route to better understanding of sphingolipid-related skeletal disorders. © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Variant rs6749447 (T > G) in the serine threonine kinase gene is associated with cardiovascular complications, the Tampere adult population cardiovascular risk study.

ABSTRACT: We have previously shown an association of STK39 (serine threonine kinase) rs6749447 (T > G) with hypertension in the Tampere adult population cardiovascular risk study in 50-year-old subjects. These 1196 subjects were followed up to the age of 65 years to determine whether rs6749447 is also associated with coronary artery disease (CAD), transient ischemic attack (TIA), or early cardiovascular death.DNA samples were collected by buccal swabs and genotypes were determined by PCR. Hypertension, TIA, and CAD were determined by questionnaire and the National Hospital Discharge Registry. Outcomes for death were collected from the National Statistics Centre. Linkage disequilibrium analysis and gene expression correlations for rs6749447 were done in silico.After following the subjects up to the age of 60 years the rs6749447 G-allele still associated with hypertension (P = .009). The variation did not associate with CAD (P = .959). The risk for TIA was 5.2-fold among G-allele carriers compared to TT genotype even after adjusting for body mass index (P = .036, 95% CI 1.11-24.59). After follow-up of the subjects to the age of 65 years, adjusting for body mass index, the G-allele was associated with 3.2-fold risk of premature cardiovascular death (P = .049, 95% CI 1.00-10.01).In conclusion, the STK39 genetic variant rs6749447 was significantly associated with TIA and premature cardiovascular death in a Finnish cohort. The in silico results of linkage disequilibrium and gene expression analyses also showed associations that were distinct from the retention of salt effect on kidneys proposed earlier for this intronic variation.

Germline EMSY sequence alterations in hereditary breast cancer and ovarian cancer families.

BACKGROUND: BRCA1 and BRCA2 mutations explain approximately one-fifth of the inherited susceptibility in high-risk Finnish hereditary breast and ovarian cancer (HBOC) families. EMSY is located in the breast cancer-associated chromosomal region 11q13. The EMSY gene encodes a BRCA2-interacting protein that has been implicated in DNA damage repair and genomic instability. We analysed the role of germline EMSY variation in breast/ovarian cancer predisposition. The present study describes the first EMSY screening in patients with high familial risk for this disease. METHODS: Index individuals from 71 high-risk, BRCA1/2-negative HBOC families were screened for germline EMSY sequence alterations in protein coding regions and exon-intron boundaries using Sanger sequencing and TaqMan assays. The identified variants were further screened in 36 Finnish HBOC patients and 904 controls. Moreover, one novel intronic deletion was screened in a cohort of 404 breast cancer patients unselected for family history. Haplotype block structure and the association of haplotypes with breast/ovarian cancer were analysed using Haploview. The functionality of the identified variants was predicted using Haploreg, RegulomeDB, Human Splicing Finder, and Pathogenic-or-Not-Pipeline 2. RESULTS: Altogether, 12 germline EMSY variants were observed. Two alterations were located in the coding region, five alterations were intronic, and five alterations were located in the 3'untranslated region (UTR). Variant frequencies did not significantly differ between cases and controls. The novel variant, c.2709 + 122delT, was detected in 1 out of 107 (0.9%) breast cancer patients, and the carrier showed a bilateral form of the disease. The deletion was absent in 897 controls (OR = 25.28; P = 0.1) and in 404 breast cancer patients unselected for family history. No haplotype was identified to increase the risk of breast/ovarian cancer. Functional analyses suggested that variants, particularly in the 3'UTR, were located within regulatory elements. The novel deletion was predicted to affect splicing regulatory elements. CONCLUSIONS: These results suggest that the identified EMSY variants are likely neutral at the population level. However, these variants may contribute to breast/ovarian cancer risk in single families. Additional analyses are warranted for rare novel intronic deletions and the 3'UTR variants predicted to have functional roles.

Minor variant of rs 16827043 in the iron regulator hemojuvelin gene (HJV) contributes to hypertension: The TAMRISK study.

It is known that iron overload may lead to an increased risk for many diseases. According to GWAS studies, iron regulatory protein HFE gene variant H63D (rs1799945) was associated with hypertension, an observation which we were able to confirm also in our TAMRISK cohort. Thus, it is possible that abnormalities in iron homeostasis may predispose to hypertension. This prompted us to study whether there is an association between hypertension and another iron overload-associated gene, hemojuvelin (HJV), which has 2 common polymorphic sites (rs 16827043, rs7536827).The study included 336 hypertensive cases and 480 controls. All participants were 50- year-old Finnish men and women, and the data was collected from the Tampere adult population cardiovascular risk study (TAMRISK). Genotypes were determined using Competitive Allelic Specific PCR (KASP).We found that the minor variant of the HJV polymorphic site rs16827043 (G-allele) is a statistically significant factor associated with hypertension among 50 year-old individuals compared with the AA genotype carriers (OR = 1.66, 95% CI: 1.06 - 2.60, P = 0.03). The risk was even higher when overweight subjects (BMI>30) were excluded from the analyses. For the other polymorphic variant rs7536827, association with hypertension was found only among normal or slightly overweight A-allele carriers.In conclusion, HJV genetic variants were associated with essential hypertension in Finnish subjects from the TAMRISK cohort. Previous studies together with the present one indicate that individuals with possible dysregulation of iron metabolism may have higher risk for hypertension than those with normal iron homeostasis.

Whole-exome sequencing of Finnish hereditary breast cancer families.

A remarkable proportion of factors causing genetic predisposition to breast cancer (BC) are unknown in non-BRCA1/2 families. Exome sequencing was performed for 13 high-risk Finnish hereditary breast and/or ovarian cancer (HBOC) families to detect variants contributing to BC susceptibility. After filtering, 18 candidate variants in DNA damage response (DDR) pathway genes were screened in 129 female HBOC patients, up to 989 female controls, and 31 breast tumours by Sanger sequencing/TaqMan assays. In addition, two variants were further studied in 49 male BC patients and 909 male controls. Second, all variants predicted to affect function in six early-onset BC patients were analysed in detail. Variants in ATM, MYC, PLAU, RAD1, and RRM2B were enriched in female HBOC patients compared with controls (odds ratio 1.16-2.16). A rare nonsynonymous variant in RAD50 was detected in a male BC patient. In addition, a very rare BRCA1 variant was identified in a single high-risk family. None of the variants showed wild-type allele loss in breast tumours. Furthermore, novel variants predicted to affect function were detected in early-onset patients in genes, which target DNA repair and replication, signalling, apoptosis, and cell cycle pathways. Family-specific enrichment of multiple DDR pathway gene defects likely explains BC predisposition in the studied families. These findings provide new information on potential BC-related pathways and an excellent premise for future studies.

Glucuronic Acid Epimerase (GLCE) Variant rs3865014 (A>G) Is Associated with BMI, Blood Hemoglobin, Hypertension, and Cerebrovascular Events, the TAMRISK Study.

Heparan sulfate proteoglycans modulate many physiological systems, and genes responsible for proteoglycan assembly and disassembly may affect their interaction. We sought to determine whether polymorphisms of the glucuronic acid epimerase (GLCE) rs3865014 and sulfatase-2 (SULF2) rs2281279, genes coding for enzymes participating in heparan sulfate side chain activity, associate with hypertension, selected cardiometabolic risk factors and cardiovascular events in the Tampere adult population cardiovascular risk study. A Finnish cohort of 339 subjects with diagnosed hypertension and 441 controls was analyzed. Samples were genotyped for GLCE rs3865014 (A>G) and SULF2 rs2281279 (T>C) polymorphisms using competitive allele-specific PCR (KASP) technique. Prevalence of ischemic heart diseases (I20-I25) and incidence of cerebrovascular diseases (I60-I69) and transient cerebral ischemic attacks (TIA) (G45) were followed up until the subjects were on the average 60 years old. GLCE rs3865014 G allele showed negative association with hypertension (p = 0.022), waist circumference (p = 0.032), BMI (p = 0.048), and positive association with hemoglobin (p = 0.029), low-density lipoprotein cholesterol (p = 0.031), and frequency of cerebrovascular events (p = 0.011). SULF2 rs2281279 showed no association with the studied parameters. The GLCE gene polymorphism rs3865014 appears to have biological relevance in human pathophysiology.

Decorin Genotypes, Serum Glucose, Heart Rate, and Cerebrovascular Events: The Tampere Adult Population Cardiovascular Risk Study.

BACKGROUND: Decorin is an extracellular matrix proteoglycan that may attenuate progression of atherosclerosis and its complications, such as stroke. Among its multitude of functions, decorin has been suggested to serve as a receptor for resistin, an adipokine involved in energy homeostasis. The GG genotype of the decorin polymorphism rs7308752 (A>G) and the CC genotype of the rs516115 (T>C) are associated with decreased plasma resistin levels. AIMS: The association of the above decorin genotypes with selected cardiometabolic risk factors and cerebrovascular events was studied in the Tampere adult population cardiovascular risk (TAMRISK) study. MATERIALS AND METHODS: A Finnish cohort of 336 subjects with diagnosed hypertension and 444 controls was analyzed. Samples were genotyped for decorin rs7308752 and rs516115 polymorphisms using a Competitive Allele-Specific PCR (KASP) technique. Cerebrovascular diseases (I60-I69), including transient cerebral ischemic attacks (G45), were followed up from 2005 to 2014. RESULTS: Subjects with either of decorin rs7308752 genotypes AG/GG had higher serum glucose (p = 0.015) and higher heart rate (p = 0.017) than those with AA genotype. Similarly, decorin rs516115 genotypes TC/CC were associated with higher serum glucose (p = 0.034) and higher frequency of cerebrovascular diseases (p = 0.015) compared to the TT genotype. However, decorin polymorphisms were not associated with hypertension or body mass index. CONCLUSIONS: These two decorin polymorphisms appear to have biological relevance in human vascular pathophysiology.

Genetic Polymorphisms of Transcription Factor NRF2 and of its Host Gene Sulfiredoxin (SRXN1) are Associated with Cerebrovascular Disease in a Finnish Cohort, the TAMRISK Study.

Oxidative stress is involved in the pathophysiology of many cardiovascular disorders, such as hypertension and atherosclerosis. NRF2 is the primary transcriptional regulator of several antioxidant genes, including that of sulfiredoxin (SRXN1). The association of genotypes of NRF2 and SRXN1 with cardiovascular conditions was studied in a Finnish cohort of 336 subjects with diagnosed hypertension and 480 normotensive controls from the Tampere adult population cardiovascular risk study (TAMRISK). Samples were genotyped for four SNPs (rs1962142, rs2706110, rs6721961 and rs6706649) in the NRF2 gene region and four SNPs (rs6053666, rs6116929, rs2008022, rs6085283) in the SRXN1 gene region using Competitive Allele Specific PCR (KASP) technique. Cardiovascular diseases were followed up from 2005 to 2014 using the Finnish National Hospital Discharge Registry (HILMO). Four out of eight studied polymorphisms: rs6721961, rs1962142, rs2706110 of NRF2, and rs6053666 of SRXN1 were associated with cerebrovascular disease. NRF2 polymorphism rs6721961 showed association with hypertension. NRF2 and SRXN1 polymorphisms, previously thought to be associated with human disease, appear to be associated particularly with cerebrovascular disease.

Functional Inducible Nitric Oxide Synthase Gene Variants Associate With Hypertension: A Case-Control Study in a Finnish Population-The TAMRISK Study.

Increased inducible nitric oxide synthase (iNOS) activity and expression has been associated with hypertension, but less is known whether the 2 known functional polymorphic sites in the iNOS gene (g.-1026 C/A (rs2779249), g.2087 G/A (rs2297518)) affect susceptibility to hypertension. The objective of this study was to investigate the association between the genetic variants of iNOS and diagnosed hypertension in a Finnish cohort.This study included 320 hypertensive cases and 439 healthy controls. All participants were 50-year-old men and women and the data were collected from the Tampere adult population cardiovascular risk study (TAMRISK). DNA was extracted from buccal swabs and iNOS single nucleotide polymorphisms (SNPs) were analyzed using KASP genotyping PCR. Data analysis was done by logistic regression.At the age of 50 years, the SNP rs2779249 (C/A) associated significantly with hypertension (P = 0.009); specifically, subjects carrying the A-allele had higher risk of hypertension compared to those carrying the CC genotype (OR = 1.47; CI = 1.08-2.01; P = 0.015). In addition, a 15-year follow-up period (35, 40, and 45 years) of the same individuals showed that carriers of the A-allele had more often hypertension in all of the studied age-groups. The highest risk for developing hypertension was obtained among 35-year-old subjects (odds ratio [OR] 3.83; confidence interval [CI] = 1.20-12.27; P = 0.024). Those carrying variant A had also significantly higher readings of both systolic (P = 0.047) and diastolic (P = 0.048) blood pressure during the follow-up. No significant associations between rs2297518 (G/A) variants alone and hypertension were found. However, haplotype analysis of rs2779249 and rs2297518 revealed that individuals having haplotype H3 which combines both A alleles (CA-GA, 19.7% of individuals) was more commonly found in the hypertensive group than in the normotensive group (OR = 2.01; CI = 1.29-3.12; P = 0.002).In conclusion, there was a significant association between iNOS genetic variant (rs2779249) and hypertension in the genetically homogenous Finnish population. Those who carried the rare A-allele of the gene had higher risk for hypertension already at the age of 35 years.

NLR family pyrin domain containing 3 (NLRP3) inflammasome gene polymorphism rs7512998 (C>T) predicts aging-related increase of blood pressure, the TAMRISK study.

BACKGROUND: The activation of NLR family pyrin domain containing 3 (NLRP3) inflammasome by cellular stress leads to activation of the inflammasome, and NLRP3 gene polymorphisms have been associated with autoinflammatory diseases. Inflammasomes have also been implicated in the initiation or progression of metabolic disorders such as atherosclerosis, type 2 diabetes and obesity. The association of NLRP3 genetic variant rs7512998 with blood pressure and hypertension was studied in a 50-year-old Finnish cohort with a subpopulation who had available data on blood pressure measurements also at the age of 45 years. RESULTS: NLRP3 gene polymorphism rs7512998 C-allele was associated with higher systolic (p = 0.006) and diastolic (p = 0.011) blood pressure compared to the TT-genotype carriers in 50-year-old subjects. In addition, by analysis of variance for repeated measures between ages of 45- and 50 years there was a significant time by genotype interaction; blood pressure increased more in subjects with the C-allele both in systolic (p = 0.035) and diastolic (p = 0.012) values. However, no association with diagnosed hypertension was found. CONCLUSION: We report for the first time that NLRP3 gene polymorphism rs7512998 was associated with systolic and diastolic blood pressure in 50-year-old subjects. In addition, an effect of this variation upon blood pressure was seen in these same subjects in a 5-year follow-up from a 45-year-old cohort to 50 years of age.

Genetic variant coding for iron regulatory protein HFE contributes to hypertension, the TAMRISK study.

Iron is essential for body homeostasis, but iron overload may lead to metabolic abnormalities and thus increase the risk for atherosclerosis and many other diseases. Major histocompatibility complex class I-like transmembrane protein (HFE) is involved in body iron metabolism. The gene coding for HFE has 3 well-known polymorphic sites of which H63D (rs1799945, C > G) has recently been associated with hypertension in a genome-wide association study (GWAS) study. In the present study, we wanted to clarify whether the genetic variant associates with hypertension in a Finnish cohort consisting of 50-year-old men and women. The study included 399 hypertensive cases and 751 controls from the Tampere adult population cardiovascular risk study (TAMRISK) cohort. Genotyping of polymorphisms was done by polymerase chain reaction using DNAs extracted from buccal swabs. We found that individuals with the mutated form of the H63D polymorphic site (G-allele) had a 1.4-fold risk (P = 0.037, 95% confidence interval [CI] 1.02-1.89) for hypertension at the age of 50 years compared with the CC genotype carriers. When obese subjects (body mass index > 30 kg/m²) were analyzed in their own group, the risk for hypertension was even stronger (odds ratio 4.15, P < 0.001, 95% CI 1.98-8.68). We also noticed that the blood pressure (BP) readings were higher in those with the minor G-allele when compared to ones having a normal genotype already at the age of 35 years. Means of systolic/diastolic BPs were 127/81 mm Hg for CC and 131/83 mm Hg for CG + GG groups (P < 0.001 for systolic and P = 0.005 for diastolic pressure). In conclusion, HFE genetic variant H63D was associated with essential hypertension in Finnish subjects from the TAMRISK cohort confirming a previous GWAS study. The effect of this SNP on BP was also confirmed from a longitudinal study.

Contribution of SLC7A1 genetic variant to hypertension, the TAMRISK study.

BACKGROUND: The rs41318021 polymorphism in the SLC7A1 gene affects endothelial NO production through changes in L-arginine transport. This variation could thus hypothetically cause dysfunction of endothelium and lead to hypertension. The association of rs41318021 with hypertension was therefore studied in a Finnish cohort. METHODS: A total of 412 hypertensive cases and 771 non-hypertensive controls from a Finnish 50-year-old cohort were included in this study. The data was collected from the Tampere adult population cardiovascular risk study (TAMRISK). DNA was extracted from buccal swabs and amplified using PCR. A subpopulation of men and women who had available follow-up data of blood pressure measurements at the age of 35-, 40-, 45- and 50 years was also analyzed. RESULTS: There was no difference between the variant frequencies of the hypertension group and normotensive group at the age of 50 years (p = 0.209). However, repeated measures analysis from the 15-year follow-up showed that subjects having gene variants CT or TT had slightly higher diastolic blood pressure than subjects having genotype CC (p = 0.047). By post-hoc analysis, this was most pronounced at the age of 35 years (p = 0.044). CONCLUSION: The rs41318021 polymorphism in the SLC7A1 gene was not associated with essential hypertension in 50-year-old subjects. However, a borderline effect of this variation upon diastolic blood pressure was seen in these same subjects in a 15-year follow-up from a 35-year-old cohort to 50 years of age.

A functional variant in the serine-threonine kinase coding gene is associated with hypertension: a case-control study in a Finnish population, the Tampere adult population cardiovascular risk study.

OBJECTIVES: Hypertension raises the risk of cardiovascular consequences to two-fold or three-fold. The incidence of hypertension is increasing worldwide. Genetic causes of blood pressure are estimated to cause half of the hypertension effect, but the genes behind this are still fairly unclear. Polymorphisms in gene STK39 (serine/threonine kinase) have in some studies been associated with hypertension, but results have differed according to genetic population. We screened the STK39 polymorphism rs6749447 in a Finnish cohort to see if it was associated with hypertension. METHODS: The study included 447 hypertensive cases and 771 controls. All participants were 50-year-old Finnish patients and the data was collected from the Tampere adult population cardiovascular risk study (TAMRISK). Genotypes were determined by polymerase chain reaction using DNAs extracted from buccal swabs. RESULTS: The risk for hypertension among G-allele carriers was 1.4-fold compared with controls (P = 0.006, 95% CI = 1.10-1.79). The genetic effect of the G-allele was even more significant when the strong effect of BMI on hypertension was taken into account: for normal weight patients (BMI < 25) the risk was two-fold (P = 0.003, 95% CI 1.3-3.1) and for normal weight or slightly overweight patients (BMI < 30), the risk was 1.6-fold (P = 0.001, 95% CI 1.2-2.2). CONCLUSION: In conclusion, there was a significant association between STK39 genetic variant rs6749447 and hypertension in a Finnish cohort.

Periodic cohort health examinations in the TAMRISK study show untoward increases in body mass index and blood pressure during 15 years of follow-up.

BACKGROUND: Obesity is a significant risk factor for hypertension and diabetes. A cohort of 50-year-old voluntary periodic health examination (PHE) participants was analyzed 15 years retrospectively. Our aim was to evaluate changes in body mass index (BMI) and blood pressure in subjects diagnosed with hypertension and/or diabetes in comparison with healthy controls. METHODS: Voluntary periodic health examinations (PHE) of the citizens have been carried out by the city of Tampere, Finland. Health data, including body mass index (BMI) and blood pressure, were recorded every five years, starting at the age of 35 (baseline). A total of 339 subjects from the 50-year-old cohort having hypertension and/or diabetes were chosen to the study group. The control group included 604 subjects from the 50-year-old cohort who had the same follow-up information but were not diagnosed with hypertension and/or diabetes. RESULTS: In the study group the mean BMI had increased from 26.1 at baseline to 28.5 at the final 15-year follow-up examination. The corresponding increase in the control group was from 23.8 at baseline to 25.5 at the final follow-up. The difference in change with time between the groups was statistically significant (p = 0.04). On the average, the controls gained 4.9 kilograms, whereas subjects in the study group gained 7.0 kilograms over the 15 years of follow-up. Systolic and diastolic blood pressures were also higher in the study group already at baseline and systolic blood pressure increased with time more in the study group than in the control group (p = 0.004). CONCLUSIONS: BMI and blood pressure were higher in the study group in comparison with the controls already at baseline at 35 years, and the differences were not favorably changed during the follow-up. Apparently, the effect of PHE had not been as efficient as planned on subjects in the study group, who were already slightly overweight at baseline.

Nrf2 gene transfer induces antioxidant enzymes and suppresses smooth muscle cell growth in vitro and reduces oxidative stress in rabbit aorta in vivo.

BACKGROUND: Reactive oxygen species (ROS) play a major role in vascular inflammation and pathophysiology of many vascular diseases such as atherosclerosis and injury-induced neointima formation after balloon angioplasty. Nuclear factor E2-related factor-2 (Nrf2) is a transcription factor orchestrating antioxidant and cytoprotective responses on oxidative and electrophilic stress, and it has been shown to have antiinflammatory effects in vascular cells in vitro. We therefore postulated that Nrf2 gene transfer would have salutary effects on vascular inflammation after angioplasty. METHODS AND RESULTS: Transduction of vascular smooth muscle cells (VSMCs) with Nrf2-expressing adenovirus increased the expression of several antioxidant enzymes including heme oxygenase-1 (HO-1) compared with beta-galactosidase (AdLacZ)-transduced controls. Moreover, Nrf2 gene transfer also inhibited vascular smooth muscle cell (VSMC) proliferation, and the effect was partially reversed by the HO inhibitor Sn(IV) protoporphyrin. In vivo, adenoviral gene transfer effectively reduced oxidative stress determined by antibody staining against oxidized epitopes of LDL, as well as inhibited vascular inflammation assessed by the macrophage cell count and monocyte chemoattractant protein-1 (MCP-1) staining. However, the antiproliferative effects of Nrf2 in vivo were counterbalanced with diminished apoptosis in neointimal VSMCs, resulting in no change in neointimal hyperplasia. CONCLUSIONS: Nrf2 gene transfer or Nrf2-inducing drugs may have therapeutic applications in vascular diseases in which inflammation and oxidative stress play a role. However, the contrasting growth inhibitory and antiapoptotic effects of Nrf2 need to be considered in pathological conditions in which SMC proliferation plays a critical role.