PRPH2-Related Retinal Dystrophies: Mutational Spectrum in 103 Families from a Spanish Cohort.

PRPH2, one of the most frequently inherited retinal dystrophy (IRD)-causing genes, implies a high phenotypic variability. This study aims to analyze the PRPH2 mutational spectrum in one of the largest cohorts worldwide, and to describe novel pathogenic variants and genotype-phenotype correlations. A study of 220 patients from 103 families recruited from a database of 5000 families. A molecular diagnosis was performed using classical molecular approaches and next-generation sequencing. Common haplotypes were ascertained by analyzing single-nucleotide polymorphisms. We identified 56 variants, including 11 novel variants. Most of them were missense variants (64%) and were located in the D2-loop protein domain (77%). The most frequently occurring variants were p.Gly167Ser, p.Gly208Asp and p.Pro221_Cys222del. Haplotype analysis revealed a shared region in families carrying p.Leu41Pro or p.Pro221_Cys222del. Patients with retinitis pigmentosa presented an earlier disease onset. We describe the largest cohort of IRD families associated with PRPH2 from a single center. Most variants were located in the D2-loop domain, highlighting its importance in interacting with other proteins. Our work suggests a likely founder effect for the variants p.Leu41Pro and p.Pro221_Cys222del in our Spanish cohort. Phenotypes with a primary rod alteration presented more severe affectation. Finally, the high phenotypic variability in PRPH2 hinders the possibility of drawing genotype-phenotype correlations.

New COL6A6 variant detected by whole-exome sequencing is linked to break points in intron 4 and 3'-UTR, deleting exon 5 of RHO, and causing adRP.

PURPOSE: This study aimed to test a newly devised cost-effective multiplex PCR assay for the molecular diagnosis of autosomal dominant retinitis pigmentosa (adRP), as well as the use of whole-exome sequencing (WES) to detect disease-causing mutations in adRP. METHODS: Genomic DNA was extracted from peripheral blood lymphocytes of index patients with adRP and their affected and unaffected family members. We used a newly devised multiplex PCR assay capable of amplifying the genetic loci of RHO, PRPH2, RP1, PRPF3, PRPF8, PRPF31, IMPDH1, NRL, CRX, KLHL7, and NR2E3 to molecularly diagnose 18 index patients with adRP. We also performed WES in affected and unaffected members of four families with adRP in whom a disease-causing mutation was previously not found. RESULTS: We identified five previously reported mutations (p.Arg677X in the RP1 gene, p.Asp133Val and p.Arg195Leu in the PRPH2 gene, and p.Pro171Leu and p.Pro215Leu in the RHO gene) and one novel mutation (p.Val345Gly in the RHO gene) representing 33% detection of causative mutations in our adRP cohort. Comparative WES analysis showed a new variant (p.Gly103Arg in the COL6A6 gene) that segregated with the disease in one family with adRP. As this variant was linked with the RHO locus, we sequenced the complete RHO gene, which revealed a deletion in intron 4 that encompassed all of exon 5 and 28 bp of the 3'-untranslated region (UTR). CONCLUSIONS: The novel multiplex PCR assay with next-generation sequencing (NGS) proved effective for detecting most of the adRP-causing mutations. A WES approach led to identification of a deletion in RHO through detection of a new linked variant in COL6A6. No pathogenic variants were identified in the remaining three families. Moreover, NGS and WES were inefficient for detecting the complete deletion of exon 5 in the RHO gene in one family with adRP. Carriers of this deletion showed variable clinical status, and two of these carriers had not previously been diagnosed with RP.

Mutational screening of splicing factor genes in cases with autosomal dominant retinitis pigmentosa.

PURPOSE: Mutations in genes encoding proteins from the tri-snRNP complex of the spliceosome account for more than 12% of cases of autosomal dominant retinitis pigmentosa (adRP). Although the exact mechanism by which splicing factor defects trigger photoreceptor death is not completely clear, their role in retinitis pigmentosa has been demonstrated by several genetic and functional studies. To test for possible novel associations between splicing factors and adRP, we screened four tri-snRNP splicing factor genes (EFTUD2, PRPF4, NHP2L1, and AAR2) as candidate disease genes. METHODS: We screened up to 303 patients with adRP from Europe and North America who did not carry known RP mutations. Exon-PCR and Sanger methods were used to sequence the NHP2L1 and AAR2 genes, while the sequences of EFTUD2 and PRPF4 were obtained by using long-range PCRs spanning coding and non-coding regions followed by next-generation sequencing. RESULTS: We detected novel missense changes in individual patients in the sequence of the genes PRPF4 and EFTUD2, but the role of these changes in relationship to disease could not be verified. In one other patient we identified a novel nucleotide substitution in the 5' untranslated region (UTR) of NHP2L1, which did not segregate with the disease in the family. CONCLUSIONS: The absence of clearly pathogenic mutations in the candidate genes screened in our cohort suggests that EFTUD2, PRPF4, NHP2L1, and AAR2 are either not involved in adRP or are associated with the disease in rare instances, at least as observed in this study in patients of European and North American origin.

Outcome of ABCA4 disease-associated alleles in autosomal recessive retinal dystrophies: retrospective analysis in 420 Spanish families.

OBJECTIVE: To provide a comprehensive overview of all detected mutations in the ABCA4 gene in Spanish families with autosomal recessive retinal disorders, including Stargardt's disease (arSTGD), cone-rod dystrophy (arCRD), and retinitis pigmentosa (arRP), and to assess genotype-phenotype correlation and disease progression in 10 years by considering the type of variants and age at onset. DESIGN: Case series. PARTICIPANTS: A total of 420 unrelated Spanish families: 259 arSTGD, 86 arCRD, and 75 arRP. METHODS: Spanish families were analyzed through a combination of ABCR400 genotyping microarray, denaturing high-performance liquid chromatography, and high-resolution melting scanning. Direct sequencing was used as a confirmation technique for the identified variants. Screening by multiple ligation probe analysis was used to detect possible large deletions or insertions in the ABCA4 gene. Selected families were analyzed further by next generation sequencing. MAIN OUTCOME MEASURES: DNA sequence variants, mutation detection rates, haplotypes, age at onset, central or peripheral vision loss, and night blindness. RESULTS: Overall, we detected 70.5% and 36.6% of all expected ABCA4 mutations in arSTGD and arCRD patient cohorts, respectively. In the fraction of the cohort where the ABCA4 gene was sequenced completely, the detection rates reached 73.6% for arSTGD and 66.7% for arCRD. However, the frequency of possibly pathogenic ABCA4 alleles in arRP families was only slightly higher than that in the general population. Moreover, in some families, mutations in other known arRP genes segregated with the disease phenotype. CONCLUSIONS: An increasing understanding of causal ABCA4 alleles in arSTGD and arCRD facilitates disease diagnosis and prognosis and also is paramount in selecting patients for emerging clinical trials of therapeutic interventions. Because ABCA4-associated diseases are evolving retinal dystrophies, assessment of age at onset, accurate clinical diagnosis, and genetic testing are crucial. We suggest that ABCA4 mutations may be associated with a retinitis pigmentosa-like phenotype often as a consequence of severe (null) mutations, in cases of long-term, advanced disease, or both. Patients with classical arRP phenotypes, especially from the onset of the disease, should be screened first for mutations in known arRP genes and not ABCA4.

Obesity in Patients with Type 1 Diabetes: Links, Risks and Management Challenges.

Obesity affects large numbers of patients with type 1 diabetes (T1D) across their lifetime, with rates ranging between 2.8% and 37.1%. Patients with T1D and obesity are characterized by the presence of insulin resistance, of high insulin requirements, have a greater cardiometabolic risk and an enhanced risk of developing chronic complications when compared to normal-weight persons with T1D. Dual treatment of obesity and T1D is challenging and no specific guidelines for improving outcomes of both glycemic control and weight management have been established for this population. Nevertheless, although evidence is scarce, a comprehensive approach based on a balanced hypocaloric diet, physical activity and cognitive behavioral therapy by a multidisciplinary team, expert in both obesity and diabetes, remains as the best clinical practice. However, weight loss responses with lifestyle changes alone are limited, so in the "roadmap" of the treatment of obesity in T1D, it will be helpful to include anti-obesity pharmacotherapy despite at present there is a lack of evidence since T1D patients have been excluded from anti-obesity drug clinical trials. In case of severe obesity, bariatric surgery has proven to be of benefit in obtaining a substantial and long-term weight loss and reduction in cardiovascular risk. The near future looks promising with the development of new and more effective anti-obesity treatments and strategies to improve insulin resistance and oxidative stress. Advances in precision medicine may help individualize and optimize the medical management and care of these patients. This review, by gathering current evidence, highlights the need of solid knowledge in all facets of the treatment of patients with obesity and T1D that can only be obtained through high quality well-designed studies.

Pancreatic ductal cells may have a negative effect on human islet transplantation.

AIM: To evaluate the effect of pancreatic ductal cells on experimental human islet transplantation. MATERIALS AND METHODS: Isolated islets were additionally purified by handpicking. Ductal cells were purified by magnetic cell sorting and then clustered into ductal pancreatospheres (DPS). Islets, DPS, and islets + DPS (100 islets + 75 DPS, or 100 islets + 200 DPS) were cultured and glucose-stimulated insulin secretion, ß-cell apoptosis, and gene expression was determined. Islets and islets + DPS preparations (800 islets + 600 DPS) were transplanted to streptozotocin-treated immunodeficient mice and glycemia, graft morphometry, and gene expression were determined. RESULTS: Insulin stimulation index was higher in islets than in islets co-cultured with DPS (5.59 ± 0.93 vs 4.02 ± 0.46; p<0.05). IL1B and CXCL11 expression was higher in 100 islets + 200 DPS than in islets (p<0.01), and IL-1ß was detected in supernatants collected from DPS and islets + DPS preparations, but not in islets. Hyperglycemia developed in 33% and 67% of mice transplanted with islets or with islets + DPS respectively. ß-cell mass was 26% lower in islets + DPS than in islets grafts (p>0.05), and the ratio ß-/endocrine non-ß-cell mass was lower in islets + DPS grafts (islets: 2.05 ± 0.18, islets + DPS: 1.35 ± 0.15; p<0.01). IL1B and IL1RN expression was significantly higher in islets + DPS grafts. CONCLUSIONS: Islet preparations enriched with ductal cells have a lower insulin stimulation index in vitro and achieved a worse metabolic outcome after transplantation. Inflammation may mediate the deleterious effects of ductal cells on islet cells.

Analysis of the PRPF31 Gene in Spanish Autosomal Dominant Retinitis Pigmentosa Patients: A Novel Genomic Rearrangement.

Purpose: The aim was to determine the prevalence of PRPF31 mutations in a cohort of Spanish autosomal dominant retinitis pigmentosa (adRP) families to deepen knowledge of the pathogenic mechanisms underlying the disease and to assess genotype-phenotype correlations. Methods: A cohort of 211 adRP patients was screened for variants in PRPF31 by using a combined strategy comprising next-generation sequencing approaches and copy-number variation (CNV) analysis. Quantitative RT-PCR and CNV analysis of the regulatory MSR1 element were also performed to assess PRPF31 gene expression. Phenotype was assessed by using ophthalmologic examination protocols. Results: Fifteen different causative mutations and genomic rearrangements were identified, revealing five novel mutations. Prevalence of PRPF31 mutations, genomic rearrangements, and lack of penetrance were 7.6%, 1.9%, and 66.7%, respectively. Interestingly, we identified a tandem duplication and a partial PRPF31 deletion in different affected individuals from the same family. PRPF31 gene expression was significantly decreased in symptomatic cases carrying either PRPF31 duplication or deletion as compared to controls. The 4 MSR1 allele in cis with the PRPF31 wild-type allele was apparently a protective factor. The mutated phenotype varied from no symptoms to typical retinitis pigmentosa with variable onset and course depending on the kind of mutation, with the duplication case the most severe. Conclusions: In view of the high genetic heterogeneity of PRPF31 mutations, the screening must include the entire gene, as well as CNV assays, to detect large rearrangements. This is the first report of a variable phenotype correlation as well as a gross duplication and deletion within the same family.

Impact and characteristics of the non-Caucasian population in hospital admissions for diabetes onset during 2003-2010. / Impacto y características diferenciales de la población de origen no caucásico en los ingresos por inicio de diabetes durante el periodo 2003-2010.

AIMS: To assess the prevalence of non-Caucasian patients in hospital admissions for onset of symptomatic diabetes mellitus during the 2003-2010 period, and to analyze the characteristics differentiating them from the Caucasian population at diagnosis and 2 years later. MATERIAL AND METHODS: A retrospective, observational study. INCLUSION CRITERIA: Patients aged 18-40 years admitted for de novo symptomatic diabetes from January 2003 to October 2010. Prevalence of patients of non-Caucasian origin was analyzed, and clinical, biochemical, immunological, and beta-cell function of both populations were compared at diagnosis and 2 years later. RESULTS: Nineteen percent of patients admitted to hospital for de novo symptomatic diabetes were non-Caucasian, with a progressive increase in recent years. Non-Caucasian patients had milder decompensation (3.0% had ketoacidosis, as compared to 15.2% in the Caucasian group, P<.05), lower presence of autoimmunity (27.2 vs. 73.1%, P<.01) and higher stimulated C-peptide levels (0.70±0.56 vs. 0.42±0.39 nmol/l, P<.05), mainly because of the subgroup with negative autoimmunity (0.82 vs. 0.25). Two years after diagnosis, less non-Caucasian patients were on intensified treatment (39.1 vs. 93.8%). CONCLUSIONS: Non-Caucasian patients had a lower prevalence of autoimmunity, better beta-cell function at diagnosis, particularly due to the subgroup with negative autoimmunity, and less need for intensive treatment 2 years after diagnosis, features which are more characteristic of type 2 diabetes mellitus.

Dominant Retinitis Pigmentosa, p.Gly56Arg Mutation in NR2E3: Phenotype in a Large Cohort of 24 Cases.

IMPORTANCE: This research is the single largest NR2E3 genotype-phenotype correlation study performed to date in autosomal dominant Retinitis Pigmentosa. OBJECTIVE: The aim of this study is to analyse the frequency of the p.Gly56Arg mutation in NR2E3 for the largest cohort of autosomal dominant Retinitis Pigmentosa patients to date and its associated phenotype. PATIENTS AND METHODS: A cohort of 201 unrelated Spanish families affected by autosomal dominant Retinitis Pigmentosa. The p.Gly56Arg mutation in the NR2E3 (NM_014249.2) gene was analysed in 201 families. In the 24 cases where the mutation had been detected, a haplotype analysis linked to the p.Gly56Arg families was performed, using four extragenic polymorphic markers D15S967, D15S1050, D15S204 and D15S188. Phenotype study included presence and age of onset of night blindness, visual field loss and cataracts; and an ophthalmoscopic examination after pupillary dilation and electroretinogram for the 24 cases. RESULTS: Seven of the 201 analyzed families were positive for the p.Gly56Arg, leading to a prevalence of 3.5%. Clinical data were available for 24 subjects. Night blindness was the first noticeable symptom (mean 15.9 years). Visual field loss onset was variable (23.3 ± 11.9 years). Loss of visual acuity appeared late in the disease´s evolution. Most of the patients with cataracts (50%) presented it from the third decade of life. Fundus changes showed inter and intrafamiliar variability, but most of the patients showed typical RP changes and it was common to find macular affectation (47.4%). Electroretinogram was impaired from the beginning of the disease. Two families shared a common haplotype. Additionally, all patients shared a 104Kb region between D15S1050 and the NR2E3 gene. CONCLUSIONS: This study highlights the importance of p.Gly56Arg in the NR2E3 gene as a common mutation associated with adRP, and provides new clues to its phenotype, which can allow for a better clinical management and genetic counselling of patients and their families.

Panel-based NGS Reveals Novel Pathogenic Mutations in Autosomal Recessive Retinitis Pigmentosa.

Retinitis pigmentosa (RP) is a group of inherited progressive retinal dystrophies (RD) characterized by photoreceptor degeneration. RP is highly heterogeneous both clinically and genetically, which complicates the identification of causative genes and mutations. Targeted next-generation sequencing (NGS) has been demonstrated to be an effective strategy for the detection of mutations in RP. In our study, an in-house gene panel comprising 75 known RP genes was used to analyze a cohort of 47 unrelated Spanish families pre-classified as autosomal recessive or isolated RP. Disease-causing mutations were found in 27 out of 47 cases achieving a mutation detection rate of 57.4%. In total, 33 pathogenic mutations were identified, 20 of which were novel mutations (60.6%). Furthermore, not only single nucleotide variations but also copy-number variations, including three large deletions in the USH2A and EYS genes, were identified. Finally seven out of 27 families, displaying mutations in the ABCA4, RP1, RP2 and USH2A genes, could be genetically or clinically reclassified. These results demonstrate the potential of our panel-based NGS strategy in RP diagnosis.

Prevalence of Rhodopsin mutations in autosomal dominant Retinitis Pigmentosa in Spain: clinical and analytical review in 200 families.

PURPOSE: We aimed to determine the prevalence of mutations in the RHO gene in Spanish families with autosomal dominant Retinitis Pigmentosa (adRP), to assess genotype-phenotype correlations and to establish an accurate diagnostic algorithm after 23 years of data collection. PATIENTS AND METHODS: Two hundred patients were analysed through a combination of denaturing gradient gel electrophoresis, single-strand conformation polymorphism, genotyping microarray and Sanger sequencing of the RHO gene. RESULTS: Overall, 42 of 200 Spanish adRP families were mutated for RHO (21.0%). Twenty-seven different RHO mutations were detected; seven of them were novel. A genotype-phenotype correlation was established with clinical data from 107 patients. The most prevalent p.Pro347Leu mutation, responsible for 4.5% (9/200) of all mutated adRP families, was associated with a phenotype of early onset and severe course diffuse RP. CONCLUSIONS: This retrospective study provides a wide spectrum of mutations in the RHO gene in Spanish patients with adRP. Also, the prevalence of mutations is similar to that reported in European population. Genotyping microarray followed by RHO sequencing is proposed as a first step in molecular diagnosis of adRP Spanish families. An increasing understanding of causal RHO alleles in adRP facilitates disease diagnosis and prognosis, especially for the prevalent p.Pro347Leu mutation.

Application of Whole Exome Sequencing in Six Families with an Initial Diagnosis of Autosomal Dominant Retinitis Pigmentosa: Lessons Learned.

This study aimed to identify the genetics underlying dominant forms of inherited retinal dystrophies using whole exome sequencing (WES) in six families extensively screened for known mutations or genes. Thirty-eight individuals were subjected to WES. Causative variants were searched among single nucleotide variants (SNVs) and insertion/deletion variants (indels) and whenever no potential candidate emerged, copy number variant (CNV) analysis was performed. Variants or regions harboring a candidate variant were prioritized and segregation of the variant with the disease was further assessed using Sanger sequencing in case of SNVs and indels, and quantitative PCR (qPCR) for CNVs. SNV and indel analysis led to the identification of a previously reported mutation in PRPH2. Two additional mutations linked to different forms of retinal dystrophies were identified in two families: a known frameshift deletion in RPGR, a gene responsible for X-linked retinitis pigmentosa and p.Ser163Arg in C1QTNF5 associated with Late-Onset Retinal Degeneration. A novel heterozygous deletion spanning the entire region of PRPF31 was also identified in the affected members of a fourth family, which was confirmed with qPCR. This study allowed the identification of the genetic cause of the retinal dystrophy and the establishment of a correct diagnosis in four families, including a large heterozygous deletion in PRPF31, typically considered one of the pitfalls of this method. Since all findings in this study are restricted to known genes, we propose that targeted sequencing using gene-panel is an optimal first approach for the genetic screening and that once known genetic causes are ruled out, WES might be used to uncover new genes involved in inherited retinal dystrophies.

Targeted Next-Generation Sequencing Improves the Diagnosis of Autosomal Dominant Retinitis Pigmentosa in Spanish Patients.

PURPOSE: Next-generation sequencing (NGS) has been demonstrated to be an effective strategy for the detection of mutations in retinal dystrophies, a group of inherited diseases that are highly heterogeneous. Therefore, the aim of this study is the application of an NGS-based approach in a Spanish cohort of autosomal dominant retinitis pigmentosa (RP) patients to find out causative mutations. METHODS: Index cases of 59 Spanish families with initial diagnosis of autosomal dominant RP and unsuccessfully studied for mutations in the most common RP causal genes, were selected for application of a NGS-based approach with a custom panel for 73 genes related to retinal dystrophies. Candidate variants were select based on frequency, pathogenicity, inherited model, and phenotype. Subsequently, confirmation by Sanger sequencing, cosegregation analysis, and population studies, was applied for determining the implication of those variants in the pathology. RESULTS: Overall 31 candidate variants were selected. From them, 17 variants were considered as mutations causative of the disease, 64% (11/17) of them were novel and 36% (6/17) were known RP-related mutations. Therefore, applying this technology16 families were characterized rendering a mutation detection rate of 27% (16/59). Of them, 5% (3/59) of cases displayed mutations in recessive or X-linked genes (ABCA4, RPGR, and RP2) allowing a genetic and clinical reclassification of those families. Furthermore, seven novel variants with uncertain significance and seven novel variants probably not causative of disease were also found. CONCLUSIONS: This NGS strategy is a fast, effective, and reliable tool to detect known and novel mutations in autosomal dominant RP patients allowing genetic reclassification in some cases and increasing the knowledge of pathogenesis in retinal dystrophies.

Contribution of mutation load to the intrafamilial genetic heterogeneity in a large cohort of Spanish retinal dystrophies families.

PURPOSE: The aim of this study was to deepen our knowledge on the basis of intrafamilial genetic heterogeneity of inherited retinal dystrophies (RD) to further discern the contribution of individual alleles to the pathology. METHODS: Families with intrafamilial locus and/or allelic heterogeneity were selected from a cohort of 873 characterized of 2468 unrelated RD families. Clinical examination included visual field assessments, electrophysiology, fundus examination, and audiogram. Molecular characterization was performed using a combination of different methods: genotyping microarray, single strand conformational polymorphism (SSCP), denaturing high pressure liquid chromatography (dHPLC), high resolution melt (HRM), multiplex ligation-dependent probe amplification (MLPA), Sanger sequencing, whole-genome homozygosity mapping, and next-generation sequencing (NGS). RESULTS: Overall, intrafamilial genetic heterogeneity was encountered in a total of 8 pedigrees. There were 5 of 873 families (~0.6%) with causative mutations in more than one gene (locus heterogeneity), involving the genes: (1) USH2A, RDH12, and TULP1; (2) PDE6B and a new candidate gene; (3) CERKL and CRB1; (4) BBS1 and C2orf71; and (5) ABCA4 and CRB1. Typically, in these cases, each mutated gene was associated with different phenotypes. In the 3 other families (~0.35%), different mutations in the same gene (allelic heterogeneity) were found, including the frequent RD genes ABCA4 and CRB1. CONCLUSIONS: This systematic research estimates that the frequency of overall mutation load promoting RD intrafamilial heterogeneity in our cohort of Spanish families is almost 1%. The identification of the genetic mechanisms underlying RD locus and allelic heterogeneity is essential to discriminate the real contribution of the monoallelic mutations to the disease, especially in the NGS era. Moreover, it is decisive to provide an accurate genetic counseling and in disease treatment.

Expanding the phenotype of PRPS1 syndromes in females: neuropathy, hearing loss and retinopathy.

BACKGROUND: Phosphoribosyl pyrophosphate synthetase (PRS) I deficiency is a rare medical condition caused by missense mutations in PRPS1 that lead to three different phenotypes: Arts Syndrome (MIM 301835), X-linked Charcot-Marie-Tooth (CMTX5, MIM 311070) or X-linked non-syndromic sensorineural deafness (DFN2, MIM 304500). All three are X-linked recessively inherited and males affected display variable degree of central and peripheral neuropathy. We applied whole exome sequencing to a three-generation family with optic atrophy followed by retinitis pigmentosa (RP) in all three cases, and ataxia, progressive peripheral neuropathy and hearing loss with variable presentation. METHODS: Whole exome sequencing was performed in two affecteds and one unaffected member of the family. Sanger sequencing was used to validate and segregate the 12 candidate mutations in the family and to confirm the absence of the novel variant in PRPS1 in 191 controls. The pathogenic role of the novel mutation in PRPS1 was assessed in silico and confirmed by enzymatic determination of PRS activity, mRNA expression and sequencing, and X-chromosome inactivation. RESULTS: A novel missense mutation was identified in PRPS1 in the affected females. Age of onset, presentation and severity of the phenotype are highly variable in the family: both the proband and her mother have neurological and ophthalmological symptoms, whereas the phenotype of the affected sister is milder and currently confined to the eye. Moreover, only the proband displayed a complete lack of expression of the wild type allele in leukocytes that seems to correlate with the degree of PRS deficiency and the severity of the phenotype. Interestingly, optic atrophy and RP are the only common manifestations to all three females and the only phenotype correlating with the degree of enzyme deficiency. CONCLUSIONS: These results are in line with recent evidence of the existence of intermediate phenotypes in PRS-I deficiency syndromes and demonstrate that females can exhibit a disease phenotype as severe and complex as their male counterparts.

Novel GUCA1A mutations suggesting possible mechanisms of pathogenesis in cone, cone-rod, and macular dystrophy patients.

Here, we report two novel GUCA1A (the gene for guanylate cyclase activating protein 1) mutations identified in unrelated Spanish families affected by autosomal dominant retinal degeneration (adRD) with cone and rod involvement. All patients from a three-generation adRD pedigree underwent detailed ophthalmic evaluation. Total genome scan using single-nucleotide polymorphisms and then the linkage analysis were undertaken on the pedigree. Haplotype analysis revealed a 55.37 Mb genomic interval cosegregating with the disease phenotype on chromosome 6p21.31-q15. Mutation screening of positional candidate genes found a heterozygous transition c.250C>T in exon 4 of GUCA1A, corresponding to a novel mutation p.L84F. A second missense mutation, c.320T>C (p.I107T), was detected by screening of the gene in a Spanish patients cohort. Using bioinformatics approach, we predicted that either haploinsufficiency or dominant-negative effect accompanied by creation of a novel function for the mutant protein is a possible mechanism of the disease due to c.250C>T and c.320T>C. Although additional functional studies are required, our data in relation to the c.250C>T mutation open the possibility that transacting factors binding to de novo created recognition site resulting in formation of aberrant splicing variant is a disease model which may be more widespread than previously recognized as a mechanism causing inherited RD.

Effect of bariatric surgery on bone mineral density: comparison of gastric bypass and sleeve gastrectomy.

The aim of our study was to compare bone mineral density (BMD) a year after Roux-en-Y gastric bypass (RYGB) and sleeve gastrectomy (SG) in age- and body mass index-matched women. In 33 morbidly obese women undergoing RYGB and 33 undergoing SG, plasma determinations of calcium, parathyroid hormone (PTH), 25-hydroxyvitamin D (25(OH) D3), and insulin-like growth factor-I (IGF-I) were made prior to and at 12 months after surgery. Dual-energy X-ray absorptiometry was performed in all patients 1 year after surgery. BMD at the femoral neck and the lumbar spine 1 year after surgery was similar in women undergoing RYGB and SG (1.01 ± 0.116 vs. 1.01 ± 0.122 g/cm(2), p = 0.993; 1.05 ± 0.116 vs. 1.08 ± 0.123 g/cm(2), p = 0.384). The percentage of patients with osteopenia and osteoporosis was not different between groups. In the linear regression analysis, age (ß = -0.628, p = 0.034) and lean mass 12 months after surgery (ß = 0.424, p = 0.021) were found to be the main determinants of femoral neck BMD. Age (ß = -0.765, p = 0.025), menopause (ß = -0.898, p = 0.033), and lean mass (ß = 0.615, p = 0.023) were determinants of BMD at the lumbar spine. No influence was found between low bone mass and type of surgery, plasma PTH, 25(OH) D3, or IGF-I. The effect of RYGB and SG on BMD was comparable a year after surgery. Menopausal women were at a higher risk of having low bone mass, but the presence of osteoporosis was uncommon.

Evaluation of bone mineral density loss in morbidly obese women after gastric bypass: 3-year follow-up.

Studies that evaluate the influence of gastric bypass (RYGP) on bone mass are limited to short-term follow-up. We analysed changes in bone mineral density (BMD) three years after surgery and evaluated the main determinants of the development of bone disease. Prospective study of 59 morbidly obese white women aged 46 ± 8 years. BMD scanning using DEXA and plasma determinations of calcium, parathyroid hormone, 25-hydroxyvitamin D and insulin-like growth factor-I were made prior, at 12 months and 3 years after surgery. In the first postoperative year BMD decreased at femoral neck (FN) 10.2 % and in the lumbar spine (LS) 3.2 %, in the third year it additionally decreased 2.7 % and 3.1 %, respectively. BMD at both sites remained above the values of women of the same age. In the follow-up, 1.7 % developed osteoporosis at FN and 6.8 % at LS. Patients with bone disease were older, the percentage of women with menopause was greater in this group and had lower initial and final values of lean mass. The percentage of BMD loss at FN remained positively associated with the percentage of lean mass loss [ß 0.304, p=0.045], and menopause [ß 0.337, p=0.025]. Major osteoporotic fracture and hip fracture risk was low even in menopausal patients (3.1 % and 0.40 %, respectively). After RYGP menopausal women and those with greater lean mass loss are at higher risk of BMD loss but progression to osteoporosis is uncommon and the risk of fracture is low.

Is plasma 25(OH) D related to adipokines, inflammatory cytokines and insulin resistance in both a healthy and morbidly obese population?

To analyse in a cohort of healthy subjects and in a group of morbidly obese patients, we studied the association amongst 25(OH) D and plasma concentrations of adipocytokines, inflammatory cytokines and insulin resistance. We also aimed to determine whether vitamin D-deficient patients showed a greater inflammatory profile. In the observational study that the authors conducted, plasma concentrations of 25(OH) D, leptin, resistin, adiponectin and interleukine-18 were determined in 134 healthy men and 127 women. In the population consisting of 44 patients with morbid obesity, plasma concentrations of 25(OH) D, leptin, resistin, adiponectin, interleukine-18, soluble tumor necrosis factor receptors 1 and 2 and C-reactive protein were analysed. In the healthy population, plasma 25(OH) D showed a negative correlation with body mass index, body fat, waist, hip circumference and with leptin. However, no significant associations were found amongst 25(OH) D and plasma concentrations of resistin, adiponectin or interleukine-18. Patients with vitamin D deficiency showed higher body mass index, fat mass percentage and higher leptin concentrations compared with subjects with normal 25(OH) D concentrations. In the morbidly obese subjects, 25(OH) D did not correlate with leptin, resistin, adiponectin, interleukine-18, soluble tumor necrosis factor receptors 1 and 2 or with C-reactive protein. In patients with morbid obesity, no differences were found in adipokines and inflammatory cytokines concentrations regarding 25(OH) D status. No associations were found either between 25(OH) D and plasma glucose and insulin resistance or with lipid profile. Plasma 25(OH) D concentrations are associated with adiposity markers but not with adipocytokines implicated in inflammation. This lack of association does not support a major role of 25(OH) D in the pro-inflammatory environment observed in morbidly obese subjects. In addition, subjects with vitamin D deficiency are not characterized by a greater inflammatory state.