Wilson disease: novel mutations in the ATP7B gene and clinical correlation in Brazilian patients

Wilson disease (WD) is a rare inherited autosomal recessive disorder caused by a defect in a metal transporting P-type ATPase, resulting in copper overload in various tissues and cells. The aim was to assess both the phenotype in Brazilian WD patients and the corresponding ATP7B genotype. Sixty subjects belonging to 46 pedigrees diagnosed as WD were included in this study. Direct sequencing of all 21 exons within ATP7B and their flanking introns was performed. Demographic, clinical, laboratory and histopathological data at the time of diagnosis were obtained. We identified twenty-five mutations, twelve of them reported for the first time. The c.3402delC mutation had the highest allelic frequency (30.8%), followed by the c.2123T>C (p.L708P) (16.7%). Exons 8 and 15 were the site of 62.5% of the mutations. The common European mutation c.3207C>A (p.H1069Q) was not present at all. Phenotype varied greatly among individuals with the same ATP7B genotype. Our data confirm the heterogeneity of ATP7B genotype in Brazilian WD patients. The mutational spectrum is compatible with the Brazilian history of Mediterranean immigration; however, new mutations, and different frequencies and phenotype associated with the previously known mutations characterize this population. Exons 8 and 15 should be preferentially screened in WD cases from Brazil. Phenotype variation among subjects with the same ATP7B genotype suggests that modifying factors play an additional role in the pathogenesis of WD.     

CXCR4-transgene expression significantly improves marrow engraftment of cultured hematopoietic stem cells

Hematopoietic stem cells (HSCs) lose marrow reconstitution potential during ex vivo culture. HSC migration to stromal cell-derived factor (SDF)-1 (CXCL12) correlates with CXC chemokine receptor 4 (CXCR4) expression and marrow engraftment. We demonstrate that mobilized human CD34+ peripheral blood stem cells (CD34+ PBSCs) lose CXCR4 expression during prolonged culture. We transduced CD34+ PBSCs with retrovirus vector encoding human CXCR4 and achieved 18-fold more CXCR4 expression in over 87% of CD34+ cells. CXCR4-transduced cells yielded increased calcium flux and up to a 10-fold increase in migration to SDF-1. Six-day cultured CXCR4-transduced cells demonstrated significant engraftment in nonobese diabetic/severe combined immunodeficient mice under conditions in which control transduced cells resulted in low or no engraftment. We conclude that transduction-mediated overexpression of CXCR4 significantly improves marrow engraftment of cultured PBSCs.     

Recently amplified satellite DNA inCallithrix argentata (Primates,Platyrrhini)

A satellite DNA has been cloned from the neotropical primateCallithrix argentata and designated CarB. The presence of the satellite was assayed in New and Old World primates by blot hybridization: CarB is highly amplified in the genomes of all three species belonging to theC. argentata species group (C. argentata, C. emiliae, C. humeralifer), but is either absent, or present in only minor amounts, in other primates, including the closely related species,C. jacchus. A completely sequenced CarB monomeric unit was 1528 bp in length and mapped to the telomeric C-band-positive regions of manyC. argentata species group chromosomes. Sequence data from eight CarB clones indicated an average difference of 3.5% when base substitutions alone were counted. The hybridization and sequence data suggest that this satellite underwent a period of amplification and dispersal in the genome of a recent ancestor of theC. argentata species group.     

Trans-Epithelial Transport,Metabolism, and Biological Activity Assessment of the Multi-Target Lupin Peptide LILPKHSDAD (P5) and Its Metabolite LPKHSDAD (P5-Met)

P5 (LILPKHSDAD) is a hypocholesterolemic peptide from lupin protein with a multi-target activity, since it inhibits both 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCoAR) and proprotein convertase subtilisin/kexin type-9 (PCSK9). This work shows that, during epithelial transport experiments, the metabolic transformation mediated by intestinal peptidases produces two main detected peptides, ILPKHSDAD (P5-frag) and LPKHSDAD (P5-met), and that both P5 and P5-met are linearly absorbed by differentiated human intestinal Caco-2 cells. Extensive comparative structural, biochemical, and cellular characterizations of P5-met and the parent peptide P5 demonstrate that both peptides have unique characteristics and share the same mechanisms of action. In fact, they exert an intrinsically multi-target behavior being able to regulate cholesterol metabolism by modulating different pathways. The results of this study also highlight the dynamic nature of bioactive peptides that may be modulated by the biological systems they get in contact with.     

Positive SARS-CoV-2 RT-qPCR of a nasal swab spot after 30 days of conservation on filter paper at room temperature

We tested the use of nasal swabs spotted onto filter paper (Whatman 3M) for the molecular diagnosis of SARS-CoV-2 infection. Spots of a positive nasal swab in conservation medium (B.1.177 strain, 21Ct) were still positive (duo E-gene/IP4) after 10, 20, and 30 days of conservation at room temperature, with Ct values of 28, 27, and 26, respectively. Direct spotting of the swab at bedside (omicron strain) still gave a positive result after 10 days in two RT-qPCR systems: 33.7 Ct using duo E-gene/IP4, and 34.8 using a specific Omicron system. Spotting of a dilution range of media spiked with the Delta (strain 2021/FR/0610, lineage B 1.617.2) and Omicron strains (strain UVE/SARS-CoV-2/2021/FR/1514) showed a threshold of 0.04 TCID₅₀ after 10 days of conservation. We show, for the first time, that this simple and low-cost conservation method can be used to store samples for RT-qPCR against SARS-CoV-2 for up to at least 1 month.     

NEOPLASTIC DISEASE AND DELETION 22q11.2: A MULTICENTRIC STUDY AND REPORT OF TWO CASES

Deletion 22q11.2 is a chromosomal abnormality detected in young patients with clinical manifestations of the DiGeorge/velocardiofacial syndrome. Conotruncal heart defects are also associated with del22q11.2. An association of these cardiac malformations with neoplasias has been observed. Our series includes two cases of malignancies, a hepatoblastoma and a renal-cell carcinoma, arising in children with complex cardiac malformations. The aim of the study was to determine if the deletion at 22q11.2 was present and could be responsible for both pathological processes. Del22q11.2 was identified in both cases. Comparative genomic hybridization revealed terminal gains on chromosomes 1q and Xq and terminal loss on 1p in the hepatoblastoma, and gains in 1p, 12q, 16p, 20q, 22q, and whole chromosome 19 and loss of Xq in the renal-cell carcinoma. Our results confirm a common genetic basis for cardiac malformations, and del22q11.2 presents a risk factor for the development of pediatric tumours.     

Correction to: Clinical and microbiological effects of non-surgical periodontal treatment in individuals with rheumatoid arthritis: a controlled clinical trial

The effect of periodontal treatment on clinical, microbiological and serological parameters of patients with rheumatoid arthritis (RA) are scarce and controversial. The aim of this study was to investigate the influence of non-surgical periodontal treatment on clinical periodontal status, subgingival bacterial levels of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola and RA activity through a controlled clinical trial on individuals with RA and periodontitis (PE). From a convenience sample, 107 individuals were considered eligible and consecutively allocated in four groups: (1) individuals without PE and RA (− PE-RA, n = 30); (2) individuals without PE and with RA (− PE + RA, n = 23); (3) individuals with PE and RA (+ PE + RA, n = 24); and (4) individuals with PE and without RA (+ PE-RA, n = 30). Full-mouth periodontal clinical examinations, microbiological analysis and Disease Activity Score (DAS-28) evaluations were performed at baseline (T1) and 45 days after non-surgical periodontal treatment (T2). At T1, individuals + PE + RA showed greater severity of PE than + PE-RA individuals. At T2, significant reductions were observed in all periodontal clinical parameters in both groups (p < 0.001) with a significant reduction in DAS-28 in + PE + RA (p = 0.011). Individuals + PE-RA and + PE-RA showed significant reductions for all bacteria (p < 0.001). Additionally, P. gingivalis demonstrated an expressively significant reduction in + PE + RA (p < 0.001). Non-surgical periodontal treatment was effective on improving the clinical periodontal condition, improving the RA clinical status and reducing the presence of periodontal pathogens. Brazilian Registry of Clinical Trials (ReBEC) protocol #RBR-8g2bc8 (https://www.ensaiosclinicos.gov.br/rg/RBR-8g2bc8/).

     

Survival of patients with malignant biliary stricture based on endoscopic brushing and on bilirubinemia

BACKGROUND: Pancreatic cancer and cholangiocarcinoma are the main causes of malignant biliary stricture. Both types of cancers have dismal survival rates, and treatment has little or no effect on prolonging the patients lives. Prognostic definition at initial endoscopic retrograde cholangiopancreatography is important to choose the most suitable management. AIM: To analyse endoscopic brushing and bilirubinemia importance in determination of the survival of patients with malignant biliary stricture. METHODS: Patients with biliary strictures diagnosed during endoscopic retrograde cholangiopancreatography were submitted to double brushing cytology. Serum samples were taken from all patients for bilirubinemia assay. Patients were followed to determine the final diagnosis and survival rates. RESULTS: A total of 50 patients with biliary stricture underwent endoscopic retrograde cholangiopancreatography (40 ultimately found to have a final diagnosis of malignant disease). Hyperbilirubinemia or cytology brushing positive for malignancy was related to a shorter survival rate. CONCLUSION: This research data demonstrate the possibility of determining the prognosis of patients with malignant biliary stricture using endoscopic brushing results and bilirubinemia levels.     

Reconstruction of Chest Wall Using a Two-Layer Prolene Mesh and Bone Cement Sandwich

Wide surgical resection is the most effective treatment for the vast majority of chest wall tumors. This study evaluated the clinical success of chest wall reconstruction using a Prolene mesh and bone cement prosthetic sandwich. The records of all patients undergoing chest wall resection and reconstruction were reviewed. Surgical indications, the location and size of the chest wall defect, diaphragm resection, pulmonary performance, postoperative complications, and survival of each patient were recorded. From 1998 to 2008, 43 patients (27 male, 16 female; mean age of 48 years) underwent surgery in our department to treat malignant chest wall tumors: chondrosarcoma (23), osteosarcoma (8), spindle cell sarcoma (6), Ewing''s sarcoma (2), and others (4). Nine sternectomies and 34 antero-lateral and postero-lateral chest wall resections were performed. Postoperatively, nine patients experienced respiratory complications, and one patient died because of respiratory failure. The overall 4-year survival rate was 60 %. Chest wall reconstruction using a Prolene mesh and bone cement prosthetic sandwich is a safe and effective surgical procedure for major chest wall defects.