Insulin-like Growth Factor Binding Proteins and Cellular Senescence Are Involved in the Progression of Non-Alcoholic Fatty Liver Disease and Fibrosis in a Mouse Model.

Background and Objectives: Non-alcoholic fatty liver disease (NAFLD) is highly prevalent worldwide. It progresses from simple steatosis to non-alcoholic steatohepatitis (NASH). Fibrosis is often present during NAFLD progression; however, factors determining which subjects develop NASH or fibrosis are unclear. Insulin-like growth factor binding proteins (IGFBPs) are a family of secreted proteins involved in senescence and scarring, mainly synthetized in the liver. Here, we aimed to study the association of IGFBPs and their induced senescence with the progression of NAFLD and liver fibrosis. Materials and Methods: A total of 16-week-old male C57BL/6 mice weighing 23 ± 3 g were fed either methionine/choline-deficient (MCD) or control diet for 2, 8, or 12 weeks. Blood and liver samples were collected, and a histological assessment of NAFLD and fibrosis was performed. Fat contents were measured. Cellular senescence was evaluated in the liver. IGFBP levels were assessed in the liver and serum. Data were expressed as mean ± SD and analyzed by a one-way ANOVA followed by Tukey's test. Lineal regression models were applied for NAFLD and fibrosis progression. p < 0.05 was considered significant. Results: IGFBP-1 and -2 were increased in serum during NAFLD. IGFBP-7 was significantly increased in the serum in NASH compared with the controls. Senescence increased in NAFLD. Serum and liver IGFBP-7 as well as SA-ß-gal activity increased as fibrosis progressed. Both IGFBP-7 and cellular senescence were significantly higher during NAFLD and fibrosis in MCD-fed mice. Conclusions: IGFBP-1, -2, and -7, through their consequent senescence, have a role in the progression of NAFLD and its associated fibrosis, being a plausible determinant in the progression from steatosis to NASH.

Epileptic spasms in clusters with hypsarrhythmia in infancy and childhood: A single age-dependent type of epilepsy or well-defined epileptic syndrome?

OBJECTIVE: In this study, we present the electroclinical features and outcomes of 92 patients with epileptic spasms (ES) in clusters without modified or classical hypsarrhythmia that started in either in infancy or in childhood; we compared both groups in terms of electroclinical features, etiology, treatment, evolution, and outcome. METHODS: Between June 2000 and July 2022, 92 patients met the electroclinical diagnostic criteria of ES in clusters without hypsarrhythmia. Patients with ES associated with other epileptic encephalopathies including West Syndrome, as well as those with the specific etiology of ES and developmental and epileptic encephalopathy associated with CDKL5 were excluded. RESULTS: The patients were divided into two groups based on the age at ES onset: those with ES onset before (Group 1) and those with ES onset after 2 years of age (Group 2). The features of ES and the type of associated seizures before and after ES onset, as well as the interictal and ictal EEG and electromyography findings were similar in both groups. The etiologies were mainly structural (40.2%), genetic (11.9%), and unknown (44.6%) in majority of the patients in both groups. Thirty-one patients were seizure-free, while in the remaining patients the seizures continued. Nine patients (9.8%) with unilateral structural lesions underwent surgery with good results. The neurological abnormalities and developmental findings prior to ES onset depended on the underlying etiology. CONCLUSION: Our series of patients may represent a well-defined epileptic syndrome or type of epilepsy with onset in infancy or childhood characterized by ES in clusters without hypsarrhythmia associated with focal and generalized seizures and EEG paroxysms without neurological deterioration.

Coping and risk perception during the COVID-19 pandemic in type 2 diabetes: Does it influence metabolic control?

Diabetes and poor glycemic control are significant predictors of severity and death in the COVID-19 disease. The perception of this risk in individuals with type 2 diabetes (T2D) could modify coping styles, leading to behaviors associated with better self-care and metabolic control. Theoretically, active coping is associated with better glycemic control in patients with T2D. Nonetheless, information during extreme risk like the COVID-19 pandemic is still limited. Our objective was to evaluate the association between coping styles and risk perception in the COVID-19 pandemic and the change in metabolic parameters. This is a prospective study that included individuals with T2D treated in a tertiary care center during the COVID-19 outbreak who returned to follow-up one year later. We assessed coping styles and risk perception with the Extreme Risk Coping Scale and the risk perception questionnaire. Clinical characteristics and metabolic parameters were registered in both visits. Groups were compared using Kruskal Wallis tests, and changes in metabolic parameters were assessed with Wilcoxon rank sum tests. Our sample included 177 participants at baseline, and 118 concluded the study. Passive coping was more frequent in women. Low-risk perception was associated with higher age, lower psychiatric comorbidities, and lower frequency of psychiatric treatment compared with other risk perception groups. Patients with active coping plus high-risk perception did not have a change in metabolic parameters at follow-up, whereas patients with other coping styles and lower risk perception had an increase in total cholesterol, LDL-cholesterol, and triglycerides. There were no differences by coping group or by risk perception in glycemic control.

In Vitro Lipid Overload Affects Cellular Proliferation, Apoptosis, and Senescence in a Time-Dependent Manner in HepG2 Hepatocytes and LX-2 Hepatic Stellate Cells.

Different cellular mechanisms influence steatotic liver disease (SLD) progression. The influence of different levels of steatogenic inputs has not been studied in hepatocytes and hepatic stellate cells (HSCs). METHODS: HepG2 hepatocytes and LX-2 HSCs were cultured in mild (MS) and severe (SS) steatogenic conditions. TGF-ß stimulation was also tested for HSCs in control (T) and steatogenic conditions (MS-T and SS-T). Steatosis was stained with Oil Red, and the proliferation was assayed via WST-8 reduction, apoptosis via flow cytometry, and senescence via SA-ß-galactosidase activity. RESULTS: Regarding hepatocytes, steatosis progressively increased; proliferation was lower in MS and SS; and the viability of both conditions significantly decreased at 72 h. Apoptosis increased in MS at 72 h, while it decreased in SS. Senescence increased in MS and diminished in SS. Regarding HSCs, the SS and SS-T groups showed no proliferation, and the viability was reduced in MS at 72 h and in SS and SS-T. The LX-2 cells showed increased apoptosis in SS and SS-T at 24 h, and in MS and MS-T at 72 h. Senescence decreased in MS, SS, and SS-T. CONCLUSIONS: Lipid overload induces differential effects depending on the cell type, the steatogenic input level, and the exposure time. Hepatocytes are resilient to mild steatosis but susceptible to high lipotoxicity. HSCs are sensitive to lipid overload, undergoing apoptosis and lowering senescence and proliferation. Collectively, these data may help explain the development of steatosis and fibrosis in SLD.

Effect of Malaria Infection on Epstein-Barr Virus Persistence in Kenyan Children.

BACKGROUND: The 2 cofactors in the etiology of Burkitt lymphoma (BL) are Epstein-Barr virus (EBV) and repeated Plasmodium falciparum malaria infections. This study evaluated EBV loads in mucosal and systemic compartments of children with malaria and controls. Age was analyzed as a covariate because immunity to malaria in endemic regions is age dependent. METHODS: Children (2-10 years) with clinical malaria from Western Kenya and community controls without malaria were enrolled. Saliva and blood samples were collected, EBV viral load was assessed by quantitative polymerase chain reaction, and EpiTYPER MassARRAY was used to assess methylation of 3 different EBV genes. RESULTS: Regardless of the compartment, we detected EBV more frequently in malaria cases compared to controls, although the difference was not significant. When EBV was detected, there were no differences in viral load between cases and controls. However, EBV methylation was significantly lower in the malaria group compared to controls in both plasma and saliva (P < .05), indicating increased EBV lytic replication. In younger children before development of immunity to malaria, there was a significant effect of malaria on EBV load in peripheral blood mononuclear cells (P = .04). CONCLUSIONS: These data suggest that malaria can directly modulate EBV persistence in children, increasing their risk for BL.

Brivaracetam as add-on therapy in children with developmental epileptic encephalopathies: A study of 42 patients.

OBJECTIVE: Here we present a multicenter series of patients with developmental epileptic encephalopathies (DEE) who were treated with brivaracetam (BRV) as add-on therapy. METHODS: Medical records of 42 patients with DEE treated with add-on BRV seen at four pediatric neurology centers in Argentina between January 2021 and July 2023 were retrospectively analyzed. RESULTS: We included 42 patients (26 males, 16 females) with a mean age of 7 years (SD, ± 3.8; median, 9; range, 2-16). The children had different types of childhood-onset treatment-resistant DEEs and received BRV as add-on therapy for a mean period of 2 years (SD, ± 1.3 years; median, 1.5 years; range, 0.5-3 years). Thirty-three patients received levetiracetam (LEV) before the introduction of BRV. In nine patients, BRV was started without prior LEV because of behavioral disturbances. Three patients (9.5 %) became seizure free and 26/42 patients (62.1 %) had a greater than 50 % decrease in seizures after a mean follow-up of 21 months. Ten patients (23.8 %) had a 25-50 % seizure reduction, while seizure frequency remained unchanged in two (4.7 %) and increased in one patient (2.4 %). The interictal EEG abnormalities improved in all the responders. Adverse effects, consisting of drowsiness, irritability, and decreased appetite, were observed in seven patients (16.6 %), but did not lead to treatment discontinuation. CONCLUSION: Brivaracetam was found to be effective, safe, and well tolerated in children with DEE. In patients on LEV with behavioral disturbances, BRV may be tried. BRV may also be given without a previous trial with LEV in patients with behavioral problems.

Differential Association of Glycation Products with Bone Mineral Density and Fat Mass in Healthy and Diabetes Type 2 Subjects from Mexican Southeastern: A Cross Sectional Study.

Background: Glycation products have been linked to decreased bone mineral density (BMD) in a number of clinical settings. This study examined the correlation between early glycation products (HbA1c and glycated albumin (ALB-g)) and advanced glycation end products (pentosidine (PTD)) with BMD in two groups of participants: those with type 2 diabetes mellitus (DM2) and those without diabetes or any other comorbidities (noDM). All of the participants had resided in southeastern Mexico for a minimum of 10 years. Material and Methods: This study included 204 participants: 112 (55%) with DM2 and 92 (45%) healthy subjects. We utilized dual X-ray absorptiometry (DXA) to measure both the total and segment-specific BMD and adipose mass. In addition, the fasting blood glucose, HbA1c, PTD, and ALB-g parameters were measured. Correlation and logistic regression analyses were conducted. Results: There was an inverse correlation between PTD and BMD in all anatomical regions among postmenopausal women (PMW) in the DM2 group, whereas in non-PMW, only the waist-to-height ratio was statistically significant. A negative correlation was observed between HbA1c levels and BMD in the arms and legs of DM2 individuals. However, in the noDM group, a negative correlation was found between HbA1c levels and BMD in the pelvis, while a positive association was observed between HbA1c and indicators of adipose tissue. ALB-g, demonstrated a negative correlation with fat mass. After performing binary logistic regressions, the following odds ratios (OR) for osteopenia/osteoporosis risk were determined: PTD OR 1.1 (p = 0.047) for DM2 PMW, HbA1c OR 1.4 (p = 0.048), and fat mass content OR 1.011 (p = 0.023) for the entire sample. Conclusions: Glycation products are associated with BMD differentially depending on the analyzed anatomical segment, but PTD, HbA1c, and fat mass are significant predictors of low bone mass. In prospective studies, this association could be determined using other techniques involving three-dimensional analysis of bone architecture to evaluate bone architecture.

Effects of Maternal HIV Infection on Early Kaposi Sarcoma-Associated Herpesvirus Seroconversion in a Kenyan Mother-Infant Cohort.

BACKGROUND: We identified whether maternal human immunodeficiency virus (HIV) infection during pregnancy affects transplacental transfer of Kaposi sarcoma-associated herpesvirus (KSHV)-specific antibodies and subsequent infant infection. METHODS: We followed pregnant Kenyan women through delivery and their infants until age 2 years. Children were classified as HIV-exposed uninfected (HEU) or HIV-unexposed uninfected (HUU) based on maternal HIV status. Maternal venous and cord blood at delivery and child venous blood every 6 months were tested for antibodies to 20 KSHV antigens by multiplex bead-based immunoassay. Multiple comparisons were adjusted using false discovery rate (FDR). RESULTS: Maternal HIV infection was significantly associated with decreased transplacental transfer of antibodies against all KSHV antigens and lower cord blood levels for 8 antigens at FDR P < .10. Neither birth to 6-month antibody level changes nor 6-month levels differed in HEU and HUU, except for ORF50. By age 24 months, 74% of children KSHV seroconverted but HEU and HUU did not differ in time to seroconversion nor 2-year seropositivity after adjustment for child malaria infection. CONCLUSIONS: Maternal HIV infection reduced a child's initial KSHV antibody levels but did not affect age of infection. Regardless of HIV exposure in utero, KSHV seroconversion in Kenyan children occurred early; associated factors must be identified.

Macrophages drive KSHV B cell latency.

Kaposi's sarcoma herpesvirus (KSHV) establishes lifelong infection and persists in latently infected B cells. Paradoxically, in vitro B cell infection is inefficient, and cells rapidly die, suggesting the absence of necessary factor(s). KSHV epidemiology unexpectedly mirrors that of malaria and certain helminthic infections, while other herpesviruses are ubiquitous. Elevated circulating monocytes are common in these parasitic infections. Here, we show that KSHV infection of monocytes or M-CSF-differentiated (M2) macrophages is highly efficient. Proteomic analyses demonstrate that infection induces macrophage production of B cell chemoattractants and activating factor. We find that KSHV acts with monocytes or M2 macrophages to stimulate B cell survival, proliferation, and plasmablast differentiation. Further, macrophages drive infected plasma cell differentiation and long-term viral latency. In Kenya, where KSHV is endemic, we find elevated monocyte levels in children with malaria. These findings demonstrate a role for mononuclear phagocytes in KSHV B cell latency and suggest that mononuclear phagocyte abundance may underlie KSHV's geographic disparity.

Evidence for Aerosol Transfer of SARS-CoV-2-Specific Humoral Immunity.

Infectious particles can be shared through aerosols and droplets formed as the result of normal respiration. Whether Abs within the nasal/oral fluids can similarly be shared between hosts has not been investigated. The circumstances of the SARS-CoV-2 pandemic facilitated a unique opportunity to fully examine this provocative idea. The data we show from human nasal swabs provides evidence for the aerosol transfer of Abs between immune and nonimmune hosts.

[Solid pseudopapillary neoplasm. Report of three cases and review of the literature]. / Neoplasia sólida pseudopapilar. Reporte de tres casos y revisión de la literatura.

Background: Solid pseudopapillary neoplasm, or Frantz-Gruber tumor, is a diagnostic challenge for the surgical pathologist. It is classified by the WHO as a malignant epithelial tumor of the pancreas, its prevalence is low, it occurs in only 1 to 2% of all malignant tumors of the pancreas, it usually affects mainly young women, its origin is still unclear, it is generally It presents as a solitary, encapsulated lesion, without invasion of peripancreatic tissues with rare cases of metastasis, which is why it is considered a low-grade malignant tumor by the WHO. The objective of this article is to present three clinical cases and to evaluate the epidemiology, clinical manifestations, morphology and immunohistochemical expression of the tumor in a review of the bibliography, as well as to compare it with the cases already reported on the subject. Clinical case: Three cases of Frantz tumor were diagnosed by the pathology department of a tertiary hospital are presented, which correspond to two women aged 17 and 34, as well as a 52-year-old man whose presentation by age and sex is rare. Conclusions: After the bibliographical review and the analysis of the cases presented, we verified the difficulty to make a correct diagnosis, since its presence is rare in the daily practice of the surgical pathologist. The morphological patterns of the solid pseudopapillary tumor are varied and can often be reminiscent of neuroendocrine tumors of the pancreas, whose presentation rate is higher.

Use of sulthiame as add-on therapy in children with non-self-limited focal epilepsies of childhood.

PURPOSE: This retrospective study aimed to evaluate the efficacy and tolerability of sulthiame (STM) as an add-on treatment in 49 patients with non-self-limited focal epilepsies of childhood (non-SeLFE) resistant to other antiseizure medications (ASM) and/or non-pharmacological treatment. METHODS: Patients with non-SeLFE who had failed to respond to at least five previous ASM, alone or in combination, were included in the study. All patients underwent neurological examination, brain magnetic resonance imaging repeated prolonged electroencephalography (EEG) or video-EEG studies, and neurometabolic studies. School achievements and/or performance on neuropsychological tests were also assessed. Sulthiame was added in doses ranging from 10 to 40 mg/kg/day. Efficacy was measured by comparing seizure frequency before and after initiating STM therapy. RESULTS: Twenty-nine of 49 patients (59.1%) who received STM as add-on therapy had a greater than 50% decrease in seizures after a mean follow-up of 35 months. One patient (2%) became seizure-free. Fourteen patients (40%) had a 25-50% seizure reduction. The mean time of response was 5 months (range, 3.5 to 6 months). No differences were found either between patients with a response of more or less than 50% or between the response of the focal seizure types (motor or non-motor, with or without consciousness impairment). CONCLUSION: In our study, STM was found to be effective and well-tolerated in children and adolescents with non-SeLFE. In the patients who responded, improvement in the EEG was seen.

Collagen matrix scaffolds: Future perspectives for the management of chronic liver diseases.

Approximately 1.5 billion chronic liver disease (CLD) cases have been estimated worldwide, encompassing a wide range of liver damage severities. Moreover, liver disease causes approximately 1.75 million deaths per year. CLD is typically characterized by the silent and progressive deterioration of liver parenchyma due to an incessant inflammatory process, cell death, over deposition of extracellular matrix proteins, and dysregulated regeneration. Overall, these processes impair the correct function of this vital organ. Cirrhosis and liver cancer are the main complications of CLD, which accounts for 3.5% of all deaths worldwide. Liver transplantation is the optimal therapeutic option for advanced liver damage. The liver is one of the most common organs transplanted; however, only 10% of liver transplants are successful. In this context, regenerative medicine has made significant progress in the design of biomaterials, such as collagen matrix scaffolds, to address the limitations of organ transplantation (e.g., low donation rates and biocompatibility). Thus, it remains crucial to continue with experimental and clinical studies to validate the use of collagen matrix scaffolds in liver disease.

Development of an ectopic huLiver model for Plasmodium liver stage infection.

Early Plasmodium falciparum and P. vivax infection requires parasite replication within host hepatocytes, referred to as liver stage (LS). However, limited understanding of infection dynamics in human LS exists due to species-specificity challenges. Reported here is a reproducible, easy-to-manipulate, and moderate-cost in vivo model to study human Plasmodium LS in mice; the ectopic huLiver model. Ectopic huLiver tumors were generated through subcutaneous injection of the HC-04 cell line and shown to be infectible by both freshly dissected sporozoites and through the bite of infected mosquitoes. Evidence for complete LS development was supported by the transition to blood-stage infection in mice engrafted with human erythrocytes. Additionally, this model was successfully evaluated for its utility in testing antimalarial therapeutics, as supported by primaquine acting as a causal prophylactic against P. falciparum. Presented here is a new platform for the study of human Plasmodium infection with the potential to aid in drug discovery.

Pancreatic Cancer: Genetic Conditions and Epigenetic Alterations.

BACKGROUND: Pancreatic cancer is a lethal proliferative disease driven by multiple genetic and epigenetic alterations. Microarrays and omics-based sequencing techniques are potent tools that have facilitated a broader understanding of the complex biological processes that drive pancreatic ductal adenocarcinoma (PDAC). In turn, these tools have resulted in the identification of novel disease markers, prognostic factors, and therapeutic targets. Herein, we provide a review of the genetic and epigenetic drivers of PDAC relative to recent discoveries that impact patient management. METHODS: A review of PubMed, Medline, Clinical Key, and Index Medicus was conducted to identify literature from January 1995 to July 2022 that is related to PDAC genetics and epigenetics. Articles in Spanish and English were considered during selection. RESULTS: Molecular, genetic, and epigenetic diagnostic tools, novel biomarkers, and promising therapeutic targets have emerged in the treatment of pancreatic cancer. The implementation of microarray technology and application of large omics-based data repositories have facilitated recent discoveries in PDAC. Multiple molecular analyses based on RNA interference have been instrumental in the identification of novel therapeutic targets for patients with PDAC. Moreover, microarrays and next-generation omics-based discoveries have been instrumental in the characterization of subtypes of pancreatic cancer, thereby improving prognostication and refining patient selection for available targeted therapies. CONCLUSION: Advances in molecular biology, genetics, and epigenetics have ushered in a new era of discovery in the pathobiology of PDAC. Current efforts are underway to translate these findings into clinical tools and therapies to improve outcomes in patients with PDAC.

Screen-related discord and dismay in low-income Mexican American families with toddlers: A qualitative study.

PURPOSE: Understanding parental experiences with managing their toddler's screen use is important to inform the design of interventions addressing early childhood screen use, yet current evidence is limited. To enhance our understanding of the context of toddler screen use, this study characterizes the screen-related discord and dismay parents experience in families with toddlers. DESIGN AND METHODS: In-depth interviews were conducted to explore everyday experiences with screen use among low-income Mexican American caregivers of toddlers (21 mothers, 10 fathers, 1 grandmother). Transcripts were content analyzed to identify prominent themes. RESULTS: Three themes were identified. Experiences of screen-related discord and dismay arose (1) between parent and child, (2) between parents, and (3) surfaced as parental internal dissonance about toddler screen use. Parent-child discord resulted from parental limit setting and child reactions to parental screen use, which often included tantrums. Parent-partner discord included patterns of agreeing to disagree and direct disagreement between partners. Parents also reported their own feelings of ambivalence and dismay as they struggled to reconcile their preferences against their toddler's actual screen use, while living in a screen-saturated world. CONCLUSIONS: Findings offer insight into types of screen-related discord and dismay low-income Mexican American parents experience as they attempt to manage their toddler's screen use. PRACTICE IMPLICATIONS: Although discord in families is normal, the screen-specific discord reported by participants warrants consideration in efforts promoting healthy screen use in families. Providers can tailor their counseling to consider the range of screen-related discord families of toddlers may experience.

Clinical and genetic study of developmental and epileptic encephalopathy in Argentinean pediatric patients.

INTRODUCTION: The aim of this study was to extend our knowledge of the genetic background of Argentinean pediatric patients with developmental and epileptic encephalopathy (DEE) applying a next generation sequencing (NGS) panel. METHODS: Thirty one patients with DEE were studied, including these phenotypes: Dravet syndrome (n:7), Dravet like syndrome (n:3), West syndrome (WS) (n:6), WS that evolved to Lennox-Gastaut syndrome (LGS) (n:4), epilepsy of infancy with migrating focal seizures (n:2), continuous spikes and waves during slow sleep evolving to LGS (n:1), LGS (n:1), myoclonic status in non-progressive encephalopathy (n:1), myoclonic atonic epilepsy (n:1), epileptic encephalopathy with multifocal spikes (n:1) and unclassified epileptic encephalopathy (n:4). Fifty-two genes frequently associated with DEE were studied by NGS in genomic DNA from peripheral blood. RESULTS: Relevant variants were detected in 12 cases; 6 novel pathogenic or likely pathogenic variants, 6 previously reported as pathogenic and 1 variant of unknown significance. Single-nucleotide heterozygous variants were identified in the SCN1A (5), GABRG2 (1), STXBP1 (2) genes, a mosaic variant in SCN2A (1) and a homozygous variant in SCN1B (1). Additionally, a heterozygous deletion involving the SCN1A, SCN2A and SCN3A genes (1), and the most frequent triplet repeat expansion in the ARX gene (1) were detected. DISCUSSION: Genetic diagnosis was made in 39% of patients. We emphasize the importance of considering mosaic variants, copy number variants and hereditary forms when designing and interpreting molecular studies, to optimize diagnosis and management of patients. Approximately 42% of the detected variants were novel, expanding the knowledge of the molecular basis of DEEs in Latin-American patients.

Introducción: El objetivo del estudio fue ampliar el conocimiento de las bases moleculares de las encefalopatías epilépticas y del desarrollo (EED) en pacientes pediátricos argentinos aplicando un panel de secuenciación de nueva generación (NGS). Métodos: Se analizaron 31 pacientes con los fenotipos clínicos de síndrome de Dravet (n:7), síndrome símil Dravet (n:3), síndrome de West (SW) (n:6), SW que evoluciona a síndrome de Lennox Gastaut (SLG)(N:4), epilepsia de la infancia con crisis focales migratorias (n:2), actividad de punta onda continua durante el sueño que evolucionan a SLG (n:1), SLG (n:1), encefalopatía no progresiva con estatus mioclónico (n:1), epilepsia mioclónica atónica (n:1), encefalopatía epiléptica con espigas multifocales (n:1) y encefalopatía epiléptica indeterminada (n:4). Se estudiaron los 52 genes más frecuentemente asociados a EED a través de NGS, en ADN extraído de sangre periférica. Resultados: Se identificaron variantes relevantes en 12 casos, de las cuales 5 fueron nuevas y 6 previamente reportadas como patogénicas o posiblemente patogénicas, mientras que una variante fue clasificada como de significado incierto. Variantes heterocigotas, de nucleótido único, se identificaron en los genes SCN1A (5), GABRG2 (1), STXBP1 (2), una variante en mosaico en SCN2A (1) y otra homocigota en SCN1B (1). Además, se detectó una deleción que involucra a los genes SCN1A, SCN2A y SCN3A (1) y la expansión de repeticiones de tripletes más frecuente en el gen ARX (1). Discusión: Se alcanzó el diagnóstico molecular en el 39% de los pacientes. Remarcamos la importancia de considerar variantes en mosaico, variantes en el número de copias y formas heredadas al momento de diseñar e interpretar los estudios moleculares, de tal forma de optimizar el diagnóstico y seguimiento de los pacientes con EED. Cabe destacar, que el 42% de las variantes detectadas fueron nuevas, ampliando nuestro conocimiento sobre las bases moleculares de las EED en población latino americana.

Clinical and genetic study of developmental and epileptic encephalopathy in argentinean pediatric patients / Estudio clínico y genético de encefalopatía epiléptica y del desarrollo en pacientes pediátricos argentinos

Abstract Introduction: The aim of this study was to extend our knowledge of the genetic background of Argentinean pediatric patients with developmental and epileptic encephalopathy (DEE) applying a next generation sequencing (NGS) panel. Methods: Thirty one patients with DEE were studied, including these phenotypes: Dravet syndrome (n:7), Dravet like syndrome (n:3), West syndrome (WS) (n:6), WS that evolved to Lennox-Gastaut syndrome (LGS) (n:4), epilepsy of infancy with migrating focal seizures (n:2), continuous spikes and waves during slow sleep evolving to LGS (n:1), LGS (n:1), myoclonic status in non-progressive encephalopathy (n:1), myoclonic atonic epilepsy (n:1), epileptic encephalopathy with multifocal spikes (n:1) and unclassified epileptic encephalopathy (n:4). Fifty-two genes frequently associated with DEE were studied by NGS in genomic DNA from peripheral blood. Results: Relevant variants were detected in 12 cases; 6 novel pathogenic or likely pathogenic variants, 6 previously reported as pathogenic and 1 variant of unknown sig nificance. Single-nucleotide heterozygous variants were identified in the SCN1A (5), GABRG2 (1), STXBP1 (2) genes, a mosaic variant in SCN2A (1) and a homozygous variant in SCN1B (1). Additionally, a heterozygous deletion involving the SCN1A, SCN2A and SCN3A genes (1), and the most frequent triplet repeat expansion in the ARX gene (1) were detected. Discussion: Genetic diagnosis was made in 39% of patients. We emphasize the importance of considering mosaic variants, copy number variants and hereditary forms when designing and interpreting molecular studies, to optimize diagnosis and management of patients. Approximately 42% of the de tected variants were novel, expanding the knowledge of the molecular basis of DEEs in Latin-American patients.

Resumen Introducción: El objetivo del estudio fue ampliar el conocimiento de las bases moleculares de las encefalopatías epilépticas y del desarrollo (EED) en pacientes pediátricos argentinos aplicando un panel de secuenciación de nueva generación (NGS). Métodos: Se analizaron 31 pacientes con los fenotipos clínicos de síndrome de Dra vet (n:7), síndrome símil Dravet (n:3), síndrome de West (SW) (n:6), SW que evoluciona a síndrome de Lennox Gastaut (SLG)(N:4), epilepsia de la infancia con crisis focales migratorias (n:2), actividad de punta onda continua durante el sueño que evolucionan a SLG (n:1), SLG (n:1), encefalopatía no progresiva con estatus mioclónico (n:1), epilepsia mioclónica atónica (n:1), encefalopatía epiléptica con espigas multifocales (n:1) y encefalopatía epiléptica indeterminada (n:4). Se estudiaron los 52 genes más frecuentemente asociados a EED a través de NGS, en ADN extraído de sangre periférica. Resultados: Se identificaron variantes relevantes en 12 casos, de las cuales 5 fueron nuevas y 6 previamente reportadas como patogénicas o posiblemente patogénicas, mien tras que una variante fue clasificada como de significado incierto. Variantes heterocigotas, de nucleótido único, se identificaron en los genes SCN1A (5), GABRG2 (1), STXBP1 (2), una variante en mosaico en SCN2A (1) y otra homocigota en SCN1B (1). Además, se detectó una deleción que involucra a los genes SCN1A, SCN2A y SCN3A (1) y la expansión de repeticiones de tripletes más frecuente en el gen ARX (1). Discusión: Se alcanzó el diagnóstico molecular en el 39% de los pacientes. Remarcamos la importancia de considerar variantes en mosaico, variantes en el número de copias y formas heredadas al momento de diseñar e interpretar los estudios moleculares, de tal forma de optimizar el diagnóstico y seguimiento de los pacientes con EED. Cabe destacar, que el 42% de las variantes detectadas fueron nuevas, ampliando nuestro conocimiento sobre las bases mole culares de las EED en población latino americana.

Influence of ligand density variations on the two peak elution behavior of a monoclonal antibody in cation exchange chromatography.

Cation exchange chromatography, as part of the monoclonal antibody purification train, is known as a mild polishing technique. However, in the last couple of years, more and more publications have shown unusual elution behavior, resulting from e.g. on-column (reversible) unfolding and aggregation of the predominantly mAb molecules. The stability of the investigated protein seems to play a significant role in this phenomenon. We have used a glycosylated IgG1 antibody as a model protein and investigated several influencing factors, including pH value and ligand density variations of three prototype Fractogel® cation exchange resins. Ligand density, pH and salt concentration are the main contributing factors in the Donnan effect, i.e. distribution of ions, between resin pore volume and bulk volume. This leads to a significantly lower pH value the protein is subjected to during the on-column hold time and therefore influences the conformational stability of our protein. Nano-DSF and kinetic SEC measurements show that the protein is destabilized at low pH values, but also, that the binding to the CEX resin and the elution with increasing salt concentration is responsible for the resulting two-peak elution behavior and partially reversible unfolding and aggregation.

Xenoimplant of Collagen Matrix Scaffold in Liver Tissue as a Niche for Liver Cells.

Hepatitis C virus-induced liver damage, chronic liver damage due to alcohol, and non-alcoholic liver disease-induced cellular alterations promote fibrosis, cirrhosis, and/or hepatocellular carcinoma. The recommended therapeutic option for advanced liver damage is liver transplantation. Extracellular matrix scaffolds have been evaluated as an alternative for tissue restoration. Studies on the biocompatibility and rejection of synthetic and natural scaffolds as an alternative to organ transplantation have been evaluated. Our group has recently described the xenoimplant of collagen matrix scaffold (CMS) in a rat model. However, no complete macroscopic and histological description of the liver parenchyma at the initial (day 3), intermediate (day 14), and advanced (day 21) stages has been obtained. In this study, we described and compared liver tissue from the CMS zone (CZ, CMS, and liver parenchyma), liver tissue from the normal zone (liver parenchyma close to the CMS), and basal tissue (resected tissue from the CMS implantation site). Our data strongly suggest that the collagen matrix xenoimplant is a good niche for hepatocytes, with no rejection, and does not affect liver function tests. The liver can regenerate after damage, but this capacity is inhibited in a chronic injury. At present, the use of CMS after liver damage has not been reported. This biomaterial could be a novel alternative in the field of regenerative medicine for liver diseases.