Aim
To investigate pathways between unhealthy and healthy dietary patterns and periodontitis in adolescents (18–19 years of age).
Materials and Methods
This population‐based study (n = 2515) modelled ...direct and mediated pathways (via biofilm and obesity) from patterns of healthy diet (fruits, fibre, vegetables, and dairy) and unhealthy diet (sugars, snacks, and salty/fast foods) with initial periodontitis (bleeding on probing BoP, probing depth PD ≥ 4 mm, clinical attachment loss CAL ≥ 4 mm), moderate periodontitis (BoP, PD ≥ 5 mm, and CAL ≥ 5 mm), and European Federation of Periodontology and the American Academy of Periodontology (EFP‐AAP) periodontitis definitions, adjusting for sex, socio‐economic status, smoking, and alcohol, through structural equation modelling (α = 5%).
Results
Higher values of healthy diet were associated with lower values of initial periodontitis (standardized coefficient SC = −0.160; p < .001), moderate periodontitis (SC = −0.202; p < .001), and EFP‐AAP periodontitis (p < .05). A higher value of unhealthy diet was associated with higher values of initial periodontitis (SC = 0.134; p = .005) and moderate periodontitis (SC = 0.180; p < .001). Biofilm mediated the association between higher values of unhealthy diet and all periodontal outcomes (p < .05).
Conclusions
Our findings suggest that both healthy and unhealthy dietary patterns may contribute to reduced or increased extent and severity of periodontitis by local and systemic mechanisms, preceding the effect of other established causes such as smoking and obesity, in younger population.
Abstract Schizophrenia was proposed as a progressive neurodevelopmental disorder. In this regard herein we attempted to determine progressive inflammatory and oxidative alterations induced by a ...neonatal immune challenge and its possible reversal by clozapine administration. For this end, Wistar rats at postnatal day (PN) 5–7 were administered the viral mimetic polyriboinosinic–polyribocytidilic acid (polyI:C) or saline. A distinct group of animals additionally received the antipsychotic drug clozapine (25 mg/kg) from PN60 to 74. At PN35 (periadolescence), 60 (adult) and 74 (adulthood) the animals were submitted to behavioral determinations of prepulse inhibition of the startle (PPI) and Y maze task for working memory evaluation. At PN35 and 74 the animals were sacrificed and the hippocampus (HC), prefrontal cortex (PFC) and striatum (ST) immunostained for Iba-1, a microglial marker, and inducible nitric oxide synthase (iNOS). At PN74 oxidative stress parameters, such as, reduced glutathione levels (GSH) and lipid peroxidation were determined. The results showed a progressive increase of microglial activation and iNOS immunostaining from PN35 to PN74 mainly in the CA2 and CA3 regions of the HC and in the ST. At PN74 neonatal challenge also induced an oxidative imbalance. These inflammatory alterations were accompanied by deficits in PPI and working memory only in adult life that were reversed by clozapine. Clozapine administration reversed microglial activation and iNOS increase, but not the alterations of oxidative stress parameters. Taken together these results give further evidences for a neuroprogressive etiology and course of schizophrenia and that clozapine may partly alleviate this process.
In this work, the acridine nucleus was used as a lead-compound for structural modification by adding different substituted thiosemicarbazide moieties. Eight new ...(Z)-2-(acridin-9-ylmethylene)-N-phenylhydrazinecarbothioamide derivatives (3a-h) were synthesized, their antiproliferative activities were evaluated, and DNA binding properties were performed with calf thymus DNA (ctDNA) by electronic absorption and fluorescence spectroscopies. Both hyperchromic and hypochromic effects, as well as red or blue shifts were demonstrated by addition of ctDNA to the derivatives. The calculated binding constants ranged from 1.74 × 10(4) to 1.0 × 10(6) M(-1) and quenching constants from -0.2 × 10(4) to 2.18 × 10(4) M(-1) indicating high affinity to ctDNA base pairs. The most efficient compound in binding to ctDNA in vitro was (Z)-2-(acridin-9-ylmethylene)-N- (4-chlorophenyl) hydrazinecarbothioamide (3f), while the most active compound in antiproliferative assay was (Z)-2-(acridin-9-ylmethylene)-N-phenylhydrazinecarbothioamide (3a). There was no correlation between DNA-binding and in vitro antiproliferative activity, but the results suggest that DNA binding can be involved in the biological activity mechanism. This study may guide the choice of the size and shape of the intercalating part of the ligand and the strategic selection of substituents that increase DNA-binding or antiproliferative properties.
Engineered 3D models employing human induced pluripotent stem cell (hiPSC) derivatives have the potential to recapitulate the cell diversity and structure found in the human central nervous system ...(CNS). Therefore, these complex cellular systems offer promising human models to address the safety and potency of advanced therapy medicinal products (ATMPs), such as gene therapies. Specifically, recombinant adeno-associated viruses (rAAVs) are currently considered highly attractive for CNS gene therapy due to their broad tropism, low toxicity, and moderate immunogenicity. To accelerate the clinical translation of rAAVs, in-depth preclinical evaluation of efficacy and safety in a human setting is primordial. The integration of hiPSC-derived CNS models in rAAV development will require, amongst other factors, robust, small-scale, high-throughput culture platforms that can feed the preclinical trials.
Herein, we pioneer the miniaturization and parallelization of a 200 mL stirred-tank bioreactor-based 3D brain cell culture derived from hiPSCs. We demonstrate the applicability of the automated miniaturized Ambr
15 Cell Culture system for the maintenance of hiPSC-derived neurospheroids (iNSpheroids), composed of neuronal and glial cells. Critical process parameters were optimized, namely, cell density and agitation mode.
Under optimized conditions, stable iNSpheroid cultures were attained in the microbioreactors for at least 15 days, with high cell viability and astrocytic and neuronal phenotype maintenance. This culture setup allowed the parallelization of different rAAVs, in different multiplicity of infections (MOIs), to address rAAV-host interactions at a preclinical scale. The iNSpheroids were exposed to rAAV2- and rAAV9-eGFP in the microbioreactors. Transgene expression was detected 14 days post-transduction, revealing different astrocyte/neuron tropism of the two serotypes.
We advocate that the iNSpheroid cultures in miniaturized bioreactors are reliable and reproducible screening tools for addressing rAAV transduction and tropism, compatible with preclinical demands.
Brucellosis is an infectious disease caused by bacteria of the genus
Brucella
, which affects domestic animals and is transmissible to humans. The objective of this study was to evaluate six methods ...of DNA extraction directly from bovine tissue to detect
Brucella
spp. The Cq values for all samples were above 30 and varied according to the extraction kit used, but four kits showed no statistical difference in sensitivity. This work demonstrates the importance of choosing the best extraction kit before validation of a molecular diagnostic technique.
Virus-based biopharmaceutical products are used in clinical applications such as vaccines, gene therapy, and immunotherapy. However, their manufacturing remains a challenge, hampered by the lack of ...appropriate analytical tools for purification monitoring or characterization of the final product. This paper describes the implementation of a highly sensitive method, capillary electrophoresis (CE)-sodium dodecyl sulfate (SDS) combined with a laser-induced fluorescence (LIF) detector to monitor the impact of various bioprocess steps on the quality of different viral vectors. The fluorescence labelling procedure uses the (3-(2-furoyl) quinoline-2-carboxaldehyde dye, and the CE-SDS LIF method enables the evaluation of in-process besides final product samples. This method outperforms other analytical methods, such as SDS-polyacrylamide gel electrophoresis with Sypro Ruby staining, in terms of sensitivity, resolution, and high-throughput capability. Notably, this CE-SDS LIF method was also successfully implemented to characterize enveloped viruses such as Maraba virus and lentivirus, whose development as biopharmaceuticals is now restricted by the lack of suitable analytical tools. This method was also qualified for quantification of rAAV2 according to the International Council for Harmonisation guidelines. Overall, our work shows that CE-SDS LIF is a precise and sensitive analytical platform for in-process sample analysis and quantification of different virus-based targets, with a great potential for application in biomanufacturing.
The majority of recombinant adeno‐associated viruses (rAAV) approved for clinical use or in clinical trials areproduced by transient transfection using the HEK293 cell line. However, this platform ...has several manufacturing bottlenecks at commercial scales namely, low product quality (full to empty capsid ratio <20% in most rAAV serotypes), lower productivities obtained after scale‐up and the high cost of raw materials, in particular of Good Manufacturing Practice grade plasmid DNA required for transfection. The HeLa‐based stable cell line rAAV production system provides a robust and scalable alternative to transient transfection systems. Nevertheless, the time required to generate the producer cell lines combined with the complexity of rAAV production and purification processes still pose several barriers to the use of this platform as a suitable alternative to the HEK293 transient transfection. In this work we streamlined the cell line development and bioprocessing for the HeLaS3‐based production of rAAV. By exploring this optimized approach, producer cell lines were generated in 3‐4 months, and presented rAAV2 volumetric production (bulk) > 3 × 1011 vg/mL and full to empty capsids ratio (>70%) at 2 L bioreactor scale. Moreover, the established downstream process, based on ion exchange and affinity‐based chromatography, efficiently eliminated process related impurities, including the Adenovirus 5 helper virus required for production with a log reduction value of 9. Overall, we developed a time‐efficient and robust rAAV bioprocess using a stable producer cell line achieving purified rAAV2 yields > 1 × 1011 vg/mL. This optimized platform may address manufacturing challenges for rAAV based medicines.
Escandell et al., described a streamlined approach for recombinant adeno‐associated viruses (rAAV) production using an HeLaS3‐based stable cell line. This work addresses some of the production system's limitations, including a cumbersome cell line development process, low titers and a complex rAAV production and purification process. This optimized platform can solve scalability issues and high cost of raw materials associated to current industry standards for rAAV manufacturing, therefy accelerating the development of rAAV‐based therapies.
Few studies, each limited to a single major city, have investigated the prevalence and seasonal patterns of different viruses among children with low respiratory tract infections (LRTI) in ...Northeastern Brazil. The aim of this study was to determine the frequency of respiratory syncytial virus (RSV) and of 7 other viruses in children for LRTI in 4 capitals from this region, and investigate their association with several risk factors, including meteorological data. From April 2012 to March 2013, 507 children, aged up to 24 months and hospitalized with LRTI in one of the participating centers at Aracajú, Salvador, Recife, and Maceió, had a sample of nasopharyngeal aspirate collected and analyzed for the following viruses by reverse-transcription polymerase chain reaction followed by hybridization on low-density microarrays: RSV, influenza, parainfluenza, adenovirus, rhinovirus, metapneumovirus, bocavirus, and coronavirus. The result was positive in 66.5% of cases, RSV was the most common virus (40.2%). Except for rhinovirus (17%), all other virus had frequency rates lower than 6%. Viral coinfections were detected in 13.8% of samples. Possible related risk factors for RSV infection were low age upon entry, attendance of daycare, low gestational age, and low educational level of the father. The relative frequency of viral infections was associated with increasing temperature and decreasing humidity separately, but the results also suggested both associated with increased frequency of RSV. Some of these findings differ from those reported for other regions in Brazil and may be used to guide policies that address LRTI.
Clinical presentation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in pediatric immunosuppressed patients is unknown. Emerging data describe a milder or asymptomatic ...course in children compared with adults in this scenario. We present the seroprevalence and clinical features of coronavirus disease 2019 in a prospective cohort of 114 immunosuppressed children and adolescents from three groups: kidney transplantation, liver transplantation, and cancer patients. Among the thirty-five (30.7%) patients who had a positive serological test for SARS-CoV-2, 77% did not report previous symptoms and none of them developed any complications of coronavirus disease 2019 (COVID-19) after 30 or more days of follow-up. Among those who were symptomatic, diarrhea, fever, and cough were the most common findings.
Conclusion
: Seroprevalence of SARS-CoV-2 infection is high among immunosuppressed children and adolescents. COVID-19 has a mild or asymptomatic course in most of these patients.
What is Known:
• The number of immunosuppressed patients with coronavirus disease 2019 is increasing.
• Viral infections have the potential for greater severity in immunocompromised children.
What is New:
• Seroprevalence for severe acute respiratory syndrome coronavirus 2 in immunocompromised pediatric patients was 31%.
• A quarter of the serology-positive patients reported mild symptoms and none of them developed multisystem inflammatory syndrome in children associated with coronavirus disease 2019.
The aim of the present work was to examine possible genetic risk factors related to the occurrence of cerebrovascular disease (CVD) in Brazilian population, the frequency of β(S)-globin gene ...haplotypes and co-inheritance with α-thalassemia (-α(3.7kb)) and single nucleotide polymorphism of methylenetetrahydrofolate reductase (MTHFR-C677T), Factor V Leiden (FV-G1691A) and prothrombin (PT-G20210A) genes in children from Rio de Janeiro. Ninety four children with sickle cell anemia (SCA) were included, 24 patients with cerebrovascular involvement and 70 patients without CVD as control group. The mean age of children at the time of the cerebrovascular event was similar to the control group. The frequency of -α(3.7kb) thalassemia was similar in both groups (p=0.751). Children with Bantu/Atypical β(S)-globin gene haplotype presented 15 times more chance (OR=15.4 CI 95% 2.9-81.6) of CVD than the other β(S)-globin gene haplotypes. The C677T polymorphism of MTHFR gene was similar in both groups (p=0.085). No mutation in the FV Leiden or PT genes was found. A large study seems necessary to establish the role of these genetic polymorphisms in Brazilian miscegenated population.
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