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[This corrects the article DOI: 10.3389/fimmu.2022.871766.].

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Probability distributions offer the best description of survival data and as a result, various lifetime models have been proposed. However, some of these survival datasets are not followed or sufficiently fitted by the existing proposed probability distributions. This paper presents a novel Kumaraswamy Odd Ramos-Louzada-G (KumORL-G) family of distributions together with its statistical features, including the quantile function, moments, probability-weighted moments, order statistics, and entropy measures. Some relevant characterizations were obtained using the hazard rate function and the ratio of two truncated moments. In light of the proposed KumORL-G family, a five-parameter sub-model, the Kumaraswamy Odd Ramos-Louzada Burr XII (KumORLBXII) distribution was introduced and its parameters were determined with the maximum likelihood estimation (MLE) technique. Monte Carlo simulation was performed and the numerical results were used to evaluate the MLE technique. The proposed probability distribution's significance and applicability were empirically demonstrated using various complete and censored datasets on the survival times of cancer and diabetes patients. The analytical results showed that the KumORLBXII distribution performed well in practice in comparison to its sub-models and several other competing distributions. The new KumORL-G for diabetes and cancer survival data is found extremely efficient and offers an enhanced and novel technique for modeling survival datasets.

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Cell line generation of mammalian cells is a time-consuming and labor-intensive process, especially because of challenges in clone selection after transfection. Antibiotics are common selection agents for mammalian cells due to their simplicity of use. However, the optimal antibiotic concentration must be determined with a kill curve experiment before clone selection starts. The traditional kill curve experiments are resource-intensive and time-consuming due to necessary sampling and offline analysis effort. This study, thus, explores the potential of online monitoring the oxygen transfer rate (OTR), as a non-invasive and efficient alternative for kill curve experiments. The OTR is monitored using the Transfer-rate Online Measurement (TOM) system and the micro(µ)-scale Transfer-rate Online Measurement (µTOM) device, which was used for mammalian cells first. It could be shown that the OTR curves for both devices align perfectly, affirming consistent cultivation conditions. The µTOM device proves effective in performing kill curve experiments in 96-deep-well plates without the need for sampling and offline analysis. The streamlined approach reduces medium consumption by 95%, offering a cost-effective and time-efficient solution for kill curve experiments. The study validates the generalizability of the method by applying it to two different CHO cell lines (CHO-K1 and sciCHO) with two antibiotics (puromycin and hygromycin B) each. In conclusion, the broad application of OTR online monitoring for CHO cell cultures in 96-deep-well plates is highlighted. The µTOM device proves as a valuable tool for high-throughput experiments, paving the way for diverse applications, such as media and clone screening, cytotoxicity tests, and scale-up experiments.

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Purpose: Sinomenine hydrochloride (SH) is used to treat chronic inflammatory diseases such as rheumatoid arthritis and may also be efficacious against Immunoglobulin A nephropathy (IgAN). However, no trial has investigated the molecular mechanism of SH on IgAN. Therefore, this study aims to investigate the effect and mechanism of SH on IgAN. Methods: The pathological changes and IgA and C3 depositions in the kidney of an IgAN rat model were detected by periodic acid-Schiff (PAS) and direct immunofluorescence staining. After extracting T and B cells using immunomagnetic beads, we assessed their purity, cell cycle phase, and apoptosis stage through flow cytometry. Furthermore, we quantified cell cycle-related and apoptosis-associated proteins by Western blotting. Results: SH reduced IgA and C3 depositions in stage 4 IgAN, thereby decreasing inflammatory cellular infiltration and mesangial injury in an IgAN model induced using heteroproteins. Furthermore, SH arrested the cell cycle of lymphocytes T and B from the spleen of IgAN rats. Regarding the mechanism, our results demonstrated that SH regulated the Cyclin D1 and Cyclin E1 protein levels for arresting the cell cycle and it also regulated Bax and Bcl-2 protein levels, thus increasing Cleaved caspase-3 protein levels in Jurkat T and Ramos B cells. Conclusion: SH exerts a dual regulation on the cell cycle and apoptosis of T and B cells by controlling cell cycle-related and apoptosis-associated proteins; it also reduces inflammatory cellular infiltration and mesangial proliferation. These are the major mechanisms of SH in IgAN.

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The present study is based on the derivation of a new extension of the Poisson distribution using the Ramos-Louzada distribution. Several statistical properties of the new distribution are derived including, factorial moments, moment-generating function, probability moments, skewness, kurtosis, and dispersion index. Some reliability properties are also derived. The model parameter is estimated using different classical estimation techniques. A comprehensive simulation study was used to identify the best estimation method. Bayesian estimation with a gamma prior is also utilized to estimate the parameter. Three examples were used to demonstrate the utility of the proposed model. These applications revealed that the PRL-based model outperforms certain existing competing one-parameter discrete models such as the discrete Rayleigh, Poisson, discrete inverted Topp-Leone, discrete Pareto and discrete Burr-Hatke distributions.

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Costly complex media components such as yeast extract and peptone are still widely used in industrial bioprocesses, despite their ill-defined composition. Side stream products such as corn steep liquor (CSL) present a compelling economical alternative that contains valuable nutrients required for microbial growth, that is, nitrogen and amino acids, but also vitamins, trace elements, and other minerals. However, as a side stream product, CSL may be subject to batch-to-batch variations and compositional heterogeneity. In this study, the Respiration Activity MOnitoring System designed for shake flasks (RAMOS) and 96-well microtiter plates (µTOM) were applied to investigate the potential and constraints of CSL utilization for two model microorganisms: E. coli and B. subtilis. Considering the dry substance content of complex nutrients involved, CSL-based media are more efficient in biomass production than the common lysogeny broth (LB) medium, containing 5 g/L yeast extract, 10 g/L peptone, and 5 g/L NaCl. At a glucose to CSL (glucose/CSL, g/g) ratio of 1/1 (g/g) and 2/1 (g/g), a secondary substrate limitation occurred in E. coli and B. subtilis cultivations, respectively. The study sheds light on differences in the metabolic activity of the two applied model organisms between varying CSL batches, which relate to CSL origin and production process, as well as the effect of targeted nutrient supplementation. Through a targeted nutrient supplementation, the most limiting component of the CSL-glucose medium used for these applied model microorganisms was identified to be ammonium nitrogen. This study proves the suitability of CSL as an alternative nutrient source for E. coli and B. subtilis. The RAMOS and µTOM technique detected differences between CSL batches, allowing easy and early identification of varying batches. A consistent performance of the CSL batches in E. coli and B. subtilis cultivations was demonstrated.

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BACKGROUND: In research and production, reproducibility is a key factor, to meet high quality and safety standards and maintain productivity. For microbial fermentations, complex substrates and media components are often used. The complex media components can vary in composition, depending on the lot and manufacturing process. These variations can have an immense impact on the results of biological cultivations. The aim of this work was to investigate and characterize the influence of the complex media component yeast extract on cultivations of Azotobacter vinelandii under microaerobic conditions. Under these conditions, the organism produces the biopolymer alginate. The focus of the investigation was on the respiration activity, cell growth and alginate production. RESULTS: Yeast extracts from 6 different manufacturers and 2 different lots from one manufacturer were evaluated. Significant differences on respiratory activity, growth and production were observed. Concentration variations of three different yeast extracts showed that the performance of poorly performing yeast extracts can be improved by simply increasing their concentration. On the other hand, the results with well-performing yeast extracts seem to reach a saturation, when their concentration is increased. Cultivations with poorly performing yeast extract were supplemented with grouped amino acids, single amino acids and micro elements. Beneficial results were obtained with the supplementation of copper sulphate, cysteine or a combination of both. Furthermore, a correlation between the accumulated oxygen transfer and the final viscosity (as a key performance indicator), was established. CONCLUSION: The choice of yeast extract is crucial for A. vinelandii cultivations, to maintain reproducibility and comparability between cultivations. The proper use of specific yeast extracts allows the cultivation results to be specifically optimised. In addition, supplements can be applied to modify and improve the properties of the alginate. The results only scratch the surface of the underlying mechanisms, as they are not providing explanations on a molecular level. However, the findings show the potential of optimising media containing yeast extract for alginate production with A. vinelandii, as well as the potential of targeted supplementation of the media.

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Cultivating microorganisms on solid agar media is a fundamental technique in microbiology and other related disciplines. For the evaluation, most often, a subjective visual examination is performed. Crucial information, such as metabolic activity, is not assessed. Thus, time-resolved monitoring of the respiration activity in agar cultivations is presented to provide additional insightful data on the metabolism. A modified version of the Respiration Activity MOnitoring System (RAMOS) was used to determine area-specific oxygen and carbon dioxide transfer rates and the resulting respiratory quotients of agar cultivations. Therewith, information on growth, substrate consumption, and product formation was obtained. The validity of the presented method was tested for different prokaryotic and eukaryotic organisms on agar, such as Escherichia coli BL21, Pseudomonas putida KT2440, Streptomyces coelicolor A3(2), Saccharomyces cerevisiae WT, Pichia pastoris WT, and Trichoderma reesei RUT-C30. Furthermore, it is showcased that several potential applications, including the determination of colony forming units, antibiotic diffusion tests, quality control for spore production or for pre-cultures and media optimization, can be quantitatively evaluated by interpretation of the respiration activity.

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BACKGROUND: The efficiency of downstream processes plays a crucial role in the transition from conventional petrochemical processes to sustainable biotechnological production routes. One promising candidate for product separation from fermentations with low energy demand and high selectivity is the adsorption of the target product on hydrophobic adsorbents. However, only limited knowledge exists about the interaction of these adsorbents and the bioprocess. The bioprocess could possibly be harmed by the release of inhibitory components from the adsorbent surface. Another possibility is co-adsorption of essential nutrients, especially in an in situ application, making these nutrients unavailable to the applied microorganism. RESULTS: A test protocol investigating adsorbent-bioprocess compatibility was designed and applied on a variety of adsorbents. Inhibitor release and nutrient adsorption was studied in an isolated manner. Respiratory data recorded by a RAMOS device was used to assess the influence of the adsorbents on the cultivation in three different microbial systems for up to six different adsorbents per system. While no inhibitor release was detected in our investigations, adsorption of different essential nutrients was observed. CONCLUSION: The application of adsorption for product recovery from the bioprocess was proven to be generally possible, but nutrient adsorption has to be assessed for each application individually. To account for nutrient adsorption, adsorptive product separation should only be applied after sufficient microbial growth. Moreover, concentrations of co-adsorbed nutrients need to be increased to compensate nutrient loss. The presented protocol enables an investigation of adsorbent-bioprocess compatibility with high-throughput and limited effort.

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Count data modeling's significance and its applicability to real-world occurrences have been emphasized in a number of research studies. The purpose of this work is to introduce a new one-parameter discrete distribution for the modeling of count datasets. Some mathematical properties, such as reliability measures, characteristic function, moment-generating function, and associated measurements, such as mean, variance, skewness, kurtosis, and index of dispersion, have been derived and studied. The nature of the probability mass function and failure rate function has been studied graphically. The model parameter is estimated using renowned maximum likelihood estimation methods. A neutrosophic extension of the new model is also introduced for the modeling of interval datasets. In addition, the proposed distribution's applicability was compared to that of other discrete distributions. The study's findings show that the novel discrete distribution is a very appealing alternative to some other discrete competitive distributions.

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BACKGROUND: The fungal natural products illudin S and M have been investigated as precursors for the development of semisynthetic anticancer agents such as Irofulven (illudin S derivative) which is currently in phase II clinical trials. Recently, illudin M derivatives have shown improved in vitro selectivity towards cancer cells encouraging further investigation. This requires a stable supply of the precursor which is produced by Basidiomycota of the genus Omphalotus. We have recently reported a robust shake flask process for the production of gram quantities of illudin M from Omphalotus nidiformis aiming to transfer that process into stirred tank bioreactors, which can be used in a commercial production set-up. However, process transfer across different systems is not straightforward and particularly challenging when the producer is morphologically complex. There are only a few reports that address the development of bioprocesses for the production of compounds from Basidiomycota as these organisms have not been extensively studied because of their complex life cycles and often are difficult to cultivate under laboratory conditions. RESULTS: The recently developed shake flask process delivering stable titers of ~ 940 mg L-1 of illudin M was investigated using off-gas analysis to identify critical parameters which facilitated the transfer from shaken into stirred tank bioreactors. Comparable titers to the shake flask process were achieved in 2 L stirred tank bioreactors (1.5 L working volume) by controlling growth of biomass with a carefully timed pH-shift combined with an improved precursor-feeding strategy. A scale-up experiment in a 15 L bioreactor (10 L working volume), resembling the process at 1.5 L resulted in 523 mg L-1 and is the starting point for optimization of the identified parameters at that scale. CONCLUSION: By identifying and controlling key process parameters, the production process for illudin M was transferred from shake flasks into 2 L stirred tank bioreactors reaching a comparable titer (> 900 mg L-1), which is significantly higher than any previously reported. The insights obtained from 10 L scale pave the way towards further scale-up studies that will enable a sustainable supply of illudin M to support preclinical and clinical development programs.

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Since their first use in the 1930s, shake flasks have been a widely used bioreactor type for screening and process development due to a number of advantages. However, the limited gas-liquid mass transfer capacities-resulting from practical operation limits regarding shaking frequency and filling volumes-are a major drawback. The common way to increase the gas-liquid mass transfer in shake flasks with the implementation of baffles is generally not recommended as it comes along with several severe disadvantages. Thus, a new design principle for shaken bioreactors that aims for improving the gas-liquid mass transfer without losing the positive characteristics of unbaffled shake flasks is introduced. The flasks consist of cylindrical glass vessels with implemented perforated concentric ring walls. The ring walls improve the gas-liquid mass transfer via the formation of additional liquid films on both of its sides, whereas the perforations allow for mixing between the compartments. Sulfite oxidation experiments revealed over 200% higher maximum oxygen transfer capacities (OTRmax) compared to conventional shake flasks. In batch cultivations of Escherichia coli BL21 in mineral media, unlimited growth until glucose depletion and oxygen transfer rates (OTR) of up to 138 mmol/L/h instead of an oxygen limitation at 57 mmol/L/h as in normal shake flasks under comparable conditions could be achieved. Even overflow metabolism could be prevented due to sufficient oxygen supply without the use of unconventional shaking conditions or oxygen enrichment. Therefore, we believe that the new perforated ring flask principle has a high potential to considerably improve biotechnological screening and process development steps.

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Somatic hypermutation (SHM) of immunoglobulin (Ig) genes is a B cell specific process required for the generation of specific and high affinity antibodies during the maturation of the immune response against foreign antigens. This process depends on the activity of both activation-induced cytidine deaminase (AID) and several DNA repair factors. AID-dependent SHM creates the full spectrum of mutations in Ig variable (V) regions equally distributed at G/C and A/T bases. In most mammalian cells, deamination of deoxycytidine into uracil during S phase induces targeted G/C mutagenesis using either direct replication of uracils or TLS mediated bypass, however only the machinery of activated B lymphocytes can generate A/T mutagenesis around AID-created uracils. The molecular mechanism behind the latter remains incompletely understood to date. However, the lack of a cellular model that reproduces both G/C and A/T mutation spectra constitutes the major hurdle to elucidating it. The few available B cell lines used thus far to study Ig SHM indeed undergo mainly G/C mutations, that make them inappropriate or of limited use. In this report, we show that in the Ramos cell line that undergoes constitutive G/C-biased SHM in culture, the low rate of A/T mutations is due to an imbalance in the ubiquitination/deubiquitination reaction of PCNA, with the deubiquitination reaction being predominant. The inhibition of the deubiquitinase complex USP1-UAF1 or the expression of constitutive fusion of ubiquitin to PCNA provides the missing clue required for DNA polymerase η recruitment and thereafter the introduction of A/T base pair (bp) mutations during the process of IgV gene diversification. This study reports the establishment of the first modified human B cell line that recapitulates the mechanism of SHM of Ig genes in vitro.

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Resumo Analisa-se como a higiene mental se tornou pauta central no debate acerca do desenvolvimento nacional (anos 1920 e 1930), detendo-se nos trabalhos desenvolvidos pela Clínica de Eufrenia (1931), criada pela Liga Brasileira de Higiene Mental, e pelo Serviço de Ortofrenia e Higiene Mental (1934), edificado na reforma Anísio Teixeira, ambos situados na cidade do Rio de Janeiro. Ainda que com abordagens distintas, trabalharam a criança como lócus para a intervenção e construção do brasileiro do futuro: forte, saudável e produtivo. As análises foram alicerçadas em alguns casos clínicos realizados pelas instituições; nos escritos de seus diretores (os médicos Mirandolino Caldas e Arthur Ramos); e nos Arquivos Brasileiros de Higiene Mental.

Abstract This article analyzes how mental hygiene became a central theme in the debate on national development in the 1920s and 1930s, examining the work of the Clínica de Eufrenia (1931) created by the Liga Brasileira de Higiene Mental and the Serviço de Ortofrenia e Higiene Mental (1934), both in Rio de Janeiro and established during the Anísio Teixeira reform. Despite following different approaches, they considered the child as a locus to intervene and construct the Brazilian of the future: strong, healthy, and productive. This analysis includes some clinical cases from these institutions, writings by their directors (the physicians Mirandolino Caldas and Arthur Ramos), and the Brazilian Mental Hygiene Archives.

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Abstract Introduction Allergic rhinitis is a form of IgE mediated inflammation of the nasal mucosa in response to specific allergens, resulting in typical symptoms. Objectives This study was designed with the primary goal of comparing the clinical efficacy of posterior nasal neurectomy with or without pharyngeal neurectomy for the treatment of moderate-to-severe perennial allergic rhinitis. Secondary study aims included a comparison of the severity of comorbidities, including chronic cough and asthma, between patients in these two surgical treatment groups. Methods A total of 52 patients were enrolled in this randomized controlled trial and were assigned to either the control group (posterior nasal neurectomy) or the experimental group (posterior nasal neurectomy + pharyngeal neurectomy). The visual analog scale and rhinoconjunctivitis quality of life questionnaire were used to compare the differences in patient symptoms between baseline and 6-, 12-, and 24-months post-treatment. In addition, patient cough and asthma symptoms were monitored during follow-up via visual analog scale and asthma control test respectively. Results No significant differences in preoperative scores were evident between groups (p> 0.05). At 6-months post-treatment, there were significant differences in visual analog scale, rhinoconjunctivitis quality of life questionnaire, and asthma control test scores relative to baseline values in experimental group and control group patients (p< 0.05), and this remained true upon 12- and 24-month follow-up. No significant differences in visual analog scale, rhinoconjunctivitis quality of life questionnaire, or asthma control test scores were observed between the two treatment groups at any postoperative follow-up time point (p> 0.05), while coughing severity was found to be significantly reduced in the experimental group relative to the control group (p< 0.05). Conclusion posterior nasal neurectomy can be safely implemented with or without pharyngeal neurectomy in order to effectively treat allergic rhinitis. Combined posterior nasal neurectomy and pharyngeal neurectomy treatment may offer greater value than posterior nasal neurectomy alone for the treatment of allergic rhinitis patients with chronic cough.

Resumo Introdução A rinite alérgica é uma forma de inflamação da mucosa nasal mediada por IgE em resposta a alérgenos específicos, resulta em sintomas típicos. Objetivos Comparar a eficácia clínica da neurectomia nasal posterior com ou sem neurectomia faríngea para o tratamento da rinite alérgica perene de moderada a grave. Além disso, comparar a gravidade das comorbidades, inclusive tosse crônica e asma, entre os pacientes nesses dois grupos de tratamento cirúrgico. Método Foram incluidos neste ensaio clínico randomizado e designados para o grupo controle (neurectomia nasal posterior) ou para o grupo experimental (neurectomia nasal posterior + neurectomia faríngea) 52 pacientes. A escala visual analógica e o questionário de qualidade de vida na rinoconjuntivite (rhinoconjunctivitis quality of life questionnaire) foram usados para comparar as diferenças nos sintomas dos pacientes entre o período inicial e 6, 12 e 24 meses após o tratamento. Além disso, a tosse e os sintomas de asma dos pacientes foram monitorados durante o acompanhamento por meio da escala visual analógica e do teste de controle da asma (asthma control test ), respectivamente. Resultados Nenhuma diferença significante nos escores pré‐operatórios foi evidenciada entre os grupos (p > 0,05). Aos seis meses pós‐tratamento, houve diferenças significantes nos escores da escala visual analógica, no questionário de qualidade de vida na rinoconjuntivite e no teste de controle de asma em relação aos valores basais dos pacientes no grupo experimental e no grupo controle (p < 0,05), o que permaneceu verdadeiro após 12 e 24 meses de acompanhamento. Não foram observadas diferenças significantes nos escores da escala visual analógica e nem no questionário de qualidade de vida para conjuntivite ou no teste de controle da asma entre os dois grupos de tratamento em qualquer momento do acompanhamento pós‐operatório (p > 0,05), enquanto a gravidade da tosse foi significantemente reduzida no grupo experimental em relação ao grupo controle (p < 0,05). Conclusão A neurectomia nasal posterior pôde ser feita com segurança com ou sem neurectomia faríngea para o tratamento eficaz da rinite alérgica. O tratamento combinado com neurectomia nasal posterior e neurectomia faríngea pode oferecer mais benefício do que a neurectomia nasal posterior isolada para o tratamento de pacientes com rinite alérgica e tosse crônica.

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Whole genome sequencing of bovine breeds has allowed identification of genetic variants in milk protein genes. However, functional repercussion of such variants at a molecular level has seldom been investigated. Here, the results of a multistep Bioinformatic analysis for functional characterization of recently identified genetic variants in Brazilian Gyr and Guzerat breeds is described, including predicted effects on the following: (i) evolutionary conserved nucleotide positions/regions; (ii) protein function, stability, and interactions; (iii) splicing, branching, and miRNA binding sites; (iv) promoters and transcription factor binding sites; and (v) collocation with QTL. Seventy-one genetic variants were identified in the caseins (CSN1S1, CSN2, CSN1S2, and CSN3), LALBA, LGB, and LTF genes. Eleven potentially regulatory variants and two missense mutations were identified. LALBA Ile60Val was predicted to affect protein stability and flexibility, by reducing the number the disulfide bonds established. LTF Thr546Asn is predicted to generate steric clashes, which could mildly affect iron coordination. In addition, LALBA Ile60Val and LTF Thr546Asn affect exonic splicing enhancers and silencers. Consequently, both mutations have the potential of affecting immune response at individual level, not only in the mammary gland. Although laborious, this multistep procedure for classifying variants allowed the identification of potentially functional variants for milk protein genes.

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The Ames test is the most commonly used mutagenicity test worldwide. It is based on a microbial system that uses histidine auxotrophic Salmonella typhimurium strains. Due to either spontaneous mutations or mutations induced by a mutagenic compound, the cells can regain their ability to grow without histidine supplementation. The degree of mutagenicity of a sample correlates with the number of cells that are able to grow in media that lack histidine. All test variants published up to now are endpoint determinations providing no information about cell growth and respiration activity during the cultivation time. This study aimed to develop an alternative type of Ames test by characterizing the respiration activity of Salmonella typhimurium over time for dynamic mutagenicity detection. It focuses on elucidating the mechanisms underlying this novel test system, and serves as a general proof of principle. Respiration activity (oxygen transfer and uptake rate) and biomass growth of Salmonella typhimurium TA 100 and TA 98 were mechanistically modeled to understand and predict the behavior of the bacteria during the Ames test. The results simulated by the model were experimentally validated by the online monitoring of respiration activity over cultivation time using a Respiration Activity MOnitoring System (RAMOS). The simulated prediction was observed to fit well to the experimental data. When a mutagenic compound was added, its mutagenicity could be detected online due to the elevated cell number and respiration of histidine prototrophic cells. Laborious manual evaluation of mutagenicity after cultivation is not necessary. Mutagenicity evaluation with the presented alternative Ames RAMOS test fitted well to results from an Ames fluctuation test. In the future, a miniaturized RAMOS device for microtiter plates should allow for a high-throughput Ames RAMOS test.

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OBJECTIVE: Silymarin is a herbal extract containing flavonolignans, and it has inhibitory effects against the growth of different cancer cell lines by inducing apoptosis. Toll-like receptors are suggested as a novel and attractive target to treat cancer. The current study aimed at examining the mechanism of silymarin-induced apoptosis in Ramos cells and investigating its effects on TLR8 expression. MATERIALS AND METHODS: The half maximal inhibitory concentration (IC50) of silymarin in Ramos cells was determined via MTT viability test while the type of cell death was tested by annexin V/propidium iodide (PI) double staining method. The activity of caspase-3 and expression of TLR8 were measured in a time-dependent manner (in IC50) by colorimetric assay and real-time polymerase chain reaction (RT-PCR), respectively. RESULTS: The results of MTT showed that IC50 of silymarin in Ramos cells was 100 µg/ml after 48 hr treatment (p<0.01). Flow cytometry by annexin V/PI, showed that silymarin induced early/late apoptosis in this cell line (p<0.05 to p<0.01). In addition, the caspase-3 colorimetric method showed that caspase-3 increased in the Ramos cell line after treatment (p<0.01). This treatment led to a reduction in TLR8 mRNA expression in a time-dependent manner (p<0.01). CONCLUSION: The results indicated a new mechanism in the anticancer activity of Toll-like receptor (TLR) signaling after silymarin treatment in Ramos cancer cell line. This plant could be used to develop anticancer agents inhibiting TLRs.

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Resumo Em 1930, o engenheiro Theodoro Ramos publicou pela livraria francesa Albert Blanchard seu livro sobre cálculo vetorial. Este artigo apresenta o contexto de edição da obra, a casa editorial onde foi impressa, o mercado para livros desse tipo na França e o aparecimento desse tema na história do cálculo. Compara conteúdos e aspectos materiais de várias obras similares editadas no período com o livro aqui estudado de maneira a evidenciar tanto suas inovações quanto adesões em relação a esse corpus de livros. Além de seus aspectos intrinsecamente matemáticos, procura capturar as representações, às vezes conflituosas, que a obra recebeu em seu percurso entre São Paulo e a Europa, seja em meio a especialistas ou a seus alunos.

Abstract In 1930, the engineer Theodoro Ramos published a book on vector calculus through the French publisher Albert Blanchard. The context of is publication, the publishing house where it was printed, the market for books of this kind in France, and the appearance of this topic in the history of calculus are discussed. Some of the contents and material aspects of several works of a similar nature published in the same period are compared with a view to bringing to light the innovations and consistencies of the book in question vis-à-vis this bibliographic corpus. Alongside its intrinsically mathematical aspects, the sometimes conflicting representations of the book by specialists and students in its trajectory between São Paulo and Europe are observed.

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The Ames test is one of the most widely used mutagenicity tests. It employs histidine auxotrophic bacteria, which can mutate back to histidine prototrophy and, thus, grow on a histidine deficient medium. These mutants develop predominantly after adding a mutagenic compound during an initial growth phase on 1 mg/L histidine. In the established test systems, an endpoint determination is performed to determine the relative number of mutants. An alternative Ames test, the Ames RAMOS test, has been developed, which enables the online detection of mutagenicity by monitoring respiration activity. The reproducibility of the newly developed test system was investigated. A strong dependence of the test results on the inoculum volume transferred from the preculture was found. The more inoculum was needed to reach the required initial OD, the more mutagenic a positive control was evaluated. This effect was attributed to the histidine transfer from the preculture to the original Ames RAMOS test. The same problem is evident in the Ames fluctuation test. High reproducibility of the Ames RAMOS test could be achieved by performing the preculture on minimal medium with a defined histidine concentration and termination after histidine depletion. By using 5 mg/L initial histidine within the minimal medium, a higher separation efficiency between negative control and mutagenic samples could be achieved. This separation efficiency could be further increased by lowering the cultivation temperature from 37 to 30 °C, i.e. lowering the maximum growth rate. The optimized Ames RAMOS test was then transferred into a 48-well microtiter plate format (µRAMOS) for obtaining a high throughput test. The online detection of mutagenicity leads to a reduction of working time in the laboratory. Due to the optimization of reproducibility and the increase in separation efficiency, a sound mutagenicity evaluation, even of weak mutagenic compounds, can be achieved.

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