Lipolytic production from solid-state fermentation of the filamentous fungus Penicillium polonicum and its applicability as biocatalyst in the synthesis of ethyl oleate.

Lipases represent versatile biocatalysts extensively employed in transesterification reactions for ester production. Ethyl oleate holds significance in biodiesel production, serving as a sustainable alternative to petroleum-derived diesel. In this study, our goal was to prospect lipase and assess its efficacy as a biocatalyst for ethyl oleate synthesis. For quantitative analysis, a base medium supplemented with Rhodamine B, olive oil, and Tween 80 was used. Solid-state fermentation utilized crambe seeds of varying particle sizes and humidity levels as substrates. In the synthesis of ethyl oleate, molar ratios of 1:3, 1:6, and 1:9, along with a total enzymatic activity of 60 U in n-heptane, were utilized at temperatures of 30 °C, 37 °C, and 44 °C. Reactions were conducted in a shaker at 200 rpm for 60 min. As a result, we first identified Penicillium polonicum and employed the method of solid-state fermentation using crambe seeds as a substrate to produce lipase. Our findings revealed heightened lipolytic activity (22.5 Ug-1) after 96 h of fermentation using crambe cake as the substrate. Optimal results were achieved with crambe seeds at a granulometry of 0.6 mm and a fermentation medium humidity of 60%. Additionally, electron microscopy suggested the immobilization of lipase in the substrate, enabling enzyme reuse for up to 4 cycles with 100% enzymatic activity. Subsequently, we conducted applicability tests of biocatalysts for ethyl oleate synthesis, optimizing parameters such as the acid/alcohol molar ratio, temperature, and reaction time. We attained 100% conversion within 30 min at 37 °C, and our results indicated that the molar ratio proportion did not significantly influence the outcome. These findings provide a methodological alternative for the utilization of biocatalysts in ethyl oleate synthesis.

Occurrence of mosaic trisomy 22 and pericentric inversion of chromosome 9 in a patient with a good prognosis.

Complete trisomy 22 is a rare chromosomal condition that is incompatible with life. However, mosaic trisomy 22 usually has prolonged survival compatibility and may present a good prognosis depending on the tissues affected. Herein, we described a male patient with the occurrence of mosaic trisomy 22 associated with the inversion of chromosome 9, with karyotype 47, XY, inv (9) (p11q13), + 22 [5] / 46, XY, inv(9) (p11q13) [45] and arr 22q11.1 ~ q13.33(16,417008-51,219,009)x2 ~ 3. It is not possible to infer, in general, the clinical characteristics associated with mosaic trisomy 22. However, the patient presented common clinical features observed in reported cases (in parentheses the percentage observed comparing all reported cases): facial dysmorphia (100%), delay in motor development/growth (82%), cardiac abnormalities (73%), ear abnormalities (55%) and facial and/or body asymmetry (55%), in addition to hypotonia, skin spots, hypoplastic nails. Given the survival and quality of life associated with multidisciplinary treatment, it can be concluded that the patient has a good prognosis. Conclusively, we're presenting the occurrence of mosaic trisomy 22 and chromosome 9 inversion in the patient with favorable prognosis. Thus, this study proposed a guide which should be inserted in databases of rare genetic conditions to help genetic counselors define mosaic trisomy 22 diagnosis.

SHP-1 phosphatase acts as a coactivator of PCK1 transcription to control gluconeogenesis.

We previously reported that the protein-tyrosine phosphatase SHP-1 (PTPN6) negatively regulates insulin signaling, but its impact on hepatic glucose metabolism and systemic glucose control remains poorly understood. Here, we use co-immunoprecipitation assays, chromatin immunoprecipitation sequencing, in silico methods, and gluconeogenesis assay, and found a new mechanism whereby SHP-1 acts as a coactivator for transcription of the phosphoenolpyruvate carboxykinase 1 (PCK1) gene to increase liver gluconeogenesis. SHP-1 is recruited to the regulatory regions of the PCK1 gene and interacts with RNA polymerase II. The recruitment of SHP-1 to chromatin is dependent on its association with the transcription factor signal transducer and activator of transcription 5 (STAT5). Loss of SHP-1 as well as STAT5 decrease RNA polymerase II recruitment to the PCK1 promoter and consequently PCK1 mRNA levels leading to blunted gluconeogenesis. This work highlights a novel nuclear role of SHP-1 as a key transcriptional regulator of hepatic gluconeogenesis adding a new mechanism to the repertoire of SHP-1 functions in metabolic control.

HSP70 mediates a crosstalk between the estrogen and the heat shock response pathways.

Cells respond to multiple signals from the environment simultaneously, which often creates crosstalk between pathways affecting the capacity to adapt to the changing environment. Chaperones are an important component in the cellular integration of multiple responses to environmental signals, often implicated in negative feedback and inactivation mechanisms. These mechanisms include the stabilization of steroid hormone nuclear receptors in the cytoplasm in the absence of their ligand. Here, we show using immunofluorescence, chromatin immunoprecipitation, and nascent transcripts production that the heat shock protein 70 (HSP70) chaperone plays a central role in a new crosstalk mechanism between the steroid and heat shock response pathways. HSP70-dependent feedback mechanisms are required to inactivate the heat shock factor 1 (HSF1) after activation. Interestingly, a steroid stimulation leads to faster accumulation of HSF1 in inactive foci following heat shock. Our results further show that in the presence of estrogen, HSP70 accumulates at HSF1-regulated noncoding regions, leading to deactivation of HSF1 and the abrogation of the heat shock transcriptional response. Using an HSP70 inhibitor, we demonstrate that the crosstalk between both pathways is dependent on the chaperone activity. These results suggest that HSP70 availability is a key determinant in the transcriptional integration of multiple external signals. Overall, these results offer a better understanding of the crosstalk between the heat shock and steroid responses, which are salient in neurodegenerative disorders and cancers.

Aspergillus sclerotiorum lipolytic activity and its application in bioremediation of high-fat dairy wastewater environments.

Oils and grease (O&G) have low affinity for water and represent a class of pollutants present in the dairy industry. Enzyme-mediated bioremediation using biocatalysts, such as lipases, has shown promising potential in biotechnology, as they are versatile catalysts with high enantioselectivity and regioselectivity and easy availability, being considered a clean technology (white biotechnology). Specially in the treatment of effluents from dairy industries, these enzymes are of particular importance as they specifically hydrolyze O&G. In this context, the objective of this work is to prospect filamentous fungi with the ability to synthesize lipases for application in a high-fat dairy wastewater environment. We identified and characterized the fungal species Aspergillus sclerotiorum as a good lipase producer. Specifically, we observed highest lipolytic activity (20.72 U g-1) after 96 h of fermentation using sunflower seed as substrate. The fungal solid fermented was used in the bioremediation in dairy effluent to reduce O&G. The experiment was done in kinetic from 24 to 168 h and reduced over 90% of the O&G present in the sample after 168 h. Collectively, our work demonstrated the efficiency and applicability of fungal fermented solids in bioremediation and how this process can contribute to a more sustainable wastewater pretreatment, reducing the generation of effluents produced by dairy industries.

Yeasts isolated from a lotic continental environment in Brazil show potential to produce amylase, cellulase and protease.

Yeasts have wide applicability in the industrial field, as in the production of enzymes used in biocatalysts. Biocatalysts are more efficient when compared to chemical catalysts, with emphasis on hydrolytic enzymes, such as amylase, cellulase and protease. Here we focused on prospecting yeasts, with a high capacity to synthesize hydrolytic enzymes, from a continental lotic ecosystem environment in Brazil. 75 yeasts were grown in Yeast Extract-Peptone-Dextrose (YPD) medium supplemented with antibacterial and their capacity for enzymatic production was tested in specific media. Accordingly, 64 yeasts showed enzyme production capacity. From those, six showed good enzyme indexes, 3 for amylase, 2 for cellulase and 1 for protease. All showed at least one hydrolytic enzyme activity for the tested enzymes (amylase, cellulase and protease), which suggested that the yeasts are metabolically active. By sequencing the 26S gene, we identified Naganishia diffluens and Apiotrichum mycotoxinivorans as the species with highest enzyme production activities. Those species showed potential for application as biological catalysts in the biotechnological scope, collaborating in a sustainable way for the development of industrial products.

Control of adipogenic commitment by a STAT3-VSTM2A axis.

White adipose tissue (WAT) is a dynamic organ that plays crucial roles in controlling metabolic homeostasis. During development and periods of energy excess, adipose progenitors are recruited and differentiate into adipocytes to promote lipid storage capability. The identity of adipose progenitors and the signals that promote their recruitment are still incompletely characterized. We have recently identified V-set and transmembrane domain-containing protein 2A (VSTM2A) as a novel protein enriched in preadipocytes that amplifies adipogenic commitment. Despite the emerging role of VSTM2A in promoting adipogenesis, the molecular mechanisms regulating Vstm2a expression in preadipocytes are still unknown. To define the molecular mechanisms controlling Vstm2a expression, we have treated preadipocytes with an array of compounds capable of modulating established regulators of adipogenesis. Here, we report that Vstm2a expression is positively regulated by PI3K/mTOR and cAMP-dependent signaling pathways and repressed by the MAPK pathway and the glucocorticoid receptor. By integrating the impact of all the molecules tested, we identified signal transducer and activator of transcription 3 (STAT3) as a novel downstream transcription factor affecting Vstm2a expression. We show that activation of STAT3 increased Vstm2a expression, whereas its inhibition repressed this process. In mice, we found that STAT3 phosphorylation is elevated in the early phases of WAT development, an effect that strongly associates with Vstm2a expression. Our findings identify STAT3 as a key transcription factor regulating Vstm2a expression in preadipocytes.NEW & NOTEWORTHY cAMP-dependent and PI3K-mTOR signaling pathways promote the expression of Vstm2a. STAT3 is a key transcription factor that controls Vstm2a expression in preadipocytes. STAT3 is activated in the early phases of WAT development, an effect that strongly associates with Vstm2a expression.

Increased DNA Damage, Instability and Cytokinesis Defects in Occupationally Exposed Car Painters.

BACKGROUND/AIM: Car painting is considered an occupational exposure job with high risk for cancer development, due to the association with harmful chemicals and mutagens. This study aimed to profile car painters occupationally exposed and determine its association with DNA damage and genomic instability. MATERIALS AND METHODS: We collected a questionnaire and buccal cells of 74 individuals (37 car painters and 37 non-exposed workers) paired by age, alcohol and smoking habits. The number of pyknotic cells, karyolitic cells, karyorrhetic cells, condensed chromatin, binucleated cells, basal cells, differentiated cells (DIFF), micronucleated cells and nuclear buds were evaluated using the Buccal Micronucleus Cytome Assay protocol. RESULTS: A statistically significant increase was observed in all parameters (p<0.05) in the exposed group, but DIFF showed a statistically significant decrease (p<0.001), compared to the control group. CONCLUSION: In association with the poor work environment and lack of personal and collective protective equipment, occupational exposure of car painters leads to high DNA damage, genomic instability and alterations in cellular kinetics.

BCL2 and miR-181a transcriptional alterations in umbilical-cord blood cells can be putative biomarkers for obesity.

Several findings suggest that in utero stressor stimuli can alter fetal development by promoting transcriptional changes, and predisposing the neonate to diseases later in life. This study aimed to investigate whether a hyperglycemic environment in pregnant women with gestational diabetes mellitus (GDM) is able to cause fetal genetic alterations and predispose neonates to obesity. Transcriptional alteration of SIRT1, TP53 and BCL2 genes, miR-181a (a SIRT1 or BCL2 regulator) and telomere length were evaluated in placental and umbilical-cord blood cells. Healthy (HP; n = 20) and GDM (n = 20) pregnant women and their respective neonates were included in the study. Additionally, obese (n = 20) and eutrophic (n = 20) adults also participated as reference populations. Gene expression data showed down-regulation of BCL2 in umbilical-cord and peripheral blood cells from GDM neonates and obese adults, respectively. The miR-181a was down-regulated only in umbilical-cord blood cells of GDM neonates. Telomere length presented no significant difference. In conclusion, our study demonstrated that the GDM hyperglycemic intrauterine environment promotes transcriptional alterations in BCL2 and miR-181a in neonate umbilical-cord blood cells. Furthermore, both GDM neonates and obese subjects share the same transcriptional alteration in BCL2. Considering the relationship between obesity development and the functions regulated by these two genes, BCL2 and miR-181a could be adopted as potential biomarkers for childhood obesity. However, further study designs are recommended to confirm this hypothesis.

Direct and Indirect Anthropogenic Contamination in Water Sources: Evaluation of Chromosomal Stability and Cytotoxicity Using the Allium cepa Test.

Increasing industrialization and urbanization has deteriorated water quality around the world. Nowadays, evaluation of the effects of chemical compounds using bioassays is a critical step in the hazard identification assessment. Thus, this work aimed to determine the genetic damage caused by different types of anthropogenic contamination in a river´s water in Brazil. Two points (P1 and P2) and two periods (referred as direct and indirect anthropogenic contamination) were evaluated in Allium cepa roots. MI was increased (p < 0.05) in both points in the indirect anthropogenic contamination and decreased in the indirect anthropogenic contamination periods. Moreover, parameters as DNA instability (CA and MN) were observed in both periods indicating substances in the water with mutagenic, genotoxic, and cytotoxic potential. Interestingly, a 20-fold increase in CA frequencies were observed in P1 and P2 in the second collection period (direct anthropogenic contamination) (p < 0.05). In conclusion, our data indicated that anthropogenic activities in the area contributed to contaminate this water source. Moreover, direct anthropogenic contamination maximized the damage, posing a possible hazard to population health.

Mutagenicity of two herbicides widely used on soybean crops by the Allium cepa test.

This study evaluated the mutagenic effects of two herbicides: Clorimurom Nortox(®) and Imazaquim Ultra Nortox(®) widely used on soybean crops in Brazil. As a test system, Allium cepa assay was used, which analyzes the frequency of micronuclei (MN), chromosomal aberrations (CA) and the mitotic index (MI). Four concentrations of each herbicide (50, 75, 100 and 125 %) were tested in triplicate using distilled water (negative control) and methyl methanesulfonate (positive control) as controls. Three experimental repetitions were realized. Clorimurom Nortox(®) showed a significantly lower MI than the negative control for the concentrations of 75, 100 and 125 %, but the CA was significantly increased at all concentrations. There was no recovery for CA or MI. The 125 % concentration of Imazaquim Ultra Nortox(®) was cytotoxic and also exerted an effect on the other parameters. The concentration of 100 % showed a statistically increased MN and there was no recovery, while the 75 % concentration significantly affected CA, with recovery observed. The two herbicides showed mutagenic damage in Allium cepa cells, which implies a careful handling of these products, to minimize the risk of human and environmental contamination.