Research progress of transcriptomics and proteomics in schizophrenia / 中华预防医学杂志

Schizophrenia is a severe psychiatric disorder with an unclear etiology and various clinical manifestations. The diagnosis and consequent treatment of schizophrenia mainly rely on clinical symptoms. Multiple risk sites associated with schizophrenia have been identified, yet objective indicators have not been found to facilitate clinical diagnosis and treatment of schizophrenia. The development of omics technology provides different perspectives on the etiology of schizophrenia and make the early identification, diagnosis and treatment of the disorder possible. This article summarizes the prevalence of schizophrenia, reviews the research results and shortcomings of transcriptomics and proteomics, as well as the latest achievements and prospects of multi-omics, aiming to reveal the use of omics in SZ, provide more comprehensive biological evidence to reveal the complex pathogenesis of schizophrenia and provide a theoretical basis for the early identification, accurate diagnosis, disease progression control, and prognosis improvement of schizophrenia.

Research progress of transcriptomics and proteomics in schizophrenia / 中华预防医学杂志

Schizophrenia is a severe psychiatric disorder with an unclear etiology and various clinical manifestations. The diagnosis and consequent treatment of schizophrenia mainly rely on clinical symptoms. Multiple risk sites associated with schizophrenia have been identified, yet objective indicators have not been found to facilitate clinical diagnosis and treatment of schizophrenia. The development of omics technology provides different perspectives on the etiology of schizophrenia and make the early identification, diagnosis and treatment of the disorder possible. This article summarizes the prevalence of schizophrenia, reviews the research results and shortcomings of transcriptomics and proteomics, as well as the latest achievements and prospects of multi-omics, aiming to reveal the use of omics in SZ, provide more comprehensive biological evidence to reveal the complex pathogenesis of schizophrenia and provide a theoretical basis for the early identification, accurate diagnosis, disease progression control, and prognosis improvement of schizophrenia.

Proteomic Difference Analysis of Whole Blood and Bloodstains / 法医学杂志

OBJECTIVES@#To study the changes of protein levels in peripheral blood after it dried.@*METHODS@#The proteins from whole blood and bloodstains were detected by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and normalized by the label-free quantification (LFQ) method. The differential proteins were analyzed by using R 4.2.1 software, limma and edgeR package. The analysis of biological function, signaling pathway and subcellular localization for the differential proteins was then performed.@*RESULTS@#A total of 623 and 596 proteins were detected in whole blood and bloodstains, respectively, of which 31 were statistically significant in the quantitative results, including 10 up-regulated and 21 down-regulated proteins in bloodstains.@*CONCLUSIONS@#The protein abundances in whole blood and bloodstains are highly correlated, and the variation of protein abundances may be related to the changes of endogenous and structural proteins in cells. The application of proteomics technology can assist the screening and identification of protein biomarkers, thereby introducing new biomarkers for forensic research.

Advances of peptide-centric data-independent acquisition analysis algorithms and software tools / 生物工程学报

Data-independent acquisition (DIA) is a high-throughput, unbiased mass spectrometry data acquisition method which has good quantitative reproducibility and is friendly to low-abundance proteins. It becomes the preferred choice for clinical proteomic studies especially for large cohort studies in recent years. The mass-spectrometry (MS)/MS spectra generated by DIA is usually heavily mixed with fragment ion information of multiple peptides, which makes the protein identification and quantification more difficult. Currently, DIA data analysis methods fall into two main categories, namely peptide-centric and spectrum-centric. The peptide-centric strategy is more sensitive for identification and more accurate for quantification. Thus, it has become the mainstream strategy for DIA data analysis, which includes four key steps: building a spectral library, extracting ion chromatogram, feature scoring and statistical quality control. This work reviews the peptide-centric DIA data analysis procedure, introduces the corresponding algorithms and software tools, and summarizes the improvements for the existing algorithms. Finally, the future development directions are discussed.

Advances of chromatogram retention time alignment algorithms in proteomics / 生物工程学报

Chromatography is a basic process in the current proteomics workflow, and the retention time alignment of the chromatogram is one of the important steps to effectively improve the identification and quantification accuracy. After years of development, a series of algorithms for retention time alignment have been developed. This review summarizes the advances of chromatographic retention time alignment algorithms and tools for proteomics analysis from the perspective of proteomics users, and discusses the development and future application directions.

Quantitative Proteomics Using Isobaric Labeling: A Practical Guide / 基因组蛋白质组与生物信息学报·英文版

In the past decade, relative proteomic quantification using isobaric labeling technology has developed into a key tool for comparing the expression of proteins in biological samples. Although its multiplexing capacity and flexibility make this a valuable technology for addressing various biological questions, its quantitative accuracy and precision still pose significant challenges to the reliability of its quantification results. Here, we give a detailed overview of the different kinds of isobaric mass tags and the advantages and disadvantages of the isobaric labeling method. We also discuss which precautions should be taken at each step of the isobaric labeling workflow, to obtain reliable quantification results in large-scale quantitative proteomics experiments. In the last section, we discuss the broad applications of the isobaric labeling technology in biological and clinical studies, with an emphasis on thermal proteome profiling and proteogenomics.

Comparison of protein precipitation methods for sample preparation prior to proteomic analysis of Chinese hamster ovary cell homogenates

BACKGROUND: Chinese hamster ovary (CHO) cells are the workhorse for obtaining recombinant proteins. Proteomic studies of these cells intend to understand cell biology and obtain more productive and robust cell lines for therapeutic protein production in the pharmaceutical industry. Because of the great importance of precipitation methods for the processing of samples in proteomics, the acetone, methanol-chloroform (M/C), and trichloroacetic acid (TCA)-acetone protocols were compared for CHO cells in terms of protein recovery, band pattern resolution, and presence on SDS-PAGE. RESULTS: Higher recovery and similar band profile with cellular homogenates were obtained using acetone precipitation with ultrasonic bath cycles (104.18 ± 2.67%) or NaOH addition (103.12 ± 5.74%), compared to the other two protocols tested. TCA-acetone precipitates were difficult to solubilize, which negatively influenced recovery percentage (77.91 ± 8.79%) and band presence. M/C with ultrasonic homogenization showed an intermediate recovery between the other two protocols (94.22 ± 4.86%) without affecting protein pattern on SDS-PAGE. These precipitation methods affected the recovery of low MW proteins (< 15 kDa). CONCLUSIONS: These results help in the processing of samples of CHO cells for their proteomic study by means of an easily accessible, fast protocol, with an almost complete recovery of cellular proteins and the capture of the original complexity of the cellular composition. Acetone protocol could be incorporated to sample-preparation workflows in a straightforward manner and can probably be applied to other mammalian cell lines as well.

Protein structure of the venom in nine species of snake: from bio-compounds to possible healing agents

Due to its various structures in bio-compounds, snake venom is the indisputable result of evolutionary stages of molecules with an increasingly complex structure, high specificity, and of great importance for medicine because of their potential. The present study proposed an underpinning examination of venom composition from nine species of venomous snakes using a useful and replicable methodology. The objective was the extension of the evaluation of protein fractions in the field up to 230 kDa to permit possible identification of some fractions that are insufficiently studied. The gel capillary electrophoresis method on the chip was performed using an Agilent 2100 bioassay with the 80 and 230-LabChip Protein kits. Interpretation of electrophoresis was performed using the Protein 2100 expert (Agilent) test software as follows: a) Protein 80 (peak size scale): 1.60, 3.5, 6.50, 15.00, 28.00, 46.00, 63.00, 95.00 kDa; b) Protein 230 (peak size scale): 4.50, 7.00, 15.00, 28.00, 46.00, 63.00, 95.00, 150.00, 240.00 kDa. The screening revealed the presence of compounds with a molecular weight greater than 80 kDa, in the case of Vipera aspis and Vipera xantina palestinae. For V. aspis, a 125 kDa molecular weight pro-coagulant protein was identified, known as being involved in the reduction of plasma clotting time without any direct activity in the fibrinogen coagulation process. The samples examined on the Protein 230-LabChip electrophoresis chip can be considered as a novelty with possible uses in medicine, requiring further approaches by advanced proteomics techniques to confirm the intimate structural features and biological properties of snake venoms.

Aspectos generales del perfil proteómico del Echinococcus granulosus / General aspects of the proteomic profile of Echinococcus granulosus

La información disponible referente a las características proteómicas del E. granulosus es escasa (no supera los 50 estudios publicados); y nos parece que la identificación proteómica, podría mejorar la comprensión de algunas características bioquímicas e inmunológicas de la Equinococosis Quística (EQ). De tal modo que el proteoma de E. granulosus aún no está bien descrito. Sólo existen reportes de algunas secuencias de proteínas. El objetivo de este manuscrito fue comentar algunos aspectos de la evidencia existente respecto de los estudios del perfil proteómico del E. granulosus. Se recomienda el estudio de al menos el quiste y su pared, el líquido hidatídico y la víscera del hospedero. Para ello, existen metodologías que han sido empleadas para estudiar las características proteómicas de la EQ. Entre ellas, destacan SDS-PAGE, electroforesis bidimensional combinada con Western Blot, inmunoanálisis, y espectrometría de masas mediante técnicas MALDI-TOF. Se han identificado una serie de proteínas en muestras de EQ. Algunas de ellas, asociadas a procesos inmunológicos, de gluconeogénesis, glucogenolisis y glucogénesis. Por otra parte, se ha documentado la liberación de exosomas al líquido hidatídico por parte de los protoescólex y la capa germinativa; estructuras en las que se han identificado factores de virulencia asociados con la supervivencia del quiste. No obstante lo anteriormente señalado, se requiere de múltiples estudios exhaustivos en la materia para comprender mejor la caracterización perfil proteómico del E. granulosus.

The information available regarding the proteomic characteristics of E. granulosus is scarce; and it seems that the proteomic identification could improve the understanding of some biochemical and immunological characteristics of cystic echinococcosis (CE). So, the proteome of E. granulosus is still not well described yet. There are only reports of some protein sequences. The objective of this manuscript was to comment on some aspects of the existing evidence regarding studies of the proteomic profile of E. granulosus. The study of at least the cyst and its wall, the hydatid fluid and the viscera of the host are recommended. There are a series of methodologies that have been used to study the proteomic characteristics of EQ. These include SDS-PAGE, bidimensional electrophoresis combined with Western Blot, immunoassay, and mass spectrometry using MALDI-TOF techniques. A series of proteins have been identified in CE samples. Some of them, associated with immune response, gluconeogenesis, glycogenolysis and glycogenesis. On the other hand, release of exosomes to the hydatid fluid by protoescolex and germinative layer has been documented (associated virulence factors have been identified in these structures). Notwithstanding the foregoing, it requires multiple exhaustive studies in the field to better understand the characterization of the proteomic profile of E. granulosus.

Diagnostic potential of saliva proteome analysis: a review and guide to clinical practice

Abstract: Proteomic techniques have become popular in medicine and dentistry because of their widespread use in analyzing bodily fluids such as blood, saliva, urine, and gingival crevicular fluids as well as hard tissues such as enamel, dentine, and cementum. This review is a guide to proteomic techniques in general dentistry, summarizing techniques and their clinical application in understanding and diagnosing diseases and their use in identifying biomarkers of various diseases.

Screening on serum biomarkers in idiopathic pulmonary fibrosis patients by proteomics technology / 中华流行病学杂志

Objective: Applying the proteomics technology to identify proteins differentially in serums of idiopathic pulmonary fibrosis patients and normal population. Methods: The study included serum samples from idiopathic pulmonary fibrosis group and normal controlled group with 30 cases of each, from the Second Hospital of Shandong University, between October 2014 and October 2015. Proteins differentially expressed in serums were quantified by the isobaric tags for relative and absolute quantization coupled with two-dimensional liquid chromatography/tandem mass spectrometry technology. The proteins were analyzed in terms of molecular function, cell location and biological processes for showing the key protein molecules which were related to the development of idiopathic pulmonary fibrosis. Results: A total of 490 kinds of proteins (with confidence coefficient above 95%) were identified by mass spectrometry and 25 kinds of differentially expressed proteins were found. Compared with the control group, we found 4 types of up-regulated proteins and 21 down-regulated ones in the serum of idiopathic pulmonary fibrosis patients. Data from the Gene ontology analysis showed that most differentially expressed proteins were in the extracellular region (92%) while pathway enrichment analysis showed that most proteins were involved in the complement and coagulation cascade pathway. Conclusion: Proteins related to the complement system coagulation cascade pathway, and the proteins function need to be further studied.

Papel das vesículas extracelulares secretadas por adenocarcinomas gástricos na resposta ao tratamento quimioterápico / Role of extracellular vesicles secreted by gastric adenocarcinomas in response to chemotherapy

O adenocarcinoma gástrico (AdG) é a terceira causa mais comum de morte por câncer no mundo e um dos tumores com maior índice de mortalidade no Brasil. Estes tumores aparecem em terceiro lugar na incidência entre homens e em quinto nas mulheres. A quimioterapia neoadjuvante (QT) com 5-fluorouracil (5-FU) pode melhorar a ressecabilidade e a sobrevida dos pacientes com AdG, porém sua eficácia está limitada pela resistência à droga. Apenas pacientes que respondem a esta terapia com toxicidade tolerável são potencialmente beneficiados, entretanto não é possível identificar e separar clinicamente estes indivíduos. Assim, identificar marcadores preditivos de resposta para selecionar os pacientes que se beneficiariam deste tratamento é relevante. Vesículas extracelulares (VEs) são componentes secretados pelas células incluindo células tumorais, cujo conteúdo é regulado e composto por moléculas que podem ter uma miríade de funções locais e a distância. Muitas destas moléculas podem ser também usadas como biomarcadores séricos por conterem informações importantes sobre o tumor. Desta forma, este trabalho tem como objetivo identificar marcadores de resistência a 5-FU em VEs secretadas por linhagens celulares humanas de câncer gástrico e avaliar o papel das VEs na quimioresistência. Para tanto, foram geradas células de AdG derivadas da linhagem AGS, resistentes a 5-FU (rAGS_FU) de onde foram isoladas, quantificadas e caracterizadas VEs. Células rAGS-FU secretam cerca de 2 vezes mais VEs que as células parentais (AGS), entretanto a distribuição destas por tamanho é semelhante. As células rAGS_FU apresentaram maior proliferação, capacidade de formação de colônias e invasão que as células AGS. Interessantemente, as VEs provenientes de células resistentes a 5-FU, rAGS_FU, são capazes de transferir à células AGS os fenótipos de resistência ao quimioterápico bem como um aumento na capacidade de formação de colônias e invasão. As células AGS que se tornam resistentes ao tratamento não têm este fenótipo revertido pela remoção das VEs da células resistentes nem pelo tratamento com VEs de células AGS parental, indicando que o fenótipo de resistência adquirido após o tratamento é irreversível, pelo menos pelo período estudado. O conteúdo proteico das VEs das células AGS e rAGS_FU e suas respectivas VEs foi comparado por proteômica. Nessa abordagem foram identificadas 1.915 proteínas nas células e 1.638 proteínas em VEs das quais 309 proteínas eram diferencialmente expressas em células e 66 em VEs. Entre as proteínas com expressão diferencial entre as duas células e também nas suas respectivas VEs, selecionamos e validamos por western blotting a proteina fascina. Esta parece ser um potencial candidato a biomarcador de resistência a 5-FU uma vez que sua expressão é indetectável na célula AGC e suas VEs e altamente expressa nas células rAGS_FU e suas vesículas. A fascina é uma proteína do citoesqueleto com um papel chave nas interações célula-célula, adesão e motilidade celulares e é associada a agressividade tumoral. Estes dados apontam o papel das VEs nos mecanismos relacionados à resistência a 5-FU em células de AdG e sugerem que fascina possa estar associada ao mecanismo de resistência a este quimioterápico e que também seja um potencial biomarcador deste fenótipo

Gastric adenocarcinoma (GAd) is the third most common cause of cancer related death worldwide, and one of the tumors with the highest mortality rates in Brazil. In men, this cancer ranks the third most common cancer, while in women, it ranks fifth. 5-fluorouracil (5-FU) based neoadjuvant chemotherapy can improve tumor resectability and patient's survival rates. Its effectiveness however, is limited by drug resistance. Thus, an effort to identify predictive markers of response to neoadjuvant therapy and select patients who could benefit from this treatment is relevant. One such approach could be to use contents of extracellular vesicles (EVs). EVs are components secreted by cells including tumor cells, whose contents are composed of molecules that can have a myriad of local and distance functions and many of these molecules can also be used as serum biomarkers since they contain important information about the tumor. This work aims to identify 5-FU resistance markers in EVs secreted by human gastric cancer cell line and to evaluate the role of EVs in chemoresistance. GAd cells resistant to 5-FU (rAGS_FU) were generated from the AGS cell line and EVs secreted by rAGS_FU cells and parental AGS cells were isolated, quantified and characterized. Our results showed that AGS_FU cells secrete about 2 times more EVs than parental AGS cells, however their size distribution is similar. The resistant rAGS_FU cells showed higher proliferation rates, capacity of colony formation and invasion properties. Interestingly, EVs from 5-FU resistant cells, rAGS_FU, are able to transfer to the AGS cells the phenotypes of resistance to chemotherapy as well as an increase in the capacity of colony formation and invasion. AGS cells that become resistant to treatment do not have this phenotype reversed by removal of the EVs from the resistant cells or by treatment with EVs from parental AGS cells, indicating that the resistance phenotype acquired after treatment is irreversible, at least for the period studied. The protein content of the AGS and rAGS_FU cells and their respective EVs was compared by proteomics. In this approach, 1,915 proteins were identified in cells and 1,638 proteins in EVs of which 309 proteins were differentially expressed in cells and 66 in EVs. Among the proteins with differential expression between the two cells and also in their respective EVs, we selected and validated by western blotting the protein fascin. Fascin protein appears to be a potential candidate for biomarker of 5-FU resistance since its expression is undetectable in AGS cells and their EVs and highly expressed in rAGS_FU cells and their vesicles. Fascin is a cytoskeletal protein with a key role in cellcell interactions, cell adhesion and motility and is associated with tumor aggressiveness. These data point to the role of EVs in the mechanisms related to 5-FU resistance in GAd cells and suggest that fascin may be associated with the mechanism of resistance to this chemotherapeutic agent and that it is also a potential biomarker of this phenotype

Missing and overexpressing proteins in domestic cat oocytes following vitrification and in vitro maturation as revealed by proteomic analysis

BACKGROUND: The domestic cat serves as an animal model for assisted reproductive studies of endangered felid species. To date, there are no data on the protein alterations following cryopreservation of oocytes in felid family. METHODS: Immature (germinal vesicle) domestic cat oocytes were vitrified in the vitrification solution containing 35% ethylene glycol, 20% DMSO and 0.5 mM sucrose. The vitrified-warmed oocytes were matured (metaphase II) in vitro and subjected to proteomic analysis using 1DE SDS-PAGE prefractionation combined with LC-MS/MS. RESULTS: A total of 1712 proteins were identified in in vitro matured oocytes. Of the 1712 proteins, 1454 proteins were found in both groups, whereas, 258 proteins were differentially expressed between control and vitrified-warmed groups. In vitrified-warmed oocytes, the missing proteins were membrane and nuclear proteins; whereas, apoptosis and DNA repair proteins were overrepresented. CONCLUSIONS: The identified missing and overexpressed proteins in vitrified-warmed oocytes represent potential markers of cryoinjuries and the developmental pathways of oocytes. The findings of differential expressed proteins may contribute to effective ways of proteome analysis of oocyte/embryo quality in order to assess safety of cryopreservation in felid species.

Standardization of a protocol for shotgun proteomic analysis of saliva

Abstract Saliva contains numerous proteins and peptides, each of them carries a number of biological functions that are very important in maintaining the oral cavity health and also yields information about both local and systemic diseases. Currently, proteomic analysis is the basis for large-scale identification of these proteins and discovery of new biomarkers for distinct diseases. Objective This study compared methodologies to extract salivary proteins for proteomic analysis. Material and Methods Saliva samples were collected from 10 healthy volunteers. In the first test, the necessity for using an albumin and IgG depletion column was evaluated, employing pooled samples from the 10 volunteers. In the second test, the analysis of the pooled samples was compared with individual analysis of one sample. Salivary proteins were extracted and processed for analysis by LC-ESI-MS/MS. Results In the first test, we identified only 35 proteins using the albumin and IgG depletion column, while we identified 248 proteins without using the column. In the second test, the pooled sample identified 212 proteins, such as carbonic anhydrase 6, cystatin isoforms, histatins 1 and 3, lysozyme C, mucin 7, protein S100A8 and S100A9, and statherin, while individual analysis identified 239 proteins, among which are carbonic anhydrase 6, cystatin isoforms, histatin 1 and 3, lactotransferrin, lyzozyme C, mucin 7, protein S100A8 and S100A9, serotransferrin, and statherin. Conclusions The standardization of protocol for salivary proteomic analysis was satisfactory, since the identification detected typical salivary proteins, among others. The results indicate that using the column for depletion of albumin and IgG is not necessary and that performing individual analysis of saliva samples is possible.

Análisis proteómico de los productos de excreción-secreción de cuatro aislados de Trichinella spiralis obtenidos de hospederos accidentales / Proteomic analysis of the excretion-secretion products of four Trichinella spiralis isolates obtained from accidental hosts

Resumen: Introducción: Trichinella spiralis es un nemátodo tisular que se aloja en el músculo esquelético de humanos y otros mamíferos y causa una serie de alteraciones fisiológicas. Las proteínas de los productos de excreción-secreción de T. spiralis juegan un papel importante en la aparición y regulación de estas alteraciones. Sin embargo, aún no se conoce el efecto de estos productos en la infección e invasión del parásito al hospedero. Métodos: Mediante un análisis electroforético en una dimensión, Western blot y espectrometría de masas, se evaluaron las diferencias y similitudes entre proteínas antigénicas y de superficie de cuatro aislados de T. spiralis obtenidos de hospederos accidentales (perros) y la cepa de referencia aislada de cerdos (MSUS/MEX/91/CM). Resultados: Utilizando ontología de genes, se encontraron cinco proteínas exclusivas de los hospederos accidentales. Después del análisis, se encontró que estas proteínas forman parte de la matriz extracelular del parásito, cuentan con actividad catalítica y están implicadas en la adhesión a las células del hospedero. La actividad antigénica de las cuatro cepas aisladas de hospederos accidentales es idéntica a la reportada para T. spiralis, visualizándose el triplete antigénico característico de 43, 45 y 47 kDa. Conclusiones: Las proteínas exclusivas de los hospederos accidentales proveen información para entender el mecanismo de acción de este parásito para penetrar el músculo y evadir la respuesta inmune en el hospedero.

Abstract: Background: Trichinella spiralis is an intestinal and tissue nematode specific for mammalian skeletal muscle, causing a series of physiological alterations. T. spiralis excretory-secretion products play an important role in the appearance and regulation of these alterations. However, the effect of these products on the infection and invasion of the parasite to the host is unknown. Methods: Differences and similarities between antigenic proteins and surface proteins of four accidental hosts isolates (dogs) of T. spiralis and the reference strain isolated from pigs (MSUS/MEX/91/CM) were assessed by electrophoresis, western blot and mass spectrometry. Results: Using gene ontology, five proteins exclusive to the accidental hosts were analyzed. The results showed that these proteins are part of the extracellular matrix of the parasite, present catalytic activity, and bind to host cells. The antigenic activity the four strains showed the antigenic triplet characteristic of T. spiralis of 43, 45 and 47 kDa. Conclusions: Five proteins exclusive to dog isolates provided information to understand the mechanism of action of this parasite to penetrate the muscle and evade the immune response in the host.

Mitochondrial proteomic profile of complex IV deficiency fibroblasts: rearrangement of oxidative phosphorylation complex/supercomplex and other metabolic pathways / Perfil proteómico mitocondrial de los fibroblastos con deficiencia del complejo IV: reordenamiento del complejo/supercomplejo de la fosforilación oxidativa y otras vías metabólicas

Abstract: Background: Mitochondriopathies are multisystem diseases affecting the oxidative phosphorylation (OXPHOS) system. Skin fibroblasts are a good model for the study of these diseases. Fibroblasts with a complex IV mitochondriopathy were used to determine the molecular mechanism and the main affected functions in this disease. Methods: Skin fibroblast were grown to assure disease phenotype. Mitochondria were isolated from these cells and their proteome extracted for protein identification. Identified proteins were validated with the MitoMiner database. Results: Disease phenotype was corroborated on skin fibroblasts, which presented a complex IV defect. The mitochondrial proteome of these cells showed that the most affected proteins belonged to the OXPHOS system, mainly to the complexes that form supercomplexes or respirosomes (I, III, IV, and V). Defects in complex IV seemed to be due to assembly issues, which might prevent supercomplexes formation and efficient substrate channeling. It was also found that this mitochondriopathy affects other processes that are related to DNA genetic information flow (replication, transcription, and translation) as well as beta oxidation and tricarboxylic acid cycle. Conclusions: These data, as a whole, could be used for the better stratification of these diseases, as well as to optimize management and treatment options.

Resumen: Introducción: Las mitocondriopatías son enfermedades multisistémicas que afectan el funcionamiento de la fosforilación oxidativa (OXPHOS). Un buen modelo de estudio para estas enfermedades es el cultivo primario de fibroblastos. En este trabajo se utilizaron fibroblastos con mitocondriopatía del complejo IV para determinar cuáles son las principales funciones afectadas en esta enfermedad. Métodos: Se realizaron cultivos primarios de fibroblastos para corroborar el fenotipo de la enfermedad. Las mitocondrias se aislaron de estas células y se extrajo su proteoma para su identificación. Las proteínas identificadas se validaron con la base de datos de MitoMiner. Resultados: Los fibroblastos conservaron el fenotipo de la enfermedad que incluye un defecto del complejo IV. El proteoma mitocondrial de estas células mostró que las proteínas más afectadas pertenecen al sistema de OXPHOS, principalmente los complejos que forman supercomplejos o respirosomas (I, III, IV y V). El defecto en el complejo IV al parecer se debió a problemas de ensamblaje que pueden evitar la formación de los supercomplejos y la eficiente canalización de sustratos. También se observó que esta mitocondriopatía afecta otros procesos relacionados con el flujo de información genética del DNA (replicación, transcripción y traducción), así como con la beta oxidación y el ciclo de los ácidos tricarboxílicos (TCA). Conclusiones: En conjunto, estos datos podrían utilizarse para una mejor clasificación de estas enfermedades, así como para la optimización de las opciones de manejo y tratamiento.

Proteomic changes in a childhood acute lymphoblastic leukemia cell line during the adaptation to vincristine / Cambios en el proteoma de una línea celular de leucemia linfoblástica aguda infantil durante la adaptación a vincristina

Abstract: Introduction: Relapse occurs in approximately 20% of Mexican patients with childhood acute lymphoblastic leukemia (ALL). In this group, chemoresistance may be one of the biggest challenges. An overview of complex cellular processes like drug tolerance can be achieved with proteomic studies. Methods: The B-lineage pediatric ALL cell line CCRF-SB was gradually exposed to the chemotherapeutic vincristine until proliferation was observed at 6 nM, control cells were cultured in the absence of vincristine. The proteome from each group was analyzed by nanoHPLC coupled to an ESI-ion trap mass spectrometer. The identified proteins were grouped into over-represented functional categories with the PANTHER classification system. Results: We found 135 proteins exclusively expressed in the presence of vincristine. The most represented functional categories were: Toll receptor signaling pathway, Ras Pathway, B and T cell activation, CCKR signaling map, cytokine-mediated signaling pathway, and oxidative phosphorylation. Conclusions: Our study indicates that signal transduction and mitochondrial ATP production are essential during adaptation of leukemic cells to vincristine, these processes represent potential therapeutic targets.

Resumen: Introducción: Aproximadamente el 20% de los pacientes mexicanos con leucemia linfoblástica aguda (LLA) infantil presentan recaídas. En este grupo, la quimiorresistencia es uno de los principales desafíos. Los estudios proteómicos pueden dar un panorama general de procesos celulares complejos como la tolerancia a fármacos. Métodos: La línea celular de LLA de linaje B, CCRF-SB, fue expuesta de manera gradual al fármaco quimioterapéutico vincristina hasta observar proliferación celular en presencia de 6 nM, como control se cultivaron células en ausencia del fármaco. Se analizó el proteoma de cada grupo mediante nanoHPLC acoplado a un espectrómetro de masas de tipo trampa de iones. Las proteínas identificadas se agruparon en categorías funcionales sobre-representadas con el sistema de clasificación PANTHER. Resultados: Encontramos 135 proteínas expresadas exclusivamente en presencia de vincristina. Las categorías funcionales más representadas fueron la señalización asociada a los receptores tipo Toll, señalización dependiente de Ras, activación de células B y T, mapa de señalización CCKR, señalización mediada por citoquinas y la fosforilación oxidativa. Conclusiones: Nuestro estudio indica que la transducción de señales y la producción de ATP mitocondrial son procesos esenciales durante la adaptación de células leucémicas a vincristina por lo que estos procesos representan potenciales blancos terapéuticos.

Characterization of Cry toxins from autochthonous Bacillus thuringiensis isolates from Mexico / Caracterización de toxinas Cry de aislados de Bacillus thuringiensis autóctonos de México

Abstract: Background: Chemical pesticides, widely used in agriculture and vector-borne disease control, have shown toxic effects on the environment and the people in contact with them. Bacillus thuringiensis is a widely used bacterium for alternative and safer control of insect pests. Its toxins are specific for insects but innocuous for mammals and may be used as powerful adjuvants when applied with vaccines. The objective of this work was to characterize some autochthonous B. thuringiensis strains, which could be used for the control of a local pest (Diatraea considerata Heinrich) that affects sugar cane crops in Sinaloa, Mexico. Also, to evaluate these strains as a source of Cry toxins, which may be used in the future as adjuvants for some vaccines. Methods: Eight strains from field-collected dead insects were isolated. These were microbiologically identified as B. thuringiensis and confirmed by amplification and sequencing of 16S rDNA. Bioassays were performed to evaluate their pathogenicity against D. considerata, and Cry toxins were identified by proteomic analyses. Results: An increased mortality among larvae infected with strain Bt-D was observed, and its toxin was identified as Cry1Ac. Conclusions: The observed data showed that the selected strain was pathogenic to D. considerata and seemed to produce Cry1Ac protein, which has been reported as an adjuvant in different types of immunization.

Resumen: Introducción: Los pesticidas químicos, ampliamente usados en agricultura y en el control de vectores transmisores de enfermedades, han mostrado efectos tóxicos sobre el medio ambiente y las personas expuestas a ellos. Bacillus thuringiensis es una bacteria ampliamente utilizada como una alternativa segura y eficaz en el control biológico de plagas agrícolas. Sus toxinas son específicas de insectos, pero inocuas para mamíferos, e incluso poseen gran potencial para ser usadas como adyuvantes en vacunas. El objetivo de este trabajo fue caracterizar cepas autóctonas de B. thuringiensis con efectividad contra el gusano barrenador (Diatraea considerata Heinrich) de la caña de azúcar en cultivos del estado de Sinaloa, México, y como fuente de proteínas Cry, con potencial de utilizarse como adyuvantes en vacunas. Métodos: Se lograron aislar ocho cepas a partir de insectos muertos en campos agrícolas, las cuales fueron identificadas microbiológicamente como B. thuringiensis, lo que se confirmó por amplificación y secuenciación del 16S rDNA. La efectividad de los aislados para el control del gusano barrenador fue evaluada mediante bioensayos y las toxinas Cry fueron identificadas por análisis proteómico. Resultados: Se observó una mortalidad elevada en las larvas infectadas con las cepas de estudio. Particularmente, la cepa Bt-D, de la cual el análisis molecular mostró que posee una toxina tipo Cry1Ac. Conclusiones: Los resultados mostraron que la cepa Bt-D posee un elevado potencial patogénico hacia D. considerata y produce la proteína Cry1Ac, de la cual existen reportes de su aplicación como adyuvante en diferentes formas de inmunización.

Omics-based biomarkers: current status and potential use in the clinic / Biomarcadores basados en tecnologías ómicas: estado actual y su potencial uso en la clínica

Abstract: In recent years, the use of high-throughput omics technologies has led to the rapid discovery of many candidate biomarkers. However, few of them have made the transition to the clinic. In this review, the promise of omics technologies to contribute to the process of biomarker development is described. An overview of the current state in this area is presented with examples of genomics, proteomics, transcriptomics, metabolomics and microbiomics biomarkers in the field of oncology, along with some proposed strategies to accelerate their validation and translation to improve the care of patients with neoplasms. The inherent complexity underlying neoplasms combined with the requirement of developing well-designed biomarker discovery processes based on omics technologies present a challenge for the effective development of biomarkers that may be useful in guiding therapies, addressing disease risks, and predicting clinical outcomes.

Resumen: En los últimos años, el uso de las tecnologías ómicas de alta densidad de datos ha permitido el rápido descubrimiento de posibles biomarcadores. Sin embargo, esto no ha tenido un impacto notable en la clínica ya que se han implementado muy pocos de esos biomarcadores. En el presente documento se describe el potencial de las tecnologías ómicas en el desarrollo de nuevos biomarcadores. Con el objetivo de dar a conocer un panorama general de la situación actual, se comentan algunos ejemplos ilustrativos de biomarcadores genómicos, transcriptómicos, proteómicos, metabolómicos y microbiómicos en el campo de la investigación en oncología. Asimismo, se señalan algunas de las recomendaciones que se han propuesto para acelerar su validación e implementación, y se comenta sobre cómo la complejidad inherente a las enfermedades se combina con la complejidad de las tecnologías ómicas, de tal modo que el desarrollo de biomarcadores predictivos, pronósticos y diagnósticos eficientes plantea retos importantes.