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Neonicotinoids (NEOs) are widely used insecticides that are ubiquitous in agricultural use. Since NEOs are found in natural waters as well as in tap water and human urine in regions where NEOs are widely used, NEOs pose a potential hazard to non-target organisms such as animals and humans. Some of the commonly detected NEOs are imidacloprid (IMD), thiamethoxam (TMX), and its metabolite clothianidin (CLO). Although previously published scientific information, including an assessment of the environmental risks, particularly for bees, had resulted in a ban on the outdoor use of these three NEOs in the EU - their use is now only permitted in closed greenhouses - these NEOs continue to be used in agriculture in many other parts of the world. Therefore, a detailed study and comparison of the effects of NEOs on the embryonic development of non-target organisms is needed to further define the risk profiles. Embryos of the South African clawed frog Xenopus laevis, a well-established aquatic model, were exposed to different concentrations of IMD, TMX, or CLO (0.1-100 mg/L) to study and compare the possible effects of a single contaminant in natural water bodies on early embryogenesis. The results included a reduced body length, a smaller orbital space, impaired cranial cartilage and nerves, and an altered heart structure and function. At the molecular level, NEO exposure partially resulted in an altered expression of tissue-specific factors, which are involved in eye, cranial placode, and heart development. Our results suggest that the NEOs studied negatively affect the embryonic development of the non-target organism X. laevis. Since pesticides, especially NEOs, pollute the environment worldwide, it is suggested that they are strictly controlled and monitored in the areas where they are used. In addition, the question arises as to whether pesticide metabolites also pose a risk to the environment and need to be investigated further so that they can be taken into account when registering ingredients.
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Mutations in PRDM15 lead to a syndromic form of holoprosencephaly (HPE) known as the Galloway-Mowat syndrome (GAMOS). While a connection between PRDM15, a zinc finger transcription factor, and WNT/PCP signaling has been established, there is a critical need to delve deeper into their contributions to early development and GAMOS pathogenesis. We used the South African clawed frog Xenopus laevis as the vertebrate model organism and observed that prdm15 was enriched in the tissues and organs affected in GAMOS. Furthermore, we generated a morpholino oligonucleotide-mediated prdm15 knockdown model showing that the depletion of Prdm15 leads to abnormal eye, head, and brain development, effectively recapitulating the anterior neural features in GAMOS. An analysis of the underlying molecular basis revealed a reduced expression of key genes associated with eye, head, and brain development. Notably, this reduction could be rescued by the introduction of wnt4 RNA, particularly during the induction of the respective tissues. Mechanistically, our data demonstrate that Prdm15 acts upstream of both canonical and non-canonical Wnt4 signaling during anterior neural development. Our findings describe severe ocular and anterior neural abnormalities upon Prdm15 depletion and elucidate the role of Prdm15 in canonical and non-canonical Wnt4 signaling.
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Objective: A simulated conversation between a physician and a family member, i.e., a medical conversation, was changed from a conventional face-to-face conversation (SS 2019) to a telehealth conversation (SS 2020) due to the COVID-19 pandemic. The medical education conversation is part of the biochemistry seminar "From Genes to Proteins" which second semester human medicine students take. The objective of this study was to analyze to what extent the switch from face-to-face to telehealth conversations affected student satisfaction and motivation. Methodology: In the seminar, students study biochemical as well as competency-oriented content, such as how to talk to family members. In the summer semester of 2019, students were trained how to talk to their patients' family members in a traditional conversation setting with the help of lay actors in a classroom format. In the summer semester of 2020, this conversation took place under comparable conditions, but in the form of an online telehealth conversation instead. Student satisfaction and motivation were surveyed by means of an evaluation questionnaire following the seminar in both semesters. Results: Both conversation formats achieved a high level of satisfaction from students (school grade A-B). For some evaluation items, such as "realistic conversation simulation", the face-to-face conversation was perceived as more satisfying (Md=5.0, IQR=1.0) than the telehealth conversation (Md=5.0, IQR=2.0). In addition, the face-to-face conversation resulted in higher subjective motivation from students (Md=5.0, IQR=1.0) than that of the telehealth conversation (Md=4.0, IQR=2.0). Conclusion: The high student satisfaction and acceptance of both didactic concepts leads to the conclusion that the simulated telehealth conversation is an adequate substitute for the simulation of a traditional face-to-face conversation with regard to the parameters that were studied.
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Educação Médica , Estudantes de Medicina , Telemedicina , Humanos , Pandemias , Encaminhamento e ConsultaRESUMO
Acetamiprid (ACT) is used extensively in agriculture worldwide, although data on ACT concentrations in natural water bodies and its impact on aquatic organisms are limited. To study whether ACT influences the embryogenesis of the South African clawed frog Xenopus laevis, embryos were incubated in ACT solutions from 0.01 to 100 mg/L. The low concentrations were chosen on the basis of concentrations already found in nature. ACT treatment leads to shorter embryo lengths, intestine malformation and reduced eye areas. It also affects the cranial cartilage and cardiac development as well as the embryo's mobility. The expression of tissue-specific marker genes is affected as well. Thus, our study suggests that pesticides may lead to an increased mortality of non-target organisms and emphasizes the importance of regular testing for ACT concentrations in nature. Our study provides an overview of ACT effects and can therefore be used as a basis for an ACT risk assessment.
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Inseticidas , Animais , Inseticidas/toxicidade , Xenopus laevis , Organismos Aquáticos , Desenvolvimento Embrionário , Embrião não MamíferoRESUMO
BACKGROUND: Glyphosate (GLY) is the most widely used herbicide in the world. Due to its mode of action as an inhibitor of the 5-enolpyruvylshikimate-3-phosphate synthase, an important step in the shikimate pathway, specifically in plants, GLY is considered to be of low toxicity to non-target organisms. However, various studies have shown the negative effects of GLY on the mortality and development of different non-target organisms, including insects, rodents, fish and amphibians. To better understand the various effects of GLY in more detail, we studied the effects of GLY without co-formulants during the embryogenesis of the aquatic model organism Xenopus laevis. RESULTS: A treatment with GLY affected various morphological endpoints in X. laevis tadpoles (body length, head width and area, eye area). Additionally, GLY interfered with the mobility as well as the neural and cardiac development of the embryos at stage 44/45. We were able to detect detailed structural changes in the cranial nerves and the heart and gained insights into the negative effects of GLY on cardiomyocyte differentiation. CONCLUSION: The application of GLY without co-formulants resulted in negative effects on several endpoints in the early embryonic development of X. laevis at concentrations that are environmentally relevant and concentrations that reflect the worst-case scenarios. This indicates that GLY could have a strong negative impact on the survival and lives of amphibians in natural waters. As a result, future GLY approvals should consider its impact on the environment.
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Desenvolvimento Embrionário , Animais , Xenopus laevis/fisiologia , África do Sul , GlifosatoRESUMO
Background and objectives: According to the World Health Organization, climate change constitutes the single greatest threat to human health. However, the health care system contributes to climate change worldwide through its high CO2 emissions. In order to make future physicians more aware of this issue and to expand medical education to include climate-related aspects, the mandatory 28 academic hours elective "Climate Change and Health" for students of human medicine in the preclinical study stage was implemented at the Medical Faculty of Ulm in the 2020/21 winter semester. Our accompanying study investigated 1. in what form the topic of climate change can be successfully integrated into the study of human medicine in a manner that includes student opinions and2. whether being required to take an elective on the topic led to changes in student environmental knowledge and awareness. Methodology: Personal individual interviews were conducted with all n=11 students after the course in a pilot that was carried out in the 2020/21 winter semester to determine course feasibility and student acceptance. The students were also able to evaluate the course using an evaluation form and were asked to complete a questionnaire on their environmental knowledge and awareness before and after the course. The course was revised on the basis of the results and offered again in the 2021 summer semester with an intervention group (n=16, participation in the mandatory elective) and a comparison group (n=25, no participation in the mandatory elective). The intervention group was asked to evaluate the course on the evaluation form. Both groups completed the environmental questionnaire at the same time. Results: The positive feedback from students for both semesters indicates a good feasibility and acceptance of the course. Student environmental knowledge was increased in both semesters. However, there were only few observable changes in student environmental awareness. Conclusion: This paper illustrates how the topic of climate change and health can be embedded into medical studies. The students considered climate change an important topic and drew added value from the course for their future work in healthcare. The study shows that knowledge transfer at the university level is an effective way to educate the young generation on climate change and its impacts.
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Educação Médica , Estudantes de Medicina , Humanos , Docentes de Medicina , Mudança Climática , Estudantes , Atenção à Saúde , CurrículoRESUMO
Objective: Climate change constitutes a major challenge. The higher education sector plays an important role in regard to climate change and the adaptation to its consequences. Various approaches toward the integration of environmental subject areas to higher education teaching have already been described in other studies, but there is a lack of data supporting the effectiveness of these approaches in changing not only the environmental knowledge of students, but also their awareness. To address this, the present study tracked whether student attitudes about the environment could be changed by implicitly addressing medically relevant environmental topics as part of an online seminar. Methods: Second semester students of molecular medicine attending a mandatory 14-hour online seminar, which was required to obtain additive key qualifications and which consisted of independent study phases as well as online class meetings, were divided into two groups: the intervention group (IG, n=27, thereof 20 in the pretest and 21 in the posttest) was exposed to medically relevant environmental topics, while the comparison group (CG, n=26, thereof 22 in the pretest and 21 in the posttest) was exposed to general, non-environmental medical topics. Surveys were conducted with standardized questionnaires before and after the seminar in order to study the influence on the students' environmental knowledge, awareness and other personal attitudes. Results: While the seminar did not significantly change the environmental awareness in either group, the environmental knowledge of the IG was significantly increased by the group's exposure to environmental topics. In addition, the IG assessed its own environmental awareness regarding sustainable working methods in a laboratory as significantly higher after the seminar than the CG did, and some students of the IG had become more interested in issues relating to sustainability. Conclusion: The approach used to communicate environmental content mainly increased the environmental knowledge of students and piqued the interest of some students in climate-related and environmental topics. However, it was not possible to change deeper personal attitudes about environmental awareness, especially everyday behavior.
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Educação Médica , Estudantes de Medicina , Estudantes de Enfermagem , Humanos , Currículo , Atitude , Inquéritos e QuestionáriosRESUMO
BACKGROUND: The issues of climate change, environmental pollution and species extinction play an increasingly central role in the public debate. At the same time, however, there is a significant gap between environmental knowledge and sustainable action (so-called value-action gap). The education system, especially at the university level, is an important institution for imparting well-founded knowledge on this topic and, as a consequence, for deriving targeted options for action. The present study surveyed the current environmental knowledge and environmental awareness, including the everyday behavior of Generation Z students of medical study programs in comparison to science-oriented study programs. METHODS: In October/November 2021, an anonymous and voluntary online survey was conducted at the University of Ulm to evaluate the environmental knowledge and awareness of students in all semesters of the study programs Human Medicine, Dentistry, Molecular Medicine, Biology and Teaching. A total of 317 students fully completed the questionnaire. RESULTS: The results confirm the current state of studies on the environmental awareness of the German population. The value-action gap is also detectable among students. Students perceive the urgency of environmental protection measures and action on climate change and also link these with emotional feelings, but in terms of behavior personal interests (still) tend to largely take precedence over environmental protection. In addition, according to our findings, the image of stereotypes and prejudices of the various courses of study is also partially confirmed in the surveyed environmental awareness. CONCLUSION: The significant differences in environmental awareness between the compared degree programs and the gap between knowledge and action call for an individual but consistent implementation of the topics of climate change and environmental protection in the curriculum in all investigated degree courses. With the knowledge and awareness gained in this way, academics as distinguished members of society can convey climate awareness and fulfill the function as a role model.
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Estudantes de Medicina , Estudantes , Humanos , Alemanha , Estudantes/psicologia , Currículo , Inquéritos e Questionários , Conhecimentos, Atitudes e Prática em Saúde , Estudantes de Medicina/psicologiaRESUMO
There is increasing concern about the health effects of pesticides that pollute natural waters. In particular, the use of neonicotinoids, such as thiacloprid (THD), is causing unease. THD is considered non-toxic to non-target vertebrates. Studies classify THD as carcinogenic, toxic to reproduction, and therefore harmful to the environment. A detailed study of possible THD effects during the amphibian embryogenesis is needed because leaching can introduce THD into aquatic environments. We incubated stage 2 embryos of the South African clawed frog in various THD concentrations (0.1-100 mg/L) at 14 °C to study the potential effects of a one-time THD contamination of waters on the early embryogenesis. We showed that THD has, indeed, negative effects on the embryonic development of the X. laevis. A treatment with THD led to a reduced embryonic body length and mobility. Furthermore, a treatment with THD resulted in smaller cranial cartilages, eyes and brains, and the embryos had shorter cranial nerves and an impaired cardiogenesis. On a molecular basis, THD led to a reduced expression of the brain marker emx1 and the heart marker mhcα. Our results underly the importance of a strict and efficient monitoring of the regulatory levels and application areas of THD.
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Desenvolvimento Embrionário , Inseticidas , Animais , Xenopus laevis , África do Sul , Neonicotinoides/toxicidade , Inseticidas/toxicidadeRESUMO
BACKGROUND: According to the World Health Organisation, climate change poses the greatest health threat to humanity. At the same time, an environmentally friendly lifestyle has a positive impact on our health, such as a plant-based diet. In order to counter climate change, society needs to be informed about climate-friendly and health-promoting measures. Therefore, an online workshop was initiated at the Medical Faculty of Ulm. In an accompanying study, it was determined whether this leads to changes in environmental knowledge and awareness among the participants. METHODS: The online workshop consisted of four 2-hour sessions. Scientific basics on climate change and possible solutions were discussed. Other focuses were on health and the health system as well as environmental psychology and climate change denial. Participants could take part in an anonymous and voluntary online survey before (pretest) and after (posttest) the workshop. RESULTS: 86 participants took part in the workshop, of whom 24 attended all appointments and completed both surveys. While hardly any changes were observed in the subsection of environmental emotion, perception and behavior, there was a significant increase in environmental knowledge in the posttest. The workshop was evaluated very positively. Furthermore, many participants were motivated to make a personal contribution to climate protection after the workshop. CONCLUSION: The workshop serves as a good example of how medical scientific findings can be shared at a societal level. The participants already showed a high level of environmental awareness in the pretest, which is why the question remains open as to how people can be sensitized to the (health) threats posed by climate change who do not proactively sign up for such an offer themselves.
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Mudança Climática , Docentes de Medicina , Humanos , Alemanha , Inquéritos e QuestionáriosRESUMO
Galloway-Mowat syndrome (GAMOS) is a very rare condition characterized by early-onset nephrotic syndrome and microcephaly with variable neurologic features. While considerable genetic heterogeneity of GAMOS has been identified, the majority of cases are caused by pathogenic variants in genes encoding the four components of the Kinase, endopeptidase, and other proteins of small size (KEOPS) complex, one of which is TP53RK. Here we describe a 3-year-old male with progressive microcephaly, neurodevelopmental deficits, and glomerular proteinuria. He was found to carry a novel homozygous TP53RK missense variant, c.163C>G (p.Arg55Gly), which was considered as potentially disease-causing. We generated a morpholino tp53rk knockdown model in Xenopus laevis showing that the depletion of endogenous Tp53rk caused abnormal eye and head development. This phenotype could be rescued by the expression of human wildtype TP53RK but not by the c.163C>G mutant nor by another previously described GAMOS-associated mutant c.125G>A (p.Gly42Asp). These findings support the pathogenic role of the novel TP53RK variant.
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Hérnia Hiatal , Microcefalia , Nefrose , Síndrome Nefrótica , Masculino , Humanos , Pré-Escolar , Microcefalia/genética , Mutação , Nefrose/genética , Hérnia Hiatal/genética , Síndrome Nefrótica/genéticaRESUMO
INTRODUCTION: Global warming is one of the major challenges faced by society. To raise students' awareness related to these issues, we embedded environmental learning goals in mandatory courses. The aim of this initiative was to determine whether the integration of environmental topics into a seminar focussing on non-environmental issues facilitates changes in the environmental knowledge and awareness of students. METHODS: Third-year molecular medicine students (intervention group, IG) were assigned short environment-related tasks in a not environment-related scientific writing seminar. Using standardized questionnaires before and after the seminar, students were assessed with a set of up to 48 questions relating to environmental affect, cognition, behavioral intention and knowledge as well as specific personal aspects. The comparison group (CG) consisted of third-year dentistry students who were assessed twice during an equivalent pre-post time period and who did not attend the seminar. The analysis was carried out with 15 students from the IG and 14 from the CG. RESULTS: Integrating environmental topics into a scientific writing seminar focussing on non-environmental issues led to a significantly increased level of awareness of the threats from climate change in the IG (p<0.05). No further significant changes in other areas of environmental awareness were observed in either group. In addition, the environmental knowledge of the IG students (p≤0.01) but not of the CG students (p=0.285) was significantly enhanced. DISCUSSION AND CONCLUSION: This study shows that factual environmental knowledge on the issues addressed can be increased by a cautious approach of environmental communication. However, more deep-seated personal attitudes and environmental awareness cannot be sustainably changed using this low intervention dose.
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Educação Médica , Estudantes de Medicina , Humanos , Alemanha , Inquéritos e Questionários , CurrículoRESUMO
Block of proliferation 1 (Bop1) is a nucleolar protein known to be necessary for the assembly of the 60S subunit of ribosomes. Here, we show a specific bop1 expression in the developing anterior tissue of the South African clawed frog Xenopus laevis. Morpholino oligonucleotide-mediated knockdown approaches demonstrated that Bop1 is required for proper development of the cranial cartilage, brain, and the eyes. Furthermore, we show that bop1 knockdown leads to impaired retinal lamination with disorganized cell layers. Expression of neural crest-, brain-, and eye-specific marker genes was disturbed. Apoptotic and proliferative processes, which are known to be affected during ribosomal biogenesis defects, are not hindered upon bop1 knockdown. Because early Xenopus embryos contain a large store of maternal ribosomes, we considered if Bop1 might have a role independent of de novo ribosomal biogenesis. At early embryonic stages, pax6 expression was strongly reduced in bop1 morphants and synergy experiments indicate a common signaling pathway of the two molecules, Bop1 and Pax6. Our studies imply a novel function of Bop1 independent of ribosomal biogenesis.
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Crista Neural , Ribossomos , Animais , Proliferação de Células , Crista Neural/metabolismo , Proteínas Nucleares/genética , Ribossomos/genética , Ribossomos/metabolismo , Proteínas de Xenopus/genética , Proteínas de Xenopus/metabolismo , Xenopus laevisRESUMO
On-site teaching at Ulm University was restricted in the summer semester (SS) 2020 due to the Corona pandemic. The biochemistry seminar "From gene to protein" in the 2nd preclinical semester, which had been successfully conducted as an Inverted Classroom (IC), had to be changed to an online concept. The aim of this study was to analyze the concept conversion in terms of students' satisfaction and knowledge acquisition. In the seminar, human medical students of the 2nd semester acquired biochemical and competency-oriented learning content. In SS2019, the course was taught using the IC concept. For the conversion to the online format in the SS2020, alternative teaching materials were developed and used. Students' satisfaction was assessed by an evaluation questionnaire and knowledge acquisition was tested by a written biochemistry exam. For both teaching concepts a high level of satisfaction was detected. Individual evaluation criteria were evaluated similarly by the students for both concepts. The online concept led up to significantly higher biochemistry exam scores. Due to the high levels of students' satisfaction in both concepts and the results of the biochemistry exam, it can be concluded that online teaching offers a suitable substitute. Based on these results, it is worth to undertake further research on digitization of studies.
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Currículo , Estudantes de Medicina , Bioquímica/educação , Humanos , Aprendizagem , Satisfação PessoalRESUMO
Background: Although the majority of medical students in Germany pursue a doctorate, only a portion of them receive a standardized scientific training, which is reflected in the quality issues seen in medical doctoral theses. The course Medical Dissertation Basics was conceptualized and scientifically monitored in order to support medical doctoral students on the one hand and to improve the quality of their scientific work on the other. Methodology: The course consists of three modules. Module I, which is an introductory module, covers time and writing management and addresses how to approach literature and the principles of scientific work as well as the chapters required in a dissertation and the dissertation presentation and defense. In the practical module II, doctoral students write sections of their dissertation chapters and receive feedback via peer and expert reviews. Module III includes training on dissertation presentations and their defense. For objective analysis purposes, a multiple-choice test was administered before and after module I. Medical students from semesters 2 to 6 served as a control group. Questionnaires were used to subjectively analyze the training and support functions of modules I-III. Results: High participation rates and the fact that the modules were taught numerous times show that doctoral students accept the courses. The objective analysis of module I showed a highly significant knowledge acquisition of the course group (N=55) in contrast to the control group (N=34). The doctoral students rated the course modules I-III with grades between 1.0 and 1.25 (grade A+/A; N=20-65 SD=0-0.44), felt well supported and estimated their learning success as high. Conclusion: The study indicates knowledge acquisition in module I and a high doctoral student satisfaction with all modules. For an objective analysis of modules II-III, a comparison of completed doctoral theses (course participants vs. non-participants) would be appropriate but would only make sense in a few years. Based on the results of our study, we recommend that other faculties implement similar courses.
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Médicos , Estudantes de Medicina , Currículo , Alemanha , Humanos , Inquéritos e Questionários , RedaçãoRESUMO
Ribosomal biogenesis is a fundamental process necessary for cell growth and division. Ribosomal protein L5 (Rpl5) is part of the large ribosomal subunit. Mutations in this protein have been associated with the congenital disease Diamond Blackfan anemia (DBA), a so called ribosomopathy. Despite of the ubiquitous need of ribosomes, clinical manifestations of DBA include tissue-specific symptoms, e.g., craniofacial malformations, eye abnormalities, skin pigmentation failure, cardiac defects or liver cirrhosis. Here, we made use of the vertebrate model organism Xenopus laevis and showed a specific expression of rpl5 in the developing anterior tissue correlating with tissues affected in ribosomopathies. Upon Rpl5 knockdown using an antisense-based morpholino oligonucleotide approach, we showed different phenotypes affecting anterior tissue, i.e., defective cranial cartilage, malformed eyes, and microcephaly. Hence, the observed phenotypes in Xenopus laevis resemble the clinical manifestations of DBA. Analyses of the underlying molecular basis revealed that the expression of several marker genes of neural crest, eye, and brain are decreased during induction and differentiation of the respective tissue. Furthermore, Rpl5 knockdown led to decreased cell proliferation and increased cell apoptosis during early embryogenesis. Investigating the molecular mechanisms underlying Rpl5 function revealed a more than additive effect between either loss of function of Rpl5 and loss of function of c-Myc or loss of function of Rpl5 and gain of function of Tp53, suggesting a common signaling pathway of these proteins. The co-injection of the apoptosis blocking molecule Bcl2 resulted in a partial rescue of the eye phenotype, supporting the hypothesis that apoptosis is one main reason for the phenotypes occurring upon Rpl5 knockdown. With this study, we are able to shed more light on the still poorly understood molecular background of ribosomopathies.
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BACKGROUND: Worldwide, amphibian populations are declining drastically. One reason might be the use of pesticides including herbicides. The herbicide glyphosate is an inhibitor of the 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase of the plant shikimate pathway, preventing the formation of aromatic amino acids and thus inducing plant death. Due to this specific action, GBH are considered nontoxic to non-target organisms. However, GBH impairs embryonic development of chickens, amphibians and fishes. So far, no detailed tissue- and organ-specific analysis of the effects of GBH during development in amphibians has been performed. RESULTS: We demonstrated that GBH Roundup® LB plus has a negative effect on embryonic development of the South African clawed frog Xenopus laevis. GBH treatment with sublethal concentrations resulted in a reduced body length and mobility of embryos. Furthermore, incubation with GBH led to smaller eyes, brains and cranial cartilages in comparison to untreated embryos. GBH incubation also resulted in shorter cranial nerves and had an effect on cardiac development including reduced heart rate and atrium size. On a molecular basis, GBH treatment led to reduced expression of marker genes in different tissues and developmental stages. CONCLUSION: GBH leads to disturbed embryonic development of Xenopus laevis.
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Herbicidas , Poluentes Químicos da Água , Animais , Galinhas , Desenvolvimento Embrionário , Glicina/análogos & derivados , Herbicidas/toxicidade , Poluentes Químicos da Água/toxicidade , Xenopus laevis , GlifosatoRESUMO
Galloway-Mowat syndrome (GAMOS) is a rare developmental disease. Patients suffer from congenital brain anomalies combined with renal abnormalities often resulting in an early-onset steroid-resistant nephrotic syndrome. The etiology of GAMOS has a heterogeneous genetic contribution. Mutations in more than 10 different genes have been reported in GAMOS patients. Among these are mutations in four genes encoding members of the human KEOPS (kinase, endopeptidase and other proteins of small size) complex, including OSGEP, TP53RK, TPRKB and LAGE3. Until now, these components have been functionally mainly investigated in bacteria, eukarya and archaea and in humans in the context of the discovery of its role in GAMOS, but the KEOPS complex members' expression and function during embryogenesis in vertebrates is still unknown. In this study, in silico analysis showed that both gene localization and the protein sequences of the three core KEOPS complex members Osgep, Tp53rk and Tprkb are highly conserved across different species including Xenopus laevis. In addition, we examined the spatio-temporal expression pattern of osgep, tp53rk and tprkb using RT-PCR and whole mount in situ hybridization approaches during early Xenopus development. We observed that all three genes were expressed during early embryogenesis and enriched in tissues and organs affected in GAMOS. More precisely, KEOPS complex genes are expressed in the pronephros, but also in neural tissue such as the developing brain, eye and cranial cartilage. These findings suggest that the KEOPS complex plays an important role during vertebrate embryonic development.
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Biologia Computacional , Nefrose , Animais , Desenvolvimento Embrionário/genética , Hérnia Hiatal , Humanos , Microcefalia , Proteínas de Xenopus/genética , Xenopus laevisRESUMO
BACKGROUND: Galloway-Mowat syndrome (GAMOS) is characterized by neurodevelopmental defects and a progressive nephropathy, which typically manifests as steroid-resistant nephrotic syndrome. The prognosis of GAMOS is poor, and the majority of children progress to renal failure. The discovery of monogenic causes of GAMOS has uncovered molecular pathways involved in the pathogenesis of disease. METHODS: Homozygosity mapping, whole-exome sequencing, and linkage analysis were used to identify mutations in four families with a GAMOS-like phenotype, and high-throughput PCR technology was applied to 91 individuals with GAMOS and 816 individuals with isolated nephrotic syndrome. In vitro and in vivo studies determined the functional significance of the mutations identified. RESULTS: Three biallelic variants of the transcriptional regulator PRDM15 were detected in six families with proteinuric kidney disease. Four families with a variant in the protein's zinc-finger (ZNF) domain have additional GAMOS-like features, including brain anomalies, cardiac defects, and skeletal defects. All variants destabilize the PRDM15 protein, and the ZNF variant additionally interferes with transcriptional activation. Morpholino oligonucleotide-mediated knockdown of Prdm15 in Xenopus embryos disrupted pronephric development. Human wild-type PRDM15 RNA rescued the disruption, but the three PRDM15 variants did not. Finally, CRISPR-mediated knockout of PRDM15 in human podocytes led to dysregulation of several renal developmental genes. CONCLUSIONS: Variants in PRDM15 can cause either isolated nephrotic syndrome or a GAMOS-type syndrome on an allelic basis. PRDM15 regulates multiple developmental kidney genes, and is likely to play an essential role in renal development in humans.