A central question in the science of science concerns how to develop a quantitative understanding of the evolution and impact of individual careers. Over the course of history, a relatively small fraction of individuals have made disproportionate, profound, and lasting impacts on science and society. Despite a long-standing interest in the careers of scientific elites across diverse disciplines, it remains difficult to collect large-scale career histories that could serve as training sets for systematic empirical and theoretical studies. Here, by combining unstructured data collected from CVs, university websites, and Wikipedia, together with the publication and citation database from Microsoft Academic Graph (MAG), we reconstructed publication histories of nearly all Nobel prize winners from the past century, through both manual curation and algorithmic disambiguation procedures. Data validation shows that the collected dataset presents among the most comprehensive collection of publication records for Nobel laureates currently available. As our quantitative understanding of science deepens, this dataset is expected to have increasing value. It will not only allow us to quantitatively probe novel patterns of productivity, collaboration, and impact governing successful scientific careers, it may also help us unearth the fundamental principles underlying creativity and the genesis of scientific breakthroughs.  Although scientific elites have attracted ensuing interest from a wide range of disciplines38, spanning across sociology, economics, psychology, and physics, most quantitative analyses have focused on the prize-winning work alone, exploring the link between age and creativity20, and allocation of credits and recognition32. On the other hand, the rich patterns unveiled in qualitative studies29, including Zuckerman’s canonical work, vividly illustrates that the careers of scientific elites encompass projects well beyond their prize-winning work. Career-level analyses of these scientific elites have remained elusive, largely due to the difficulty in obtaining large-scale, high-resolution individual career histories.Recent advances in the science of science have revealed a series of highly reproducible patterns governing the fundamental dynamic of scienceet al.39 introduced an open database on scientific output of Vietnamese researchers in social sciences and humanities; and Morrison et al.40 developed a name disambiguation method for inventors and assignees on 8.47 million patents. While these open-accessed datasets are mostly about ordinary scientists, researchers have also been interested in Nobel laureates. For example, Jones et al.19 collected a biographical dataset of 525 Nobel Prize winners, and Fortunato et al.30 curated data on dates of birth, the year of Nobel prizes and year(s) of publication(s) of prize-winning work. Chan et al.41 collected a dataset consisting of 34,448 publications of 192 Nobel laureates between 1970 and 2000 based on the Scopus dataset. Li et al.42 collected a fraction of papers published by Nobel laureates during the period of 1901–2012 using the Web of Science data. These efforts are highly complementary to the dataset presented here, highlighting the wide interest in the quantitative study of Nobel laureates.Many studies have been devoted to building open-access datasets on scientific productivity and careers of scientists. For example, Vuong There have also been practical utilities for such datasets. For example, Clarivate Analytics has developed tools that use similar proprietary publication and citation databases to predict future Nobel laureates. While the focus of Clarivate Analytics’ work is on predicting future laureates, the goal of our paper is to collect a comprehensive dataset capturing careers of individual Nobel laureates over the past century, which could then serve as an empirical starting point for future quantitative studies.43. Despite a plethora of data capturing the various contributions of scientific elites, such information is often located in unstructured, isolated sources. Here, by combining unstructured information collected from Nobel Prize official websites, laureates’ personal and university websites, Wikipedia entries, and publication and citation records from the MAG, we constructed a unique dataset of career histories for nearly all Nobel laureates in Physics, Chemistry, and Physiology or Medicine from 1900 to 2016 . We validated this dataset using four different approaches to ensure the reliability of our results, including comparison with manually collected CVs, selected Google Scholar (GS) profiles, additional affiliation information, and random selection of 60 Nobel laureates (20 for each field) for manual verification. The total data collection and validation procedure took more than 1000 hours.Here we build an open-access dataset on the scientific careers of Nobel laureatesThe curated data could serve as critical input that feeds into several promising research directions. (1) The data make available quantitative patterns of productivity, collaboration, and impact governing the careers of scientific elites, offering a unique opportunity to systematically identify quantitative signals tracing the careers of elite scientists. (2) Combining our datasets with publication records that capture the careers of ordinary scientists offers opportunities for an array of fascinating comparative studies. Such studies would deepen our understanding of the factors driving exceptional scientific careers, helping us answer the broad question of what makes great scientists great.44. Briefly, we find that careers of Nobel laureates are characterized by remarkably similar patterns as those of ordinary scientists. For example, apart from the prize-winning paper, all other important works in Nobel careers closely follow the random impact rule11, a finding that is contrary to the common belief that Nobel laureates tend to do critical work early in their careers. Further, the laureates also show a tendency toward collaborative research in larger teams, which runs counter to the iconic image of lone geniuses making solo contributions.As a more concrete example, we present two new findings using the curated dataset in the associated commentaryThese results only represent some initial examples of how such datasets can help advance our quantitative understanding of career dynamics. Indeed, our dataset may help uncover a set of reproducible principles underlying individual creativity, offering insights into the conditions and environments that best facilitate scientific creativity and the genesis of scientific breakthroughs.We constructed the publication records for almost all Nobel laureates in physics, chemistry, and physiology or medicine from 1900 to 2016 . We first collected information manually from Nobel Prize official websites, their university websites, and Wikipedia. We then matched it algorithmically with big data, tracing publication records from the MAG database. Figure 45. The dataset is updated on a weekly basis and contains 174,910,379 papers, 210,983,748 authors, 228,843 fields of study, 4,028 conferences, 47,963 journals, and 25,558 institutions as of June 2018.The MAG data contains scientific publication records, interlinked through the citation relationships between them, together with information about authors, institutions, publication venues , and fields of studyThe Nobel Foundation’s website. The Nobel Foundation’s website (nobelprize.org) offers a rich source of information about Nobel laureates. It contains numerous site-accessible documents, photos, audiotapes, videotapes, films, and articles for each laureate.https://www.wikipedia.org/). All Nobel laureates have dedicated pages which offer detailed biographical information including biographies, scientific activities, selected publication records, awards, and honors, etc.Wikipedia pages. Wikipedia is a multilingual, web-based, free encyclopedia based on a model of openly editable and viewable content  The Nobel laureate is in the author list of the reference paper, i.e., the paper has at least one author with the same last name and first name  with the Nobel winner. For an author with a middle name or middle initial, such information must be consistent as well; 2) The paper was published within the same period of prize-winning papers; (3) Institution and co-author information of the reference is consistent with other auxiliary information about the prize-winning work; (4) The topic of the reference paper is consistent with the Nobel Prize motivation. If there are multiple papers that satisfy the criteria (1–4), we assume that prize-winning papers garner higher impact . Following these procedures, we manually identified and collected the prize-winning papers for all laureates in our sample , Chemistry (1998–2013), Medicine (2006–2013), and Economics (1995–2013). Our data collection also takes into account these existing efforts.In rare cases where prize-winning papers cannot be reliably identified from the references of the Nobel lectures, we consulted other sources: (1) Nobel laureates’ Wikipedia page usually offers a detailed personal biography, which contains their prize-winning contributions and mentions of prize-winning papers. (2) Important works are usually selected by the Nobel laureates and displayed on their personal homepage or lab website, which can help us to identify the prize-winning papers. (3) Existing studies have collected some of the prize-winning papers. For example, Shen and BarabasiWe matched all prize-winning papers with the MAG database. Together, we gathered 874 prize-winning papers for 545 Nobel laureates, including 283 prize-winning papers for 193 laureates in Physics, 259 prize-winning papers for 163 laureates in Chemistry, and 332 prize-winning papers for 189 laureates in Medicine.48. This seemingly simple task represents a major unsolved problem for information and computer sciences, and a major roadblock to studies of individual careers. Here we exploit one important feature of the MAG data. Indeed, one major advantage of the MAG dataset is that author profiles have been processed through a well-designed disambiguation algorithm that optimizes the accuracy of a profile49. That is, each disambiguated profile in MAG may not contain all the papers published by an individual, but papers that are included in the profile do belong to the same person with high accuracy. In other words, MAG may split one scientist’s publication record into several different profiles, optimizing accuracy at the expense of recall. Note that authors themselves can also claim and assemble different profiles into one (https://www.microsoft.com/en-us/research/project/academic/articles/microsoft-academic-uses-knowledge-address-problem-conflation-disambiguation/), offering additional crowd intelligence to assist in the disambiguation procedure. This also suggests that if we could intelligently combine these profiles, we may be able to curate individual profiles with both high precision and recall.A key challenge in analyzing scholarly databases is to identify the individual(s) who wrote a paper and, conversely, to identify all of the works that belong to a given individual50, a method similar to label propagation in machine learning51 to merge various MAG author profiles into one. Figure Manually collecting laureates’ papers. As part of identifying prize-winning papers, we have collected and identified several of the laureates’ papers, including the prize-winning papers, Nobel lecture papers, and other papers collected from their Wikipedia page, the Nobel official website, and their own homepage. These manually collected laureates’ papers are referred to as ground-truth papers.Matching laureates’ papers into the MAG dataset as seeds. We then match these collected laureates’ papers into the MAG dataset using the following rules: Two papers are identical if they share the same Digital Object Identifier (DOI). In cases where the DOI is missing, they are considered to be the same paper if the following matching rules are satisfied: (1) the two papers were published within ±1 years; (2) the two papers have the same number of authors; (3) author sequence of the two papers is the same; (4) the text similarity, defined as the cosine similarity between the titles of the paper after removing stop words and punctuation, is higher than 0.75.11: (1) the author’s last name is identical to the laureate’s last name; (2) initial of the first name is identical to the laureate’s first initial. If an author’s full first name is available, it must be the same as the laureate’s; (3) for those authors who have middle names, the middle initial must be the same as the laureate’s. If the author’s full middle name is available, it also must be consistent with the laureate’s middle name. (4) For each laureate, all the publication records under the potential MAG author’s profiles constitute the candidate pool of papers for the laureate.Creating a pool of all candidate MAG profiles for each laureate. We then created a pool of all candidate MAG profiles for each laureate according to the same rules followed by the literatureBuilding a citation network within the candidate paper pool through the “Seed + Expand” process. One assumption for the “Seed + Expand” process is that scientists are more likely to cite their own published work due to topical relevance or intellectual similarity between the papers. Thus, their own papers are usually connected through a citation network. Starting with the seed papers, we take candidate papers that have cited at least one of the seed papers and expand the pool of seeds iteratively. The process stops when there are no papers left in the candidate paper pool to be added to the network Fig. .Merging candidate MAG author profiles. For each Nobel laureate, we have a pool of candidate MAG authors. We then merge all the candidate MAG author profiles that have at least one paper in the citation network. Combining the merged MAG author’s profiles yields the entire publication history for each laureate.In this paper, we adopt a “Seed + Expand” procedureTogether, we curated publication records for each Nobel laureate, totaling 93,394 journal papers for 545 Nobel laureates.The manually collected ground-truth papers play an important role in the name disambiguation procedure. These ground-truth papers are matched into the MAG dataset as seeds, which are the starting point of the “Seed + Expand” process. Here we conducted cross-validation with additional information such as co-authors, affiliations, timing, a paper’s keywords, and even e-mail information, if available, to double check each manually collected laureates’ paper. We did a lot of tinkering to ensure the accuracy and correctness of the collected papers, trying to minimize the human errors accounted for in the “Seed + Expand” procedure. Nevertheless, although all these steps help reduce any potential human errors in the data curation process, thereby ensuring the accuracy of the collected data, readers should take note of the possibility that there may still be errors that remain unaccounted for given the manual processes.43. It contains four comma-separated values (CSV) files named “Prize-winning paper record,” “Physics publication record,”We built an open-access dataset on publication records for Nobel laureates in Physics, Chemistry, and Medicine, which is available at Harvard Dataverse“Chemistry publication record,” and “Medicine publication record.” The details are further described in Table https://www.nobelprize.org/prizes/medicine/1923/macleod/facts/); the experiment that resulted in the discovery was mainly conducted by Frederick Banting and Charles Best in John MacLeod’s laboratory in 1921 and was first published in the February 1922 issue of The Journal of Laboratory and Clinical Medicine under the names Frederick Banting and Charles Best; Macleod declined co-authorship because he considered it Banting’s and Best’s work (https://en.wikipedia.org/wiki/John_Macleod_(physiologist)).“Prize-winning paper record” contains information about prize-winning papers, including “Field,” “Laureate ID,” “Laureate name,” “Prize year,” “Title,” “Pub year,” “Paper ID,” and “Additional information” Table . “Field”The three files named “Physics publication record,” “Chemistry publication record,” and “Medicine publication record” contain the publication records of Nobel laureates for Physics, Chemistry, and Medicine, respectively. Each file includes the following data fields Table : “LaureaTable To understand the reliability of the curated dataset, we take the following four different approaches to validate the data.We manually collected 30 laureates’ CVs from their personal websites, lab homepages, or university homepages which contain their full list of publications. Compared against these publication histories as the gold standard, our data shows a high precision and recall at 82.3% and 92.2% respectively.Google provides scholar profiles for scientists to create, maintain and update their own publication records. Armed with Google’s proprietary name disambiguation algorithms, Google Scholar profiles may represent a comprehensive collection of individual publication histories. Keep in mind, however, that GS profiles tend to cover currently active scientists, and only a modest fraction of Nobel laureates have their own GS profiles. We collected GS publication records for 29 Nobel laureates. Since GS indexes not only journal publications but also conferences, patents, reports, meeting abstracts, talks, reviews, and even slides, here we consider only journal publications. Compared against the 29 GS profiles, our data shows precision and recall of 87.2% and 84.2% respectively.More than 80% of publications we collected contain author affiliations, allowing us to use additional affiliation information to verify the reliability of our data. For each Nobel laureate, we first collected his/her affiliations from his/her Wikipedia and University homepage. Then we went through the curated publication list to check if the affiliation information recorded in the paper was consistent with the laureate’s career history. We find our data has high accuracy (97.6%).https://academic.microsoft.com/). We randomly selected 60 Nobel laureates (20 for each field) to manually check the reliability of our data.The MAG matched the biography information for a selected fraction of Nobel laureates from the Wikipedia to the author profiles. We can access this information from the MAG API  can be fulfilled automatically with established procedures and rules. The highly automated process of our method guarantees the reproducibility of the dataset and method.Together, we present a novel systematic dataset recording career information of Nobel laureates in science. Although the curated data offers, to our knowledge, one of the most comprehensive publication records of Nobel laureates so far, the goal of publishing this dataset is to allow all interested readers to help further refine and improve the quality of the data.Download metadata file


Allograft interstitial fibrosis was characterized by massive extracellular matrix deposition caused by activated fibroblasts and myofibroblasts. Epithelial‐mesenchymal transition (EMT) is recognized as an important source of myofibroblasts contributing to the pathogenesis of allograft interstitial fibrosis. Smad ubiquitination regulatory factor 1 (Smurf1) has been recently reported to be involved in the progression of EMT. Our study was to detect the effect of Bortezomib and Smurf1 in the EMT and allograft interstitial fibrosis. Biomarkers of EMT, as well as Smurf1, were examined in human proximal tubular epithelial cells (HK‐2) treated with tumour necrosis factor‐alpha (TNF‐α) in various doses or at various time points by Western Blotting or qRT‐PCR. We knockdown or overexpressed Smurf1 in HK‐2 cells. Furthermore, rat renal transplant model was established and intervened by Bortezomib. Allograft tissues from human and rats were also collected and prepared for HE, Masson's trichrome, immunohistochemical staining and western blotting assays. As a result, we found that TNF‐α significantly promoted the development of EMT in a time‐dependent and dose‐dependent manner through Smurf1/Akt/mTOR/P70S6K signalling pathway. More importantly, Bortezomib alleviated the progression of EMT and allograft interstitial fibrosis in vivo and in vitro by inhibiting the production of TNF‐α and expression of Smurf1. In conclusion, Smurf1 plays a critical role in the development of EMT induced by TNF‐α. Bortezomib can attenuate the Sumrf1‐mediated progression of EMT and renal allograft interstitial fibrosis, which could be suggested as a novel choice for the prevention and treatment of renal allograft interstitial fibrosis. The study protocol involving human kidney tissues was approved by the local ethics committee of the First Affiliated Hospital of Nanjing Medical University. Written informed consent was obtained from all transplant recipients and nephrectomy patients.2.2Allograft kidney sections were obtained from 30 patients, which were collected from transplanted kidney nephrectomy or kidney biopsy of recipients. All patients, who had undergone kidney transplantation at our centre from January 2001 to October 2017 was diagnosed with CAD according to their allograft biopsy results. Patients diagnosed with CAD was classified as CAD group. In addition, 25 normal kidney samples obtained from more than 5 cm away from the tumour tissues were collected from radical nephrectomy and considered as control group. The baseline characteristics of patients in the CAD group and control group are given in Table 2.3Adult male F344 and Lewis rats (200‐250 g) were procured from Charles River Laboratories . The animal centre provided the rats with the clean tap water and standard rat chow. Animal handling procedures were in accordance with guidelines published by the US National Institutes of Health, and animal ethics established by Nanjing Medical University.2.4Orthotopic left kidney transplantation was performed between F344 and Lewis rats as previously described.2.5iv injected to isogeneic or allogeneic recipient rats twice a week.From the first day after the kidney transplantation, Bortezomib  was At 4 weeks, 8 weeks, 12 weeks and 16 weeks, organs were harvested and divided into two parts, which was fixed in paraffin or snap‐frozen in N2 and stored at −80°C.2.6The levels of rat serum TNF‐α were quantified by the rat TNF‐α ELISA kit . Rat serum and cell culture medium supernatant level of TGF‐β1 were quantified by the TGF‐β1 ELISA kit . The assays were performed as described in the manufacturer's instruction.2.7Histological analysis was performed by using haematoxylin and eosin (H&E) and Masson trichome staining. H&E and Masson trichome staining were performed as detailed elsewhere, which was used to evaluate the severity of chronic kidney rejection and the area of renal interstitial fibrosis separately.2.8Immunohistochemical staining assays were performed to evaluate the expression and distribution of TNF‐α, α‐SMA, Smurf1, Fibronectin, E‐cadherin, collagen I. Paraffin sections (4 μm) were deparaffinized and rehydrated. Then, antigen retrieval was performed before primary antibody incubation, including suppression of endogenous peroxidase activity and 10% skim milk blocking. Kidney sections were incubated overnight with anti‐collagen I , anti‐Smurf1 , anti‐ TGF‐β1 , anti‐ TNF‐α , anti‐α‐SMA , anti‐Fibronectin  and anti‐E‐cadherin  primary antibodies at 4°C. After that, slices were incubated with biotinylated goat anti‐mouse/rabbit IgG  for 1 hr Digital images of immunohistochemistry staining were captured and analysed by two authors independently with the help of a light microscope .2.9Immunofluorescence double‐staining assay were performed as previously described, using antibodies against TGF‐β1 , α‐SMA , E‐cadherin .2.10Concentrations of rat blood creatinine and urea nitrogen were tested by instructions of manufacturer of the kits .2.11Total RNA was extracted from cells with the RNA extraction kits . cDNA was synthesized with a PrimeScriptTMRT reagent kit . qRT‐PCR was performed with a SYBR Green PCR kit (TaKaRa Biotechnology) on a DNA Engine Opticon 2 System . The specific primers used were as follows:Smurf1: 5′‐ CTACCAGCGTTTGGATCTAT‐3′ (F)5′‐ TGTCTCGGGTCTGTAAACT‐3′ (R);CDH1: 5′‐CGAGAGCTACACGTTCACGG‐3′ (F)5′‐GGGTGTCGAGGGAAAAATAGG‐3′(R);ACTA2: 5′‐AAAAGACAGCTACGTGGGTGA‐3′ (F)5′‐GCCATGTTCTATCGGGTACTTC‐3′(R);Fibronectin1: 5′‐ CGGTGGCTGTCAGTCAAAG‐3′(F)5′‐AAACCTCGGCTTCCTCCATAA‐3′(R);Actin: 5′‐TGACGTGGACATCCGCAAAG‐3′ (F)5′‐CTGGAAGGTGGACAGCGAGG‐3′ (R).mRNA expression was normalized to b‐actin expression. Every experiment described previously was repeated at least three times.2.12Western blotting was performed according to the protocol described by Liu et al2.13HK‐2s were cultured in Dulbecco's modified Eagle's medium (DMEM)/F12 medium containing 10% foetal bovine serum and 1% penicillin‐streptomycin in a humidified atmosphere containing 5% CO2 at 37°C. For TNF‐α treatment, the cells were starved without serum overnight and then treated with 50 ng/mL TNF‐α for 0, 0.5, 1, 2, 4, 6, 8, 12, 24, 48 hours or 10, 20, 50, 100 ng/mL TNF‐α for 24 hours. MK‐2206 (10 μM) was previously given to HK‐2 cells for 0.5 hour before the treatment of TNF‐α, in order to determine the effects of Akt pathway on the Smurf1‐meditated EMT. To evaluate the anti‐fibrogenic effect, cells were divided into five groups and then starved without serum overnight. Four of them were co‐treated with TNF‐α (50 ng/mL) and Bortezomib  for 24 hours.Construction and verification of stable Smurf1‐knockdown and Smurf1‐overexpression cell lines:HK‐2 cells were inoculated into 12‐well plates, then added with 1.5 μL Smurf1 short hairpin ribonucleic acid (shRNA) viruses or 2.0 μL Smurf1 overexpressing viruses. Control viruses were added at the same time. The cells were further cultured with medium containing 2 μL polybrene for 6 hours. After that, medium was removed and cells were cultured with non‐polybrene medium until 48 hours. The infection rate of viruses was observed under a fluorescence microscope , and puromycin was added to kill the uninfected cells. The expression level of Smurf1 protein was determined by western blotting.2.14t test. Values of P < 0.05 were considered statistically significant. All the assays were repeated at least three times independently.All data were presented as the mean ± SD values determined from three independent experiments. After demonstration of homogeneity of variance with Bartlett test, inter‐group comparisons were made using one‐way analysis of variance (ANOVA). Multiple means were compared using Tukey's test. The differences between two groups were determined by Student 33.1In this study, we used human specimens to confirm the high expression of Smurf1 in the CAD patients with allograft renal interstitial fibrosis. Histological studies with HE and Masson staining revealed significant renal interstitial fibrosis from CAD group (n = 30) to control group (n = 25)  for 24 hours. TNF‐α up‐regulated the protein and mRNA expression of fibronectin and α‐SMA, leading to a dose‐dependent decline in the expression of E‐cadherin at the same time Figure A‐C. Prot3.3To confirm the role of Smurf1 in the EMT induced by TNF‐α, we examined the effect of Smurf1 silencing on these events by constructing Smurf1 stable knockdown HK‐2 cells. As shown in the Figure 3.4To evaluate the signalling pathways involved in TNF‐α‐induced EMT in HK‐2 cells, we examined phosphorylation of factors involved in the canonical signalling pathways by western blotting. We observed the activation of the Akt‐mTOR‐P70S6K signalling pathways when HK‐2 cells were treated with TNF‐α Figure A. The Ak3.5In this study, we applied Bortezomib to explore the role of the proteasome inhibitor in ameliorating renal interstitial fibrosis in allograft kidney after transplantation. Rat renal transplant model was established by transplantation from F344 rat donors to Lewis rat recipients. 0.2 mg/kg Bortezomib was injected to the allo‐recipient rats through tail vein every three days. After 4 weeks of transplantation, allograft kidney was infiltrated with inflammatory cells and the lesion area was up to almost 80% according to the results of HE staining Figure A, 5C. Bo3.6The outcome of immunohistochemistry assay revealed the remarkably high expression of TNF‐α and Smurf1 in allo‐recipient group Figure A. After 3.7In order to test the impact of Bortezomib on allograft renal function and long‐term survival of allogeneic recipients, the native right kidneys of recipients were removed 2 weeks after left kidney transplantation. One week after the resection of right kidney, the renal function including the blood concentration of creatinine and urea nitrogen, started to be continuously deteriorated until their death. However, the increase of blood creatinine and urea nitrogen was inhibited and they remained at a relatively low level by the treatment of Bortezomib Figure A‐B. The 4In present study, we reported that Bortezomib could attenuate the fibrosis of renal interstitial in kidney transplantation via regulating the EMT induced by TNF‐α‐Smurf1‐Akt‐mTOR‐P70S6K pathway. The effects of Bortezomib are not only to block the progression of the EMT induced by TNF‐α‐Smurf1‐Akt‐mTOR‐P70S6K, but also to reduce the secretion of TNF‐α. To our best knowledge, it is the first study that elucidate the role and underlying mechanism of Bortezomib, which could attenuate the fibrosis of renal interstitial in kidney transplantation. Smurf1, as a key of the EMT and renal interstitial fibrosis induced by TNF‐α, plays a critical role in this progression. Our results offer novel insights into the treatment and prevention of allograft renal interstitial fibrosis. Bortezomib could be a new way in the treatment and prevention of transplant renal interstitial fibrosis. Furthermore, Smurf1 could provide us with a new drug target of the treatment and prevention of transplant kidney interstitial fibrosis.TGF‐β1 has been widely used in the induction of EMT in HK‐2 cells. Our previous research confirmed TNF‐α also could promote the progression of EMT in HK‐2 cells. Immunohistochemistry staining of TGF‐β1 revealed the expression of TGF‐β1 in CAD group was remarkably higher than Control group Figure A‐B. Bortvitro findings revealed that the progression of fibrosis and EMT of HK‐2 was induced by the TNF‐α‐Smurf1‐Akt‐mTOR‐P70S6K pathway. In our vitro study, overexpression of Smurf1 without the stimulation of TNF‐α could not lead to the progression of EMT in HK‐2 cells. Unlike TNF‐α and TGF‐β1，Smurf1 served as a regulatory factor rather than inducer of the EMT progression in HK‐2 cells.In order to determine the relationship between TNF‐α, Smurf1 and EMT in the transplant kidneys, our Furthermore, we found the anti‐fibrogenic effects of Bortezomib, which alleviated the fibrosis and EMT of HK‐2 cells by downregulating the expression of Smurf1. Bortezomib was reported that it prevents oncogenesis and bone metastasis of prostate cancer by inhibiting Smurf1 expression.vivo findings revealed that Bortezomib could reduce the inflammatory response and renal interstitial fibrosis in allograft kidney. Bortezomib blocked the progression of EMT induced by TNF‐α‐Smurf1‐Akt‐mTOR‐P70S6K. Another positive effect of Bortezomib was to reduce the secretion of TNF‐α. The treatment of Bortezomib could prolong the survival of allo‐recipients. Bortezomib, as a proteasome inhibitor, was used to prevent the production of donor specific antibody in treatment of ABMR. The anti‐fibrogenic effect and mechanism of Bortezomib was never reported in transplant kidney. So Bortezomib could be a potential therapeutic drug in prevention and treatment of transplant kidney fibrosis.To confirm the anti‐fibrogenic effect of Bortezomib, our Taken together, the current studies proved that Bortezomib attenuates the fibrosis of renal interstitial and Smurf1‐meditated EMT induced by TNF‐α‐Akt‐mTOR‐P70S6K pathway. The results of our study provide novel insight into Smurf1 and Bortezomib. Smurf1 could be an additional target for the treatment and prevention of kidney interstitial fibrosis and CAD in kidney transplant recipients. Bortezomib can attenuate the Sumrf1‐mediated progression of EMT and renal allograft interstitial fibrosis, which could be suggested as a novel choice for the prevention and treatment of renal allograft interstitial fibrosis.The authors confirm that there are no conflicts of interest.Study design: M Gu and RY Tan; Data Collection: JJ Zhou, H Cheng, ZJ Wang and H Chen; Software material: H Cheng, CJ Suo; Data Analysis: JY Zhang, HC Zhang，YH Yang and L Geng; Manuscript preparation: JJ Zhou, H Cheng and ZJ Wang. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file.


Stenosis or obstruction of the celiac artery (CA) is known as celiac artery stenosis (CAS) and is usually accompanied by the formation of arterial anastomosis between the superior mesenteric artery (SMA) system and the CA system. Arterial bypass is mainly achieved through the gastroduodenal artery (GDA); therefore, the division of the GDA during pancreaticoduodenectomy (PD) could pose a problem in patients with CAS.We reported a case of PD presenting complete occlusion of the CA, in which perfusion to organs in the CA system was maintained via peri-pancreatic arterial communication. There were complicated arterial anastomoses around the pancreas, which were clearly visualized on a three-dimensional reconstruction of the arterial system using multi-detector computed tomography. Among these complicated anastomoses, one well-developed anastomosis between the SMA and the splenic artery through the dorsal pancreatic artery (DPA) was identified. The DPA was considered to work as a potential collateral pathway from the SMA to organs in the CA system after division of the GDA. During surgery, Doppler ultrasonography detected hepatopetal arterial flow even after the GDA clamping; therefore, we performed typical PD with division of the GDA. The postoperative course of the patient was uneventful, and there was no sign of ischemic complications in the CA system organs including the liver, stomach or spleen.Three-dimensional reconstruction of the arterial system using multi-detector computed tomography and the intraoperative GDA clamping test were useful to determine whether it was possible to divide the GDA in PD, in the case of CAS. The celiac artery (CA) is the first major branch of the abdominal aorta supplying blood to the liver, stomach, spleen, duodenum, and pancreas. Stenosis or obstruction of the CA, known as celiac artery stenosis (CAS), can result from various factors, including median arcuate ligament syndrome (MALS), atherosclerosis, and invasion of malignancies or pancreatitis . HoweverPancreaticoduodenectomy (PD) is a surgical procedure to resect pancreatic head tumor, including pancreatic adenocarcinoma, distal bile duct cancer, and ampullary carcinoma . The proAn 80-year-old man with epigastric pain visited our hospital in January 2019. A magnetic resonance cholangiopancreatography revealed several stones in the common bile duct (CBD), and he was diagnosed with choledocholithiasis. Endoscopic lithotripsy was performed, and cholangiography showed narrowing and wall-irregularity of the lower CBD Fig. a. A 3D iPD was performed in April 2019. After taping of the GDA Fig. , CHA, anPostoperative laboratory data showed slight elevation of hepatic enzyme immediately after the surgery Fig. , but we CAS is a relatively common finding, identified in 12.5–24% of patients undergoing abdominal angiography . The typSevere stenosis or obstruction of the CA often induces the formation of collateral arterial pathways from the SMA system to the CA system to compensate the decrease of splanchnic arterial flow of the upper abdomen. The main collateral pathway is reported to be the GDA or DPA in most cases, whereas various arteries, such as the DPA, jejunal artery, omental artery, left gastric artery, arc of Buhlar, and Riolan or Drummond artery can be a collateral pathway , 5. SongIn the context of CAS, the formation of collateral arterial pathways is surgically relevant for arterial division in PD. Although the GDA is the main collateral pathway from the SMA system to the CA system in most of the cases of CAS, the GDA must be divided in PD. Therefore, various procedures were attempted to avoid organ ischemia after PD in patients with CAS . There aIn conclusion, we described a case of PD with complete occlusion of the CA, in which we maintained perfusion via the peri-pancreatic arterial communication. A 3D reconstruction of the arterial system from MDCT and intraoperative GDA clamping test were useful to determine whether it was possible to divide the GDA.


Tacrolimus, a calcineurin (CaN) inhibitor, has been used for treatment of refractory allergic ocular disease, although its role in optic nerve degeneration remains to be elucidated. In this study, we investigated whether tacrolimus modulates tumor necrosis factor (TNF)-mediated axonal degeneration and whether it alters nuclear factor of activated T cells (NFATc), a downstream effector of CaN signaling. Immunoblot analysis showed no significant difference in CaNAα protein levels in optic nerve on day 3, 7, or 14 after TNF injection compared with PBS injection. However, a significant increase in NFATc1 protein level was observed in optic nerve 7 days after TNF injection. This increase was negated by simultaneous administration of tacrolimus. Administration of tacrolimus alone did not change the NFATc1 protein level in comparison to that observed after PBS injection. A significant increase in TNF protein level was observed in optic nerve 14 days after TNF injection and this increase was prevented by tacrolimus. Immunohistochemical analysis showed the immunoreactivity of NFATc1 to be increased in optic nerve after TNF injection. This increased immunoreactivity was colocalized with glial fibrillary acidic protein and was suppressed by tacrolimus. Treatment of tacrolimus significantly ameliorated the TNF-mediated axonal loss. These results suggest that tacrolimus is neuroprotective against axon loss in TNF-induced optic neuropathy and that the effect arises from suppression of the CaN/NFATc1 pathway.The online version of this article (10.1007/s11064-019-02804-6) contains supplementary material, which is available to authorized users. Glaucoma is, in developed countries, one of the most common diseases that can lead to irreversible blindness. It is characterized by a slowly progressive retinal ganglion cell (RGC) pathology in which axonal degeneration of RGCs in the optic nerve precedes death of the RGC body. The only reliable clinical treatment for glaucoma is lowering intraocular pressure. However, means of neuroprotection are being studied as a complementary treatments.2+/calmodulin (CaM). It is composed of two subunits, CaNA and CaNB. CaNA which has a CaM-binding domain, acts as a 60-kDa catalytic subunit. CaNB, a 19-kDa regulatory subunit, has a Ca2+-binding domain [Calcineurin (CaN) is a serine/threonine phosphatase, dependent on Cag domain , 4.2+ activates CaN, and the CaN combines with the Ca2+/CaM, leading to dephosphorylation of nuclear factor of activated T cells (NFATc) in the cytoplasm. Dephosphorylated NFATc translocates to the nucleus to regulate the transcription of cytokines such as interleukin (IL)-2 and tumor necrosis factor (TNF) [2+/CaM-dependent NFATc transcription factors: NFATc1 (NFAT2), NFATc2 (NFAT1), NFATc3 (NFAT4), and NFATc4 (NFAT3) [Increased intracellular Caor (TNF) , 6. Ther (NFAT3) . All are (NFAT3) . Increas (NFAT3) Tacrolimus, a CaN inhibitor, forms a complex with immunophilin (FKBP) and inhibits CaN enzyme activity . TacroliTNF is a cytokine that is synthesized and released from microglia and astrocytes around several neurons. An association between TNF-related neurotoxicity and glaucomatous optic neuropathy has been reported . For exaFor experiments, male Wistar rats of 50 to 55 days old were used. All experiments were conducted in accordance with statements of the Association for Research in Vision and Ophthalmology statement on the Use of Animals in Ophthalmic and Vision Research. Animals were kept in a controlled condition with a temperature of 23 ± 1 °C, humidity of 55 ± 5%, and light between 6 am and 6 pm.Intravitreal injection of TNF  or tacrolimus  was performed as previously described . Brieflyg for 15 min at 4 °C after 3, 7 or 14 days after the intravitreal injections. Optic nerve specimens (n = 2) were collected for each sample. Each retina was used as one sample. Protein concentrations were measured with the Bio-Rad Protein Assay kit . Protein samples  were subjected to SDS-PAGE on 10% polyacrylamide gels and transferred to an enhanced chemiluminescent membrane . Tris buffered saline-0.1% Tween-20 (T-TBS) containing 5% skim milk was used for blocking of membranes. Membranes were reacted with CaNAα antibody , NFATc1 antibody , TNF antibody  or anti-β-actin antibody  in T-TBS. The membranes were then reacted to peroxidase-labeled anti-rabbit secondary antibody , peroxidase-labeled anti-goat secondary antibody  or peroxidase-labeled anti-mouse secondary antibody  diluted 1:5000 in T-TBS. Immunoblots were detected by a chemiluminescence membrane detection system .One hundred and two rats were used for immunoblot analysis as described previously . BrieflyTwelve rats were used for immunohistochemical analysis as described previously . After i2 each . An average of the number of axons per eye was calculated, and this average value was calculated as a number per square millimeter. For each experimental condition at least 5 eyes were used for the analysis.Morphometric analysis of each optic nerve in samples from 23 rats was performed as described previously . Two weep value) of < 0.05 was considered statistically significant.Data are presented as mean ± SEM. Differences between groups were analyzed by one-way ANOVA and then Dunnett’s test. A probability value . Although treatment of 10−6 M tacrolimus had a certain axonal protective effect . However, treatment with 10−5 M tacrolimus . These protective effects were shown to appear in a concentration-dependent manner.We confirmed degeneration in the optic nerve axons 14 days after TNF injection Fig. b compareect Fig. c, the neThe present study demonstrated a significant increase in NFATc1 expression in optic nerves after TNF injection. NFATc1 is present in glial cells in optic nerve. Tacrolimus prevented the increase in NFATc1 expression and ameliorated the loss of axons of the optic nerve after TNF injection. These results suggest that the protective effect of tacrolimus may be involved in its inhibitory effect on NFATc1 expression.2+, CaNA activity is reversible. However, under a pathophysiological condition such as kainate- or glutamate-induced excitotoxicity [2+ influx, CaNA activity is lasting. Calpain, a Ca/CaM dependent protein, puts CaN into a constitutively activated state by cleaving the self-suppressing region of CaNA [2+ concentration is limited in the TNF model and that the increase is not enough to occur CaN cleavage or alter the CaN protein level. Nonetheless, we cannot deny the possibility of CaN activation in the TNF model because lack of agreement between the protein levels and the enzyme activity of CaN has been reported [In a previously reported study, cleaved CaN was found in retinal protein in the rat IOP elevation model but not in retinal protein in the optic nerve crush model . In our toxicity , which creported .2+ may result in CaN/NFAT activation, thereby increasing in TNF in certain cell death. Consistent with this idea, the present study demonstrated that increased NFATc1 protein level was observed 7 days and increased TNF protein level was observed 14 days after TNF injection, suggesting that NFATc1 exists upstream of TNF. It was reported that NFATc1β, an isoform of NFATc1, upregulated TNF production in NIH3T3 cells [Recent studies have reported NFATc3 expression in human lamina cribrosa cells  and NFATT3 cells . On the T3 cells .In the present study, we found that tacrolimus significantly prevented the TNF-induced increase in NFATc1 expression in optic nerve. Morphometric analysis revealed that tacrolimus significantly prevented TNF-induced axon loss. These findings are consistent with those of a previously reported study demonstrating that tacrolimus attenuates the activation of astrocytes and microglia of brain in a rat cerebral ischemia model . That stIn conclusion, tacrolimus may protect against axon loss in TNF-induced optic neuropathy by inhibiting the CaN/NFATc1 pathway.Supplementary material 1 (TIFF 2709 kb)Supplementary material 2 (TIFF 5688 kb)Below is the link to the electronic supplementary material.


Heart failure (HF) represents a major epidemic with high morbidity and mortality rates, imposing a significant burden on healthcare systems worldwide  in HFpEF development. LVDD has long been considered the major causative factor of HFpEF; however, many previous trials aiming to reduce long-term mortality by enhancing diastolic function, namely by improving LV relaxation and/or halting the progression of LV hypertrophy through the antagonism of the renin-angiotensin-aldosterone system, have failed to report favorable outcomes , cardiac computerized tomography, and magnetic resonance imaging Butler, . MoreoveIn addition to network analysis, another method that has more recently been implemented to assess the underlying heterogeneity in HF, amongst various other diseases, is “phenomapping,” which involves machine-based learning to analyze large sums of phenotypic data in order to categorize patients into distinct subgroups based on a select number of clinical features (Katz et al., The three classes of HF are all characterized by distinct pathophysiological processes, which in turn contribute to the heterogeneity in the expressed phenotype. Whilst there is currently less unknown about HFrEF, literature pertaining to HFpEF and the newly classified HFmrEF still incites many questions with respect to pathogenesis and optimal therapeutic strategies. Specifically, more prospective studies, as opposed to cross-sectional studies, that follow HFpEF and HFmrEF patients for a lengthy time period are required to assess whether or not the development of HF is truly, in fact, a continuum that progresses from HFpEF through to HFmrEF and eventually HFrEF. Overall, the diversity in the observed clinical features, namely of HFpEF and HFmrEF, poses challenges to the creation of a universal treatment regimen that will fit all patients. As such, future management of these two conditions, in particular, will likely necessitate the implementation of a patient-specific method that targets unique metabolic derangements present in distinct patient subgroups. Although tools such as network analysis and machine-based learning that attempt to collate and find links between available data have proven to yield informative results, a greater understanding of the underlying mechanisms involved in the development of HFpEF and HFmrEF is still required before this individualistic approach can be employed holistically in the clinical setting.All authors listed have made a substantial, direct and intellectual contribution to the work, and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Children with juvenile idiopathic arthritis (JIA) have impaired physical abilities. TNF-α plays a crucial role in this pathogenesis, but it is also involved in the use of lipids and muscle health. Objective of this study was to explore substrate oxidation and impact of TNF blockade on energy metabolism in children with JIA as compared to healthy children.Fifteen non-TNF-blockaded and 15 TNF-blockaded children with JIA and 15 healthy controls were matched by sex, age, and Tanner stage. Participants completed a submaximal incremental exercise test on ergocycle to determine fat and carbohydrate oxidation rates by indirect calorimetry.− 1) when compared to the controls ; but was higher in JIA children under TNF blockade  when compared to JIA children untreated by TNF blockade. At the same relative exercise intensities, there was no difference in carbohydrate oxidation rate between three groups.The maximal fat oxidation rate during exercise was lower in JIA children untreated by TNF blockade (134.3 ± 45.2 mg.minLipid metabolism during exercise was found to be impaired in children with JIA. However, TNF treatment seems to improve the fat oxidation rate in this population.NCT02977416, registered on 30 November 2016.In ClinicalTrials.gov, reference number  Children with juvenile idiopathic arthritis (JIA) have impaired aerobic and anaerobic capacities  and reduTumor necrosis factor alpha (TNF-α) has a crucial role in the pathogenesis of JIA, inducing expression of other pro-inflammatory cytokines such as interleukin (IL)-1β, IL-6 and IL-8, leading to an overall inflammatory response. But is also involved in tissue insulin resistance lipid utilization and muscular atrophy . FurtherThe two main energy sources oxidized during aerobic exercise are fats and carbohydrates (CHOs), and their relative use depends on the exercise modalities, primarily the intensity of exercise , but alsHence, compared to healthy children, children with JIA could be characterized as having impaired oxidation of energy substrates during submaximal exercise due to a reduction in lipid oxidation. Therefore, this study compares substrate oxidation during exercise in children with JIA treated with a TNF blockade and those not treated with a TNF blockade versus healthy controls to determine: whether the pathology and TNF treatment affect these metabolic responses during submaximal exercise.Thirty children (15 non-TNF blockade and 15 under TNF blockade medication) aged 7 to 18 years old with JIA  and 15 healthy controls were enrolled in the study. Participants in the three groups were matched by age, pubertal stage, and gender. All participants were followed between January 2017 and June 2019 at the pediatric unit of the Clermont-Ferrand University Hospital in France. We firstly included patients treated by TNF blockade, then we included matched JIA patients without TNF blockade and healthy controls. Healthy controls were recruited from the entourage  of the patients.Participants were excluded if they had a physician-diagnosed infection, had received oral corticosteroids within the previous 3 months, had chronic disease (other than JIA for patients). All treatments had been administered for at least 3 months at the time of evaluation. Disease status (active or inactive) was evaluated according to the American College of Rheumatology criteria . The phyThe study was approved by the governing ethics committee . All participants and their parents gave their informed consent and were free to withdraw from the study at any time.2peak was considered to have been reached when the RER was ≥1.05 and the participant achieved his or her age-predicted maximal heart rate (HRmax: 220 − age), according to the methodology validated by Riddell et al. [Participants were asked to refrain from consuming any food or liquid, with the exception of water, 3 h before the visit. Participants were in minimal fasting state of 3 h. They also avoided high fat food and refrained from strenuous physical activity for at least 24 h before the visit. Participants arrived the hospital laboratory for indirect calorimetry test in the morning. After sitting quietly for 20 min, participants performed a graded exercise test to volitional fatigue on an electromagnetically-braked cycle ergometer with continuous gas collection and heart rate monitoring. Following a 2-min warm-up involving unloaded pedaling, Tanner stages 1 and 2 participants started cycling at 10 W and the work rate was increased by 10 W every 3 min. Tanner stages 3 and 4 participants started at 20 W and the work rate was increased by 15 W every 3 min. In cases where the heart rate was unstable, this stage was extended for up to 5 min to obtain a stable heart rate of ±5 beats. When the respiratory exchange ratio (RER) was ≥1.00—indicating the absence of fat oxidation—, the work rate was increased by the same increments at 1-min intervals until volitional fatigue was reached. The VOl et al. .2 consumption (VO2) and CO2 production (VCO2) were measured breath-by-breath through a mask connected to an O2 and CO2 analyzer .All the tests were performed on a Cyclus 2 ergometer . The OVentilatory parameters were averaged every 1 min during the submaximal exercise test and during the subsequent 10-min recovery period. The heart rate was continuously monitored over the duration of the tests .2 and VCO2 values were averaged over the last minute of each work rate, with the results then being used to calculate fat oxidation over a wide range of exercise intensities for each participant [− 1) = 1.6946 x VO2–1.7012 x VCO2 and CHO (g.min− 1) = 4.585 VCO2–3.2255 VO2.Indirect calorimetry is known to be the standard method to quantify substrate oxidation rates at rest and during exercise . The VO2ticipant  by emploticipant . Lipids − 1) vs. exercise intensity (expressed as % VO2peak). Each individual curve was then used to determine the peak fat oxidation rate (MFO) and the exercise intensity that was associated with the Fatmax [For each individual, a best-fit polynomial curve was constructed for their fat and CHO oxidation rate  and forα = 0.05. Continuous data were expressed as the mean ± standard deviation (SD) or as the median (interquartile range) according to the statistical distribution . To account for the between- and within-patient variability due to several measures being taken for the same participant, random-effects models for the correlated data were undertaken rather than the usual statistical tests, as they would have been inappropriate due to an unverified assumption of independence. Time-point evaluations, groups (with/without a TNF blockade and controls) and their interactions were considered as fixed effects, whereas the participant (patient) was considered as a random effect (slope and intercept). A Sidak’s post hoc test for multiple comparisons was applied. The normality of the residuals from these models was studied as described above using the Shapiro–Wilk test. When appropriate, the data were log-transformed to achieve normality of the dependent endpoint. Concerning non-repeated measures, quantitative variables  were compared between groups by ANOVA or by using the Kruskal–Wallis test when assumptions required for the ANOVA were not met . When appropriate , a post hoc test to account for multiple comparisons was performed: A Tukey–Kramer post-ANOVA and a Dunn test after the Kruskal–Wallis test. As less than 5% of the data were missing, the missing data were not dealt with.Statistical analyses were performed using Stata software version 13 . Tests were two-sided with the type-I error set at 2peak, VO2peak (/kg of body mass), disease activity, physical activity levels, and resting metabolism between the three groups.The characteristics of the participants are given in Table p = 0.61). Five patients received no medication, two were treated only with nonsteroidal anti-inflammatory drugs (NSAIDs), two were treated with methotrexate (MTX) plus NSAIDs, six were treated only with MTX, two with MTX plus adalimumab, four with etanercept, six with MTX plus etanercept, two with infliximab, and one was treated with adalimumab plus NSAIDs. The mean duration of the TNF blockade treatment was 25.8 ± 19.1 months.In JIA patients, the disease duration was the same in the treated and untreated TNF blockade groups .The fat and CHO oxidation rates relative to exercise intensities are displayed in Fig. 2peak, fat oxidation rate was higher in the healthy controls than JIA children treated with TNF blockade (p = 0.14)  in the healthy controls, JIA children with TNF blockade and children JIA without TNF blockade, respectively. The heart rate reached at MFO was 135 ± 18, 127 ± 17, and 127 ± 13 beat.min− 1 in the healthy controls, JIA with TNF blockade and JIA without TNF blockade, respectively (p = 0.40). Furthermore, MFO was reached at 53.6 ± 12.3, 49.9 ± 9.9, and 50.6 ± 11.5 of the %VO2peak in healthy controls, JIA with TNF blockade and JIA without TNF blockade respectively (p = 0.67).The heart rate, power, and %VO− 1 and relative to total body mass .Maximal fat oxidation rates expressed in mg.minass Fig.  were higThe first main finding from the present study is that during submaximal exercise, the fat oxidation rate was higher in healthy controls than in JIA patients. Secondly patients under TNF blockade had an intermediate fat oxidation rate profile that was higher than in JIA untreated by TNF blockade but lower than in the healthy controls. These results indicate an apparent dysregulation of fat metabolism during exercise in JIA patients that improves with TNF blockade treatment.2peak per total body mass values were comparable in the three groups, suggesting that the JIA children did not exhibit cardiopulmonary deconditioning in comparison to the controls. Our maximal fat oxidation rates were consistent with other studies in healthy children, although our values in mg.min− 1.kg− 1 were found to be lower since they were normalized to the total body mass and not to total lean mass [The VOean mass . The valean mass , the criean mass .2peak), the rate of plasma FFA uptake is sufficient to account for most of the fat metabolism. However, during strenuous work, there is a greater contribution from muscle triglycerides to total fat oxidation [Metabolic flexibility is the ability of the skeletal muscle to adapt its fuel preference to substrate availability and to its energy requirements . Substraxidation . Furtherxidation . Our resxidation . An impaTNF-α may influence skeletal muscle metabolism via different actions. First, TNF-α is known to stimulate muscle catabolism and alter contractile function in inflammatory diseases . It coulHowever, as there is no difference in the carbohydrate oxidation rate, it does not seem to be a quantitative but rather a qualitative muscle impairment. Skeletal muscle is an insulin-sensitive organ that plays a crucial role in maintaining systemic glucose homeostasis . InflammIn our study, a higher MFO was observed in healthy controls when compared with JIA groups. Moreover, JIA with TNF blockade presented a higher MFO when compared with JIA without TNF blockade, although FATmax (intensity of exercise at which MFO is achieved) was similar between JIA groups and also between healthy controls and JIA groups. A high level of interindividual variation exists in both maximal rates of fat oxidation and the intensity at which maximal rates of fat oxidation occur (FATmax) . But alsFinally, hormones can influence the lipid oxidation rate and the balance of substrates during exercise. For instance, leptin activates AMPK and increases fat oxidation within the skeletal muscle . InteresOur results show that, there is a qualitative alteration of muscle metabolism in JIA. This could have long-term repercussions on health, especially in a cardiovascular point of view. Since lower MFO may be associated with lower insulin sensitivity and, thus, potentially CVD risk on the long term .The relatively small number of patients could be considered as a limitation. A second limitation of this study is the heterogeneity of our patient group. However, we matched patients by pubertal status, gender, and level of physical activity to reduce any bias. In addition, in future studies, the evaluation of their body composition should be taken into account  as well as measurements of their blood lipid, glucose, and insulin levels. In addition, the population under study had different impact diseases . WhetheJuvenile idiopathic arthritis was found to be associated with a metabolic dysregulation of the fat oxidation rate during submaximal exercise, also in an inactive disease  with lower maximal fat oxidation as compared to healthy peers. When children were treated by TNF blockade, fat oxidation rate during submaximal exercise was improved. This metabolic involvement could result from infra-clinical inflammation in JIA patients, which seems to be more controlled by TNF blockade treatment. Impaired metabolic control of lipid is associated with overweight and cardiometabolic risk, but lipid oxidation capacity can be improve by regular physical activity. Whether such alterations can be reversed by training needs to be assessed.


Upregulation of the expression of tumor necrosis factor  has a significant role in the development of autoimmune diseases. The fluorescent antibodies binding TNF may be used for personalized therapy of TNF-dependent diseases as a tool to predict the response to anti-TNF treatment. We generated recombinant fluorescent proteins consisting of the anti-TNF module based on the variable heavy chain (VHH) of camelid antibodies fused with the far-red fluorescent protein Katushka (Kat). Two types of anti-TNF VHH were developed: one (BTN-Kat) that was bound both human or mouse TNF, but did not neutralize their activity, and a second (ITN-Kat) that was binding and neutralizing human TNF. BTN-Kat does not interfere with TNF biological functions and can be used for whole-body imaging. ITN-Kat can be evaluated in humanized mice or in cells isolated from humanized mice. It is able to block human TNF (hTNF) activities both in vitro and in vivo and may be considered as a prototype of a theranostic agent for autoimmune diseases. Therapeutic neutralization of the inflammatory cytokines, in particular TNF, has revolutionized the treatment of autoimmune diseases including rheumatoid arthritis (RA), Crohn’s disease, spondyloarthritis (SpA), and others. However, a significant number of the patients ~50%) with RA and SpA do not respond, showing only marginal improvement, or initially responding but then relapsing ,2,3,4. M0% with RCamelidae family [Since 1989, when a novel type of antibody devoid of the light chains was identified in the sera of various members of the e family , a numbee family . Also, ie family ,17 and ce family . All thee family . Nanobode family .In this study two fluorescent sensors specific to TNF, both fused to far-red protein Katushka, were evaluated. One is based on VHH derived from Bactrian camel  and bindDNA fragments encoding single-domain anti-TNF antibodies VHH41  or ahTNF10 molecular mass against elution volume. The same data were used to calculate the predicted molecular mass of the complex using the plot of molecular mass against elution volumes.ITN-Kat (5 mg/mL) or BTN-Kat (5 mg/mL) was applied to a Superose 6 column  equilibrated in gel-filtration buffer . The flow rate was kept at 0.5 mL/min. Experimental and standard proteins were solubilized in gel-filtration buffer. The Superose 6 column was calibrated with standard proteins . Elution volumes of ITN-Kat were found to be 12 mL, 4 mL, and 11 mL, and those of BTN-Kat were 11 mL, 3 mL, and 10.9 mL. To estimate their molecular masses, we used the plot of logHuman TNF knock-in mice (huTNFKI) described earlier ,27 were Blood was taken from the buccal sinus of mice using sterile medical needles. To isolate the serum, blood was incubated at room temperature for 20–30 min and centrifuged at +4 °C and 14,000 rpm in clot activator tubes . After that, the serum was transferred to fresh test tubes. TNF concentration was measured in sera by the Human-TNF ELISA Ready-SET-Go® .w/v SDS solution in DMSO, and OD was measured at 560 nm. The percentage of living cells was calculated and fitted to a nonlinear regression curve using Prism 5  software. One unit was defined as the amount of TNF inhibitor sufficient to mediate half-maximal protection from cytotoxicity in the presence of 100 U/mL human TNF.The inhibiting activity of BTN-Kat and ITN-Kat in TNF-mediated cytotoxicity was analyzed on WEHI-164 cell line . The celd-Galactosamine   400 ng/g and 800 μg/g, respectively. Mice were euthanized when moribund. Kaplan–Meier survival curves were plotted, and pairwise statistical comparison of BTN-Kat, ITN-Kat, and infliximab was performed.TNF-humanized mice were individually weighted and received intraperitoneal injection of 150, 300, or 450 pmol/g of BTN-Kat; 75, 150, or 300 pmol/g of ITN-Kat; or 150 pmol/g of infliximab or PBS buffer followed 30 min later by an otherwise lethal dose of lipopolysaccharide and v/v) horse serum, and 30% (v/v) of L929 conditioned medium, and then detached with ice-cold PBS and counted. Activation of TNF production by the macrophages was performed using LPS . The cells mortality was obtained by measuring the percentage of dead macrophages by Trypan blue exclusion. Fcγ receptor was blocked by anti-Fcγ receptor antibodies , and then cells were stained with anti-F4/80-FITC antibodies  and BTN-Kat or ITN-Kat by intracellular cytokine staining protocol using BD Fixation and Permeabilization Solution Kit with BD GolgiPlug™ . The samples were analyzed on Cytoflex S flow cytometer . The fluorescent signal from Katushka was detected with 585 nm laser excitation and 660/20 nm emission filter. Twenty thousand cells were evaluated per test; in a list mode, data were analyzed using CytExpert 2.0 software .Bone marrow-derived macrophages (BMDM) from TNF-humanized mice or from C57Bl/6 mice were cultured for 10 days in DMEM, L-glutamune, Pen/Strep, 20% , was incubated with 1× RBC lysis buffer  and then was resuspended in 100 μL of FACS buffer (PBS with 2% FBS), with Fixable Live/Dead Stain  and an appropriate combination of fluorescent antibodies specific to CD45, F4/80, CD3, CD45R , and FcR block . After that, cells were incubated with BTN-Kat and ITN-Kat on ice without permeabilization, or stained by intracellular cytokine staining protocol using BD Fixation and Permeabilization Solution Kit with BD GolgiPlug™ . Data were analyzed using CytExpert 2.0 .d-Gal (800 μg/g) or the same volume of PBS. As a control, mice were injected with PBS 30 min prior to the injection of LPS/d-Gal instead of the proteins or the PBS only. Fluorescence imaging was performed on the IVIS-Spectrum system  in the epi-luminescence mode; the fluorescence was excited at 605/30 nm and detected at 660/20 nm [d-Gal, the mice were euthanized by isoflurane, and whole-body fluorescence images of the animals were acquired. Quantitative analysis was performed in the Living Image Software 4.2  by calculation of the fluorescence intensity averaged over the abdominal region and normalizing to the values of autofluorescence measured from the mice injected with PBS alone. To analyze the uptake of the BTN-Kat or ITN-Kat by the liver, ex vivo fluorescence imaging was performed.150 pmol/g of the fluorescent sensors were injected intraperitonealy (i/p), and after 30 minu animals were i/p injected with LPS (400 ng/g) and 60/20 nm . Before p ≤ 0.05.Statistically significant differences of values were determined using STATISTICA 10  using the Mann–Whitney U-test. Differences were considered statistically significant at Two genetically encoded fluorescent sensors based on two distinct TNF-binding modules and far-red protein Katushka A were de50) = 9.73·103 pM). The clinically utilized chimeric monoclonal antibody infliximab was used as a positive control (EC50 = 2.83·103 pM) . Cells were incubated with a mixture of TNF and inhibitor in a range of concentrations overnight. In contrast to BTN-Kat, ITN-Kat demonstrated a dose-dependent hTNF inhibitory activity  .Additionally, in vitro TNF binding activity of BTN-Kat and ITN-Kat was examined using flow cytometry. Bone marrow-derived macrophages (BMDM) derived from TNF humanized mice were activated by bacterial LPS and then stained with BTN-Kat and ITN-Kat using an intracellular cytokine staining protocol that resulted in specific staining of the TNF-expressing cells. BTN-Kat demonstrated lower staining than ITN-Kat in BMDM cells from huTNFKI mice A; howeveThe ability of BTN-Kat and ITN-Kat to inhibit systemic TNF was evaluated in TNF-humanized mice in the experimental model of acute hepatotoxicity .Mice injected with ITN-Kat at concentration 150 pmol/g survived B, as didd-Gal. Thus, the model of acute hepatotoxicity is characterized by an inflammatory process in the abdominal cavity, and, as expected, the highest level of fluorescence of the sensors was detected in the abdominal area. The peak of the fluorescence intensity was observed at 1 h post-injection of the LPS/d-Gal. In this group fluorescence signal was significantly higher (p < 0.05) than in the control mice injected with PBS and LPS/d-Gal or PBS alone   C. The renjection E.d-gal-induced acute hepatotoxicity. The results showed that the main source of TNF corresponded to F4/80-positive cells. At the same time, the maximal number of TNF-positive cells was observed at 1 h after the LPS/d-gal injection, in correlation with the TNF level in the blood  mice  used herd-Gal. Soluble TNF is the main hepatotoxic mediator in this toxicity model [d-gal-induced acute hepatotoxicity is the myeloid cell compartment  and fluorescent sensor-inhibitor (ITN-Kat), utilizing two single-domain anti-TNF antibodies. We evaluated their ability to bind and neutralize TNF in vitro, and to serve as imaging labels in vitro and in the whole body non-invasive analysis. We concluded that BTN-Kat is a convenient tool for studying the dynamics of TNF expression without interfering with its biological functions, while ITN-Kat is a prototype theranostic agent for TNF-dependent autoimmune diseases.


Toxoplasma gondii (T. gondii) play a critical role during parasite invasion into host cells. In this study, virus-like particles (VLPs) vaccines containing ROP4 and/or ROP13 together with influenza M1 were generated. ROP4+ROP13 VLPs were produced by combining ROP4 VLPs with ROP13 VLPs, and ROP(4 + 13) VLPs by co-infecting insect cells with recombinant baculovirus expressing ROP4 or ROP13. Mice intranasally immunized with ROP(4 + 13) VLPs showed significantly higher levels of IgG, IgG1, IgG2a and IgA antibody responses in sera compared to ROP4+ROP13VLPs. Upon challenge infection by oral route, mice immunized with ROP(4 + 13) VLPs elicited higher levels of IgG and IgA antibody responses in fecal, urine, intestine and vaginal samples as well as CD4+ T, CD8+ T cells, and germinal center B cell responses compared to other type of vaccines, ROP4 VLPs, ROP13 VLPs, and ROP4+ROP13 VLPs. ROP(4 + 13) VLPs vaccination showed a significant decrease in the size and number of cyst in the brain and less body weight loss compared to combination ROP4+ROP13 VLPs upon challenge infection with T. gondii ME49. These results indicated that the ROP(4 + 13) VLPs vaccination provided enhanced protection against T. gondii infection compared to ROP4+ROP13 VLPs, providing an important insight into vaccine design strategy for T. gondii VLPs vaccines.Rhoptry organelle proteins (ROPs) secreted by  T. gondii is a widespread parasite that infects humans and other warm-blooded animals. T. gondii infections are caused by ingesting raw meat and water contaminated with T. gondii oocyst, bradyzoite and tachyzoite. In pregnant women, T. gondii infection can lead to abortions, stillbirths, and neonatal deaths. In addition, T. gondii infection results in toxoplasmic encephalitis and a decrease of the survival rate in immunocompromised individuals such as acquired immunodeficiency syndrome (AIDS) patients [T. gondii [patients ,2. In va. gondii ,4. In th. gondii .T. gondii infection is usually treated with pyrimethamine and sulfadiazine. However, an increasing drug resistance in T. gondii ME49 has rendered the drugs ineffective [T. gondii human vaccine and thus many studies are underway to develop a vaccine. Recent efforts based on DNA or protein vaccines failed due to their inability to confer effective protection against T. gondii ME49 [T. gondii ME49 infection using alternative methods.ffective . Currentdii ME49 ,8,9,10. T. gondii VLPs and Clonorchis sinensis VLPs [The present study utilized the recombinant baculovirus system, which is crucial for producing virus-like particles (VLPs) vaccines . Comparesis VLPs ,14,15,16sis VLPs ,18. In tT. gondii ROP4 and ROP13 are proteins secreted by the rhoptries, which are apical secretory organelles of T. gondii. The ROP4 protein is associated with the vacuole membrane and becomes phosphorylated in the infected host. T. gondii invasion of the host cell involves the secretion of recently identified ROP13 protein, which interacts with the various host cytoplasmic compartments [T. gondii ME49.In this work, ROP4 and ROP13 proteins were specifically used as surface antigens for VLPs. artments ,19. SincT. gondii ME49. We have generated multiple VLPs using T. gondii ROP4 and ROP13. ROP4 and ROP13 VLPs solely expressed either ROP4 or ROP13 antigen, whereas ROP4+ROP13 VLPs were produced by combining ROP4 VLPs and ROP13 VLPs. ROP(4 + 13) VLPs were produced by co-infecting insect cells with recombinant baculoviruse expressing ROP4 or ROP13. We found that ROP(4 + 13) VLPs immunization elicited the highest levels of T. gondii-specific mucosal and systemic immunity, and CD4+, CD8+ T cell and germinal center B cell responses compared to other types of VLPs. This result indicates that ROP(4 + 13) VLPs exhibited the best vaccine efficacy compared to ROP4 VLPs, ROP13 VLPs, and ROP4+ROP13 VLPs. To evaluate the protective efficacy of the VLP vaccines, mice were intranasally immunized with four differently designed vaccines, followed by oral challenge with n = 10) were purchased from KOATECH . T. gondii RH and ME49 were maintained in mice by intraperitoneal injection. All animal experiments and husbandry involved in these studies presented in this manuscript were conducted under the guidelines of the Kyung Hee University IACUC (permit number: KHUASP (SE)-18-050, 7 June 2018). T. gondii RH were collected 5 days after infection from peritoneal fluids and T. gondii ME49 cyst was collected from the brains of mice 30 days after infection [Spodoptera frugiperda (Sf9) insect cells were used to generate recombinant baculoviruses (rBVs) and T. gondii VLPs were cultured in SF900-II serum-free medium  at 27 °C in spinner flasks at 135 rpm.Seven weeks old BALB/c female mice  after obtaining T. gondii RNA. The primers used for T. gondii ROP4 and ROP13 were as follows: 5-AAA GCATGC ACC ATG GGG CAC CCT ACC TCT TT-3 and 5-TTA GGTACC TCA CGT TTC CGG TGG TGG CAT-3, 5-AAA GGATCC ACC ATG AAG AGA ACA GAG CTT TG-3 and 5-TTA CTCGAG TCA CAA TAG CCT CAA GGA ATT-3, respectively. Amplified genes were cloned into the pFastBac with Sph I/Kpn I (ROP4) and BamH I/Xho I (ROP13) sites. Afterwards, pFastBac transformants containing ROP4 or ROP13 were transfected into SF9 cells using cellfectin II . For T. gondii VLP vaccine generation, Sf9 cells were co-infected with ROP4, ROP13, and M1 rBVs. Sf9 cell culture supernatants were harvested on day 3 post-infection and subsequently centrifuged at 6000 rpm for 30 min. Supernatants were collected and centrifuged at 30,000 rpm for 30 min. VLPs were purified through a sucrose gradient as described [escribed ,21. VLPsT. gondii ROP4, ROP13 or M1 proteins were characterized by western blots and transmission electron microscopy (TEM) . Influenza virus anti-M1 monoclonal antibody  or T. gondii-specific polyclonal antibody were used for detection of ROP4, ROP13 or M1 proteins by western blot. T. gondii-specific polyclonal antibody was obtained from mice infected with T. gondii ME49. Briefly, peripheral blood collected from mice via retro-orbital plexus puncture were centrifuged at 7000 RPM for 10 min and the resulting polyclonal antibodies were stored at −20 °C until use. Morphology of VLPs was confirmed through TEM analysis [analysis . T. gondii ME49 cysts and monitored daily to record body weight changes for 60 days. Mice were sacrificed four weeks after challenge infection with T. gondii ME49 to collect intestine, urine, feces, vaginal secretion, brain, spleen, and mesenteric lymph nodes (MLN). Groups of 10 mice were intranasally (IN) immunized twice with ROP4 VLPs (100 μg), ROP13 VLPs (100 μg), mixed ROP4 VLPs (50 μg) and ROP13 VLPs (50 μg), and ROP(4 + 13) VLPs (100 μg) at 4-week intervals. Four weeks after the last immunization, mice were orally challenged with 450 T. gondii-specific antibody levels  were determined by enzyme-linked immunosorbent assay (ELISA) using T. gondii RH coating antigen (4 μg/mL). T. gondii RH coating antigens were prepared by sonicating of tachyzoites of T. gondii RH in PBS as described [escribed . Immune T. gondii-specific immunoglobulin G (IgG) and IgA antibody responses by enzyme-linked immunosorbent assay (ELISA) as previously described [After 30 days of challenge infection, spleens were collected from mice and homogenized. Splenocytes were passed through the cell strainer and the red blood cells were removed as described . MLN wasescribed ,23. 6 cells/well) were incubated at 37 °C for 4 days in 96 well cell culture plates coated with T. gondii RH antigen (4 μg/mL). After 4 days, splenocytes were removed from the wells of plates and the plates were reacted with HRP-conjugated mouse IgG and IgA antibodies as described [T. gondii RH (4 μg/mL). Cytokine concentrations of IL-6 and IFN-γ were quantified using ELISA kits following the manufacturer’s instructions .Isolated mouse splenocyte  antibodies in staining buffer  after Fc receptor blocking as described previously [Splenocytes and mesenteric lymph nodes (MLN) were collected at week 4 after challenge infection with eviously . Cell poT. gondii ME49 cysts were isolated from the brains of mice 4 weeks after challenge infection. Brain tissues were homogenized with a syringe in 400 μL PBS. After mixing homogenates with 45% Percoll solution, samples were centrifuged at 12,100 rpm for 20 min using a swing-bucket rotor. Brain homogenates were subdivided into 3 layers following centrifugation: T. gondii ME49 cyst, RBC, and brain tissues. The T. gondii ME49 cysts layer was carefully collected and centrifuged at 6000 rpm for 20 min with repeated PBS washing. Five microliters of the collected cysts were placed on a slide glass and cysts were counted from 5 different areas under the microscope .post hoc test or 2-way ANOVA with Bonferroni’s test. The data was considered statistically significant if p value * < 0.05, ** < 0.01, *** < 0.001.Statistical analysis was performed using GraphPad prism version 5 . Data was analyzed using one-way ANOVA with Tukey’s T. gondii polyclonal antibodies and anti-M1 monoclonal antibodies. Also, the successful formation of spherical VLP vaccines with ROP4 . It was noted that mice immunized with ROP(4 + 13) VLPs showed the highest levels of T. gondii-specific IgG (p < 0.05).Mice were intranasally immunized with ROP4 VLPs, ROP13 VLPs, ROP4+ROP13 VLPs, and ROP(4 + 13) VLPs at weeks 0 and 4. As seen in ific IgG A, IgG1  VLPs. Also, IgA . These results indicate that mucosal immunity was successfully induced by IN immunization with ROP4 VLPs, ROP13 VLPs, ROP4+ROP13 VLPs, and ROP(4 + 13) VLPs.After challenge infection with lso, IgA B,D,F,H aT. gondii-specific IgG and IgA antibodies secreted into splenocyte culture supernatants were determined by ELISA. ROP(4 + 13) VLPs immunization exhibited higher levels of T. gondii IgG (p < 0.05). Also, to confirm VLPs vaccine efficacy, the cellular immune response was determined by measuring IFN-γ and IL-6 cytokines secreted from splenocytes cultures supernatants with T. gondii RH stimulation. The observation of the increase in the production of IFN-γ . Also, ROP(4 + 13) VLPs immunization induced higher levels of activation of CD8+ T cell (p < 0.05). It is interesting to note that MLN exhibited greater quantities of germinal center B cell responses compared to spleen . In addition, as seen in T. gondii ME49, naïve challenge mice showed a rapid body weight loss, whereas mice immunized with ROP(4 + 13) VLPs showed 2% of body weight loss. As a result, as shown in T. gondii ME49 invasion. The best way to demonstrate the efficacy of the T. gondii ROP 4 and/or ROP13. We found that ROP(4 + 13) VLPs generated by co-infecting insect cells with recombinant baculoviruses presenting ROP4 and ROP13 conferred better protection than ROP4+ROP13 VLPs generated by combining ROP4 and ROP13 VLPs. Currently, a number of T. gondii antigens  have been evaluated as potential vaccine candidates against toxoplasmosis [T. gondii infection in mice [T. gondii ME49 [T. gondii significantly reduced parasite burden against highly virulent T. gondii tachyzoites (GT1) infection [T. gondii contribute to protection against T. gondii infection. VLPs targeting multi-antigens showed better parasite - specific IgG, IgA antibody responses, germinal center B cell responses, and CD4+, CD8+ T cell responses compared to VLPs vaccines targeting single antigen [T. gondii infection. Interestingly, multi-antigen VLPs also significantly reduce proinflammatory cytokine levels and provide lower apoptosis responses, compared to single antigen VLP vaccines. Thus, in this study, we hypothesized that both ROP4 and ROP13 proteins containing VLPs platform can induce high levels of vaccine efficacy. As expected, ROP(4 + 13) VLPs vaccination showed the lowest body weight loss compared to ROP4 VLPs, ROP13 VLPs and combination of ROP4+ROP13 VLPs, further demonstrating better protection than a combination of ROP4 and ROP13 VLPs. This is consistent with our previous study, in which multi-antigen VLPs showed better protection than combination VLPs [In this study, we evaluated the vaccine efficacy induced by influenza virus - like nanoparticles containing lasmosis . Among tlasmosis . Rhoptry in mice . ROP4 VLdii ME49 . Recentlnfection . Enhance antigen ,27. Thesion VLPs . T. gondii ME49 as a challenge infection pathogen, using an oral route since human infection is naturally acquired through the oral route through the ingestion of meat, water or other food products contaminated with cysts or oocyst of T. gondii [T. gondii parasite-specific IgG, IgG1, IgG2a, and IgA antibody responses from sera, as well as mucosal IgG and IgA responses of vaginal secretions, urine, feces, and intestines were higher in the ROP(4 + 13) VLPs-vaccinated group compared to other groups. Higher levels of IgG and IgA antibody responses detected from mucosal samples, particularly the intestines and/or feces from ROP(4 + 13) VLPs-vaccinated mice might have contributed to better protection compared to other vaccination groups. Interestingly, ROP(4 + 13) VLPs vaccination showed better immunity than ROP4+ROP13 VLPs, ROP4 VLPs and ROP13 VLPs, in which ROP4 VLPs vaccination showed similar levels of immune responses to ROP13 in inducing mucosal antibody responses, ASC responses and germinal center B cell responses. Our results indicated that cellular responses are also important, contributing protection against T. gondii infection. CD4+ and CD8+ T cell responses in ROP(4 + 13) VLPs vaccination showed higher than other VLPs, in which ROP(4 + 13) VLPs vaccination induced the best vaccine efficacy. These immunological parameters were exactly correlated with the protection induced by VLPs immunization, in which cyst size, cyst counts, and body weight loss was lowest and the survival rate of mice was highest in ROP(4 + 13) VLPs vaccination, followed by ROP4+ROP13 VLPs and ROP4 VLPs or ROP13 VLPs in order. It is well known that mucosal vaccination induces antigen-specific humoral and cell-mediated immune responses in both the systemic and mucosal compartments . In our . gondii . We founT. gondii VLPs induces better protection than IM immunization by eliciting systemic and mucosal immunity [IN route immunization with immunity . Parasitimmunity  and alsoimmunity . Germinaimmunity . CompareT. gondii ME49 challenge infection. Among them, ROP(4 + 13) VLPs immunization showed the best protective immunity against a T. gondii ME49 challenge infection, in which mucosal immunity might play an important role. In conclusion, we generated ROP4 VLPs, ROP13 VLPs, ROP4 + ROP13 VLPs, and ROP(4 + 13) VLPs and found that they all have protective efficacy against a


Anti-TNF (tumor necrosis factor) medications work by inhibiting the production of TNF or its effect on target organs. TNF is a cell-signaling protein, or cytokine, involved in systemic inflammation and is one of the cytokines that make up the acute phase reactants. TNF inhibitors are available for the treatment of a number of rheumatic and other immune-mediated diseases. Treatment of rheumatoid arthritis with anti-TNFα  agents may lead to autoantibody formation and flares of vasculitis. Although medications are a common cause of renal injury, anti-TNFα medications very rarely cause renal complications. We present a case of a patient who presented with nausea and flu-like illness and was ultimately found to have etanercept-induced nephropathy. Anti-TNF (tumor necrosis factor) medications work by inhibiting the production of TNF or its effect on target organs. TNF is a cell-signaling protein, or cytokine, involved in systemic inflammation and is one of the cytokines that make up the acute phase reactants. TNF is produced chiefly by activated macrophages, although it can be produced by other cells such as CD4+ (cluster of differentiation 4) lymphocytes, natural killer (NK) cells, neutrophils, mast cells, eosinophils, and neurons . TNF levEtanercept, otherwise known as enbrel, is a TNF inhibitor that was approved by the Food and Drug Administration (FDA) in 1998 for the treatment of rheumatoid arthritis, polyarticular juvenile idiopathic arthritis in patients aged two or older, psoriatic arthritis, ankylosing spondylitis, and plaque psoriasis in patients aged four or older . The mosA 56-year-old female with a past medical history significant for hypothyroidism, hyperlipidemia, and seropositive rheumatoid arthritis diagnosed in 2014 presented to the nephrology clinic upon referral for evaluation of acute kidney injury, proteinuria, and hematuria. The patient first presented to her primary care physician in July 2017 due to a flu-like illness accompanied by nausea, stomach upset, and dark urine. She was prescribed a two-week course of ciprofloxacin by her primary care physician for a presumed urinary tract infection and then prescribed a two-week course of trimethoprim-sulfamethoxazole due to persistent symptoms.In August 2017, the patient presented to the emergency department with persistent nausea, vomiting, fatigue, and anorexia. Laboratory workup at this visit revealed an elevated creatinine of 1.79 mg/dL from a baseline creatinine of 0.75 mg/dL in February 2017. The patient received intravenous (IV) fluids and ondansetron. Further workup was non-contributory, and the patient was subsequently discharged to home with a follow-up appointment with her primary care physician. The following month, the patient presented to the emergency department with similar symptoms and was admitted to the hospital. Her creatinine upon presentation was 2.00 mg/dL, which improved to 1.56 mg/dL with IV fluids. She was discharged to home the following day with a plan to follow up with outpatient nephrology.The patient presented to the nephrology clinic in September 2017 with persistent generalized fatigue, malaise, nausea, and occasional vomiting. She reported that her lack of energy affected her ability to perform her job. The patient’s medications included levothyroxine 75 mcg daily, methotrexate 7.5 mg per week, etanercept 50 mg subcutaneous every week since January 2016, simvastatin 20 mg daily, famotidine, folic acid, vitamin D and biotin. Vital signs revealed a temperature of 37°C, a pulse of 72 beats per minute, a blood pressure of 130/92, and a respiratory rate of 18. Physical examination did not reveal any abnormalities. Initial laboratory workup was significant for a blood urea nitrogen (BUN) level of 29 mg/dL , creatinine level of 1.67 mg/dL  and a urine protein/creatinine ratio of 3.30   alpha and beta, pamidronate, and lithium have been shown to cause both minimal change disease and focal segmental glomerulosclerosis. Additionally, non-steroidal anti-inflammatories and cyclooxygenase-2 inhibitors have been shown to cause minimal change disease, and sirolimus and anabolic steroids have been shown to cause focal segmental glomerulosclerosis. Certain anti-angiogenesis drugs, such as mitomycin-C, gemcitabine, interferon, antiplatelet agents, and quinine have been shown to cause thrombotic microangiopathy .There are currently five TNF inhibitors available for the treatment of rheumatoid arthritis, psoriatic arthritis and psoriasis, spondylarthritis, inflammatory bowel disease, and a number of other rheumatic and other immune-mediated diseases. Biosimilar forms of several TNF inhibitors have become commercially available in some countries, and additional biosimilar agents are being developed. These medications include etanercept, infliximab, adalimumab, certolizumab, and golimumab. Etanercept is a soluble p75 TNF receptor fusion protein that consists of two p75 TNF receptors bound to the Fc portion of immunoglobulin G (IgG). TNF is considered a key inflammatory cytokine in the pathogenesis of rheumatoid arthritis and its dysregulated secretion both induces and maintains synovitis in patients with this disease. As such, TNF inhibitors such as etanercept are widely used in the treatment of this disease .Studies suggest that TNF inhibitors work by causing cell cycle arrest, apoptosis, and suppression of cytokine production in both monocytes and macrophages in synovial fluid and blood . In addiBaseline testing for antinuclear antibody (ANA), anti-dsDNA (double-stranded DNA), and antineutrophil cytoplasmic antibody (ANCA) should be performed in all patients with rheumatoid arthritis prior to commencing etanercept inhibitor therapy in order to help identify new, drug-induced autoimmune processes in those patients who eventually develop glomerular disease. TNF inhibitors such as etanercept should be used with caution and patients on this therapy should have their renal function routinely monitored. Although it is rare, TNF inhibitor-induced nephrotoxicity should be considered in patients who are using TNF inhibitors and present with acute kidney injury and abnormal urinalysis. If patients do develop renal disease, a final decision about drug continuation or withdrawal should be entertained after discussing the risks of renal failure and the benefits of treatment of autoimmune disease with the patient.We present a rare case of etanercept-related crescentic glomerulonephritis in a patient being treated for rheumatoid arthritis. While the exact mechanism of this adverse reaction is still under investigation, TNF inhibitor-related nephrotoxicity is supported by the temporal relationship between onset of glomerular disease in patients without previous renal disease and improvement after the withdrawal of TNF inhibitor therapy.