Search

BRAFV600E mutation is the most frequent genetic alteration in papillary thyroid cancer (PTC). β-Catenin (Ctnnb1) is a key downstream component of canonical Wnt signaling pathway and is frequently overexpressed in PTC. BRAFV600E-driven tumors have been speculated to rely on Wnt/β-catenin signaling to sustain its growth, although many details remain to be elucidated. In this study, we investigated the role of β-catenin in BrafV600E-driven thyroid cancer in a transgenic mouse model. In BrafV600E mice with wild-type (WT) Ctnnb1 (BVE-Ctnnb1WT or BVE), overexpression of β-catenin was observed in thyroid tumors. In BrafV600E mice with Ctnnb1 knockout (BVE-Ctnnb1null), thyroid tumor growth was slowed with significant reduction in papillary architecture. This was associated with increased expression of genes involved in thyroid hormone synthesis, elevated 124iodine uptake, and serum T4. The survival of BVE-Ctnnb1null mice was increased by more than 50% during 14-month observation. Mechanistically, downregulation of MAPK, PI3K/Akt, and TGFβ pathways and loss of epithelial–mesenchymal transition (EMT) were demonstrated in the BVE-Ctnnb1null tumors. Treatment with dual β-catenin/KDM4A inhibitor PKF118–310 dramatically improved the sensitivity of BVE-Ctnnb1WT tumor cells to BRAFV600E inhibitor PLX4720, resulting in significant growth arrest and apoptosis in vitro, and tumor regression and differentiation in vivo. These findings indicate that β-catenin signaling plays an important role in thyroid cancer growth and resistance to BRAFV600E inhibitors. Simultaneously targeting both Wnt/β-catenin and MAPK signaling pathways may achieve better therapeutic outcome in BRAFV600E inhibitor-resistant and/or radioiodine-refractory thyroid cancer.

Protein homeostasis (proteostasis) is crucial for proper cellular function, including the production of peptides with biological functions through controlled proteolysis. Proteostasis has roles in maintenance of cellular functions and plant interactions with the environment under physiological conditions. Plant stress continues to reduce agricultural yields causing substantial economic losses; thus, it is critical to understand how plants perceive stress signals to elicit responses for survival. As previously shown in Arabidopsis thaliana, thimet oligopeptidases (TOPs) TOP1 (also referred to as organellar oligopeptidase) and TOP2 (also referred to as cytosolic oligopeptidase) are essential components in plant response to pathogens, but further characterization of TOPs and their peptide substrates is required to understand their contributions to stress perception and defense signaling. Herein, label-free peptidomics via liquid chromatography-tandem mass spectrometry was used to differentially quantify 1111 peptides, originating from 369 proteins, between the Arabidopsis Col-0 wild type and top1top2 knock-out mutant. This revealed 350 peptides as significantly more abundant in the mutant, representing accumulation of these potential TOP substrates. Ten direct substrates were validated using in vitro enzyme assays with recombinant TOPs and synthetic candidate peptides. These TOP substrates are derived from proteins involved in photosynthesis, glycolysis, protein folding, biogenesis, and antioxidant defense, implicating TOP involvement in processes aside from defense signaling. Sequence motif analysis revealed TOP cleavage preference for non-polar residues in the positions surrounding the cleavage site. Identification of these substrates provides a framework for TOP signaling networks, through which the interplay between proteolytic pathways and defense signaling can be further characterized.

Sperm-specific phospholipase C zeta (PLCζ) initiates intracellular calcium (Ca2+) transients which drive a series of concurrent events collectively termed oocyte activation. Numerous investigations have linked abrogation and absence/reduction of PLCζ with forms of male infertility in humans where oocyte activation fails. However, very few studies have examined potential relationships between PLCζ and advancing male age, both of which are increasingly considered to be major effectors of male fertility. Initial efforts in humans may be hindered by inherent PLCζ variability within the human population, alongside a lack of sufficient controllable repeats. Herein, utilizing immunoblotting, immunofluorescence, and quantitative reverse transcription PCR (qRT-PCR) we examined for the first time PLCζ protein levels and localization patterns in sperm, and PLCζ mRNA levels within testes, from mice at 8 weeks, 12 weeks, 24 weeks, and 36 weeks of age, from two separate strains of mice, C57BL/6 (B6; inbred) and CD1 (outbred). Collectively, advancing male age generally diminished levels and variability of PLCζ protein and mRNA in sperm and testes, respectively, when both strains were examined. Furthermore, advancing male age altered the predominant pattern of PLCζ localization in mouse sperm, with younger mice exhibiting predominantly post-acrosomal, and older mice exhibiting both post-acrosomal and acrosomal populations of PLCζ. However, the specific pattern of such decline in levels of protein and mRNA was strain-specific. Collectively, our results demonstrate a negative relationship between advancing male age and PLCζ levels and localization patterns, indicating that aging male mice from different strains may serve as useful models to investigate PLCζ in cases of male infertility and subfertility in humans.

To our knowledge, this is the first inclusive assessment of the status of Sparidae fisheries in the territorial waters of Kuwait, integrating information on catch trends, consumer preferences, recreational fishing behaviour, and molecular based identification of fish species by DNA barcoding Fisheries landing data were obtained from the official fisheries bulletin released by the Kuwait Central Statistical Bureau. Surveys were conducted to assess consumer preferences on seabream species, landed seabream species, and the behaviour of recreational anglers towards seabream species. DNA barcodes were generated to authenticate commercial seabream using the mitochondrial cytochrome c oxidase subunit I (COXI) gene and then compared with GenBank sequence entries; these sequences were then used to construct a neighbour‐joining (NJ) phylogenetic tree. Yellowfin seabream Acanthopagrus latus was the prime and most favoured commercial seabream species (45%), followed by Sparidentex hasta, which was the top‐rated secondary, unassessed seabream species. Approximately 54% of households conduct recreational fishing more than twice a month, and most (77%) primarily target S. hasta and Ac. latus. Consumer demand for secondary seabreams, which were not assessed and are listed in the bulletin under the category ‘others’, was evident. The NJ phylogenetic tree revealed that Arabian Gulf seabreams, including those of Kuwait, are genetically different from their counterparts inhabiting neighbouring waters. The data presented here highlight the urgency to modify the existing fisheries list using accurate identification tools, such as DNA barcoding, for the sustainable and conservation‐oriented management of local fisheries, which are in decline.

Triple-negative breast cancer (TNBC) is a very aggressive subtype with high recurrence rate and no molecular targets for therapies. This subtype is characterized by high expression/secretion of the proinvasive/metastatic interleukin-6 (IL-6) cytokine. In the present study, we have shown that tocilizumab inhibits the IL-6/STAT3 signaling and suppresses the cancer/inflammatory epigenetic IL-6/STAT3/NF-κB positive feedback loop. Furthermore, tocilizumab inhibited the proliferative and the migratory/invasiveness capacities as well as the epithelial-to-mesenchymal transition (EMT) process in TNBC cells. Importantly, tocilizumab suppressed the stemness-related characteristics of TNBC cells, through the inhibition of the Wnt/β-catenin breast cancer stem cell-related pathway. Additionally, we have shown that tocilizumab suppresses the paracrine activation of normal breast stromal fibroblasts to myofibroblats. Moreover, tocilizumab sensitized TNBC cells to the cytotoxic effect of cisplatin in vitro. Furthermore, pharmacological inhibition of IL-6 by tocilizumab had great inhibitory effect on tumor growth and the EMT process in humanized orthotopic breast tumors in mice. In addition, tocilizumab potentiated the proapoptotic effect of cisplatin in humanized breast tumors. Together, these findings indicate that tocilizumab can suppress the prometastatic capacity of TNBC cells and enhances the cytotoxic effect of cisplatin against these cells. Therefore, tocilizumab could be of great therapeutic value for these hard-to-treat TNBC patients.

Aniridia is a rare eye disorder, which is caused by mutations in the paired box 6 (PAX6) gene and results in vision loss due to the lack of a long-term vision-saving therapy. One potential approach to treating aniridia is targeted CRISPR-based genome editing. To enable the Pax6 small eye (Sey) mouse model of aniridia, which carries the same mutation found in patients, for preclinical testing of CRISPR-based therapeutic approaches, we endogenously tagged the Sey allele, allowing for the differential detection of protein from each allele. We optimized a correction strategy in vitro then tested it in vivo in the germline of our new mouse to validate the causality of the Sey mutation. The genomic manipulations were analyzed by PCR, as well as by Sanger and next-generation sequencing. The mice were studied by slit lamp imaging, immunohistochemistry, and western blot analyses. We successfully achieved both in vitro and in vivo germline correction of the Sey mutation, with the former resulting in an average 34.8% ± 4.6% SD correction, and the latter in restoration of 3xFLAG-tagged PAX6 expression and normal eyes. Hence, in this study we have created a novel mouse model for aniridia, demonstrated that germline correction of the Sey mutation alone rescues the mutant phenotype, and developed an allele-distinguishing CRISPR-based strategy for aniridia., Graphical Abstract, Aniridia is a rare genetic disease that reduces vision and progresses to blindness due to unmet therapeutic needs. The authors first created a new aniridic mouse model that allows quantification of genome-based therapies. They then successfully developed a CRISPR strategy to correct this mouse’s germline mutation and completely rescued the blindness.

Membrane trafficking is a complex, essential process in eukaryotic cells responsible for protein transport and processing. Deficiencies in vacuolar protein sorting (VPS) proteins, key regulators of trafficking, cause abnormal intracellular segregation of macromolecules and organelles and are linked to human disease. VPS proteins function as part of complexes such as the homotypic fusion and vacuole protein sorting (HOPS) tethering complex, composed of VPS11, VPS16, VPS18, VPS33A, VPS39 and VPS41. The HOPS-specific subunit VPS41 has been reported to promote viability of dopaminergic neurons in Parkinson’s disease but to date has not been linked to human disease. Here, we describe five unrelated families with nine affected individuals, all carrying homozygous variants in VPS41 that we show impact protein function. All affected individuals presented with a progressive neurodevelopmental disorder consisting of cognitive impairment, cerebellar atrophy/hypoplasia, motor dysfunction with ataxia and dystonia, and nystagmus. Zebrafish disease modelling supports the involvement of VPS41 dysfunction in the disorder, indicating lysosomal dysregulation throughout the brain and providing support for cerebellar and microglial abnormalities when vps41 was mutated. This provides the first example of human disease linked to the HOPS-specific subunit VPS41 and suggests the importance of HOPS complex activity for cerebellar function., Sanderson et al. describe ten individuals with biallelic variants in the gene encoding the vacuolar protein sorting protein VPS41, presenting with cerebellar dysfunction and abnormal membrane-bound vesicles. Mutant zebrafish recapitulate the cerebellar dysfunction phenotype, further implicating VPS41 as a human disease gene.

Context Hypophosphatemic rickets (HR) is a group of rare hereditary renal phosphate wasting disorders caused by mutations in PHEX, FGF23, DMP1, ENPP1, CLCN5, SLC9A3R1, SLC34A1, or SLC34A3. Objective A large kindred with 5 HR patients was recruited with dominant inheritance. The study was undertaken to investigate underlying genetic defects in HR patients. Design Patients and their family members were initially analyzed for PHEX and FGF23 mutations using polymerase chain reaction sequencing and copy number analysis. Exome sequencing was subsequently performed to identify novel candidate genes. Results PHEX and FGF23 mutations were not detected in the patients. No copy number variation was observed in the genome using CytoScan HD array analysis. Mutations in DMP1, ENPP1, CLCN5, SLC9A3R1, SLC34A1, or SLC34A3 were also not found by exome sequencing. A novel c.979–96 T>A mutation in the SGK3 gene was found to be strictly segregated in a heterozygous pattern in patients and was not present in normal family members. The mutation is located 1 bp downstream of a highly conserved adenosine branch point, resulted in exon 13 skipping and in-frame deletion of 29 amino acids, which is part of the protein kinase domain and contains a Thr-320 phosphorylation site that is required for its activation. Protein tertiary structure modelling showed significant structural change in the protein kinase domain following the deletion. Conclusions The c.979–96 T>A splice mutation in the SGK3 gene causes exon 13 skipping and deletion of 29 amino acids in the protein kinase domain. The SGK3 mutation may cause autosomal dominant HR.

The efficacy of Agroecosystem Compounds, fipronil, spinosad and chlorantraniliprole as alternatives to the conventional insecticides on the mortality rates and haemolymph protein, lipid and carbohydrate contents were assessed in Schistocerca gregaria and Locusta migratoria migratorioides. One day old 5th nymphal instars of each of the mentioned species were treated by various concentrations of tested compounds by the feeding technique. The results indicated that Locusta migratoria migratorioides is more sensitive to all tested insecticides than Schistocerca gregaria. In both species, fipronil was the most effective followed by spinosad, and chlorantraniliprole was the least effective. LC30 concentrations of the three compounds were used to treat the 5th instar hoppers of both species to examine the effects of sub lethal concentrations of the mentioned insecticides on the haemolymph total proteins, total lipids and total carbohydrates. It was found that haemolymph proteins, lipids and carbohydrates contents were dramatically reduced for the treated hoppers when compared to the untreated ones of both species. Malformed hopper and wrinkled winged adult were seen in insects treated by sub lethal concentrations of fipronil and spinosad.

Ciliopathies are clinical disorders of the primary cilium with widely recognized phenotypic and genetic heterogeneity. In two Arab consanguineous families, we mapped a ciliopathy phenotype that most closely matches Joubert syndrome (hypotonia, developmental delay, typical facies, oculomotor apraxia, polydactyly, and subtle posterior fossa abnormalities) to a single locus in which a founder homozygous truncating variant in FAM149B1 was identified by exome sequencing. We subsequently identified a third Arab consanguineous multiplex family in which the phenotype of Joubert syndrome/oral-facial-digital syndrome (OFD VI) was found to co-segregate with the same founder variant in FAM149B1. Independently, autozygosity mapping and exome sequencing in a consanguineous Turkish family with Joubert syndrome highlighted a different homozygous truncating variant in the same gene. FAM149B1 encodes a protein of unknown function. Mutant fibroblasts were found to have normal ciliogenesis potential. However, distinct cilia-related abnormalities were observed in these cells: abnormal accumulation IFT complex at the distal tips of the cilia, which assumed bulbous appearance, increased length of the primary cilium, and dysregulated SHH signaling. We conclude that FAM149B1 is required for normal ciliary biology and that its deficiency results in a range of ciliopathy phenotypes in humans along the spectrum of Joubert syndrome.

Normal breast fibroblasts (NBFs) support and maintain the architecture of the organ, and can also suppress tumorigenesis. However, the mechanisms involved are not fully understood. We have shown here that NBFs suppress breast carcinogenesis through secretion of osteoprotegerin (OPG), a soluble decoy receptor for the Receptor Activator of NF-κB ligand (RANKL). Indeed, NBFs and human recombinant OPG (rOPG), suppressed breast cancer cells proliferation and motility through inhibition of the epithelial-to-mesenchymal transition (EMT) process both in vitro and in vivo. Additionally, rOPG inhibited the IL-6/STAT3 and NF-κB pathways as well as the OPG gene, which turned out to be STAT3-regulated. This was confirmed using denosumab, a RANKL-targeted antibody, which also inhibited NF-κB, down-regulated OPG and repressed EMT in breast cancer cells grown in 2D and 3D. Importantly, both rOPG and denosumab targeted cancer stem cells (CSCs). This was mediated through inhibition of the CSC-related pathway β-catenin. Moreover, rOPG reduced tumor growth and inhibited breast CSC biomarkers in orthotopic humanized breast tumors. Therefore, normal mammary fibroblasts can suppress carcinogenesis through OPG, which constitutes great potential as preventive and/or therapeutic molecule for breast carcinomas.

Made available in DSpace on 2021-06-25T15:08:19Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-03-09 Wellcome Trust From 1985 to 2016, the prevalence of underweight decreased, and that of obesity and severe obesity increased, in most regions, with significant variation in the magnitude of these changes across regions. We investigated how much change in mean body mass index (BMI) explains changes in the prevalence of underweight, obesity, and severe obesity in different regions using data from 2896 population-based studies with 187 million participants. Changes in the prevalence of underweight and total obesity, and to a lesser extent severe obesity, are largely driven by shifts in the distribution of BMI, with smaller contributions from changes in the shape of the distribution. In East and Southeast Asia and sub-Saharan Africa, the underweight tail of the BMI distribution was left behind as the distribution shifted. There is a need for policies that address all forms of malnutrition by making healthy foods accessible and affordable, while restricting unhealthy foods through fiscal and regulatory restrictions. Imperial Coll London, London, England Harvard TH Chan Sch Publ Hlth, Boston, MA USA Middlesex Univ, London, England World Hlth Org, Geneva, Switzerland Minist Hlth, Victoria, Seychelles Univ Lausanne, Lausanne, Switzerland Eduardo Mondlane Univ, Maputo, Mozambique Victor Babes Univ Med & Pharm Timisoara, Timisoara, Romania Univ Sydney, Sydney, NSW, Australia Natl Inst Biomed Innovat Hlth & Nutr, Tokyo, Japan Univ Auckland, Auckland, New Zealand Seoul Natl Univ, Seoul, South Korea ICMR Natl Inst Nutr, Hyderabad, India Capital Med Univ, Beijing An Zhen Hosp, Beijing, Peoples R China Univ Peruana Cayetano Heredia, Lima, Peru Univ Tunis El Manar, Tunis, Tunisia Univ Oulu, Oulu, Finland Univ Zagreb, Zagreb, Croatia Univ Ljubljana, Ljubljana, Slovenia Caja Costarricense Seguro Social, San Jose, Costa Rica Al Quds Univ, East Jerusalem, State Of Palest, Palestine Natl Ctr Publ Healthcare, Nur Sultan, Kazakhstan Minist Hlth, Kuala Lumpur, Malaysia Qatar Univ, Doha, Qatar Birzeit Univ, Birzeit, State Of Palest, Israel Usmanu Danfodiyo Univ Teaching Hosp, Sokoto, Nigeria Inst Mexicano Seguro Social, Mexico City, DF, Mexico Flinders Univ S Australia, Adelaide, SA, Australia Mahidol Univ, Salaya, Nakhon Pathom, Thailand BRAC James P Grant Sch Publ Hlth, Dhaka, Bangladesh Univ Copenhagen, Copenhagen, Denmark Copenhagen Univ Hosp, Copenhagen, Denmark Food & Nutr Res Inst, Taguig, Philippines Urmia Univ Med Sci, Orumiyeh, Iran Inst Nacl Ciencias Med & Nut, Mexico City, DF, Mexico Univ Amsterdam, Amsterdam, Netherlands Shahrekord Univ Med Sci, Shahrekord, Iran Noncommunicable Dis Res Ctr, Tehran, Iran Univ Oslo, Oslo, Norway Univ Bremen, Bremen, Germany Republican Ctr Hlth Promot, Bishkek, Kyrgyzstan Natl Ctr Diabet Endocrinol & Genet, Amman, Jordan Princess Nourah Bint Abdulrahman Univ, Riyadh, Saudi Arabia Kuwait Inst Sci Res, Kuwait, Kuwait King Abdulaziz Univ, Jeddah, Saudi Arabia Dasman Diabet Inst, Kuwait, Kuwait Luxembourg Inst Hlth, Strassen, Luxembourg Aldara Hosp & Med Ctr, Riyadh, Saudi Arabia King Abdullah Int Med Res Ctr, Riyadh, Saudi Arabia Bispebjerg & Frederiksberg Hosp, Copenhagen, Denmark Barcelona Inst Global Hlth CIBERESP, Barcelona, Spain World Hlth Org Reg Off Eastern Mediterranean, Cairo, Egypt Bombay Hosp & Med Res Ctr, Mumbai, Maharashtra, India Res Ctr Social Determinants Hlth, Tehran, Iran UMR CNRS MNHN 7206 Ecoanthropologie, Paris, France Univ Lille, Lille, France Lille Univ Hosp, Lille, France Western Norway Univ Appl Sci, Sogndal, Norway Norwegian Sch Sport Sci, Oslo, Norway Madras Diabet Res Fdn, Chennai, Tamil Nadu, India Zahedan Univ Med Sci, Zahedan, Iran Natl Inst Publ Hlth, Tunis, Tunisia Univ Porto, Inst Publ Hlth, Porto, Portugal Norwegian Inst Publ Hlth, Oslo, Norway Univ Massachusetts, Amherst, MA 01003 USA Abt Associates Inc, Kathmandu, Nepal Univ Iceland, Reykjavik, Iceland Univ Yaounde I, Yaounde, Cameroon Univ Fed Pelotas, Pelotas, RS, Brazil Univ Med 1, Yangon, Myanmar Banska Bystrica Reg Author Publ Hlth, Banska Bystrica, Slovakia Tel Aviv Univ, Tel Aviv, Israel Hebrew Univ Jerusalem, Jerusalem, Israel Univ Porto, Med Sch, Porto, Portugal Neyshabur Univ Med Sci, Neyshabur, Iran Res Inst Endocrine Sci, Tehran, Iran Indian Council Med Res, New Delhi, India Natl Inst Publ Hlth, Copenhagen, Denmark Univ Sci & Technol, Sanaa, Yemen Med Univ Lodz, Lodz, Poland Med Univ Gdansk, Gdansk, Poland Univ Autonoma Madrid CIBERESP, Madrid, Spain Univ Rzeszow, Rzeszow, Poland Univ Palermo, Palermo, Italy Pan Amer Hlth Org, Washington, DC USA Mohammed V Univ Rabat, Rabat, Morocco Univ Pernambuco, Recife, PE, Brazil Baqai Inst Diabetol & Endocrinol, Karachi, Pakistan Univ Fed Santa Catarina, Florianopolis, SC, Brazil Dalhousie Univ, Halifax, NS, Canada Jordan Univ Sci & Technol, Irbid, Jordan Univ Fed Maranhao, Sao Luis, Brazil Farabi Kazakh Natl Univ, Alma Ata, Kazakhstan Univ Coll Dublin, Dublin, Ireland Christian Med Coll & Hosp, Vellore, Tamil Nadu, India Fed Minist Social Affairs Hlth Care & Consumer Pr, Vienna, Austria Cafam Univ Fdn, Bogota, Colombia Kazakh Natl Med Univ, Alma Ata, Kazakhstan Lithuanian Univ Hlth Sci, Kaunas, Lithuania Univ Sao Paulo, Sao Paulo, Brazil BJ Med Coll, Ahmadabad, Gujarat, India Chirayu Med Coll, New Delhi, India Sunder Lal Jain Hosp, Delhi, India Hosp Sick Children, Toronto, ON, Canada Shandong Univ Tradit Chinese Med, Jinan, Peoples R China Shanghai Jiao Tong Univ, Sch Med, Shanghai, Peoples R China Univ Republica, Montevideo, Uruguay Inst Med Res & Med Plant Studies, Yaounde, Cameroon Ufa Eye Res Inst, Ufa, Russia Nepal Hlth Res Council, Kathmandu, Nepal Univ Montenegro, Niksic, Montenegro Univ Southern Denmark, Copenhagen, Denmark Univ Gothenburg, Gothenburg, Sweden Univ Fed Rio de Janeiro, Rio De Janeiro, Brazil Natl Inst Publ Hlth & Environm, Bilthoven, Netherlands Univ Turin, Turin, Italy UCL, London, England Liverpool John Moores Univ, Liverpool, Merseyside, England Nanyang Technol Univ Singapore, Singapore, Singapore German Inst Human Nutr, Potsdam, Germany Endocrinol Res Ctr, Moscow, Russia Ctr Educ Med & Invest Clin, Buenos Aires, DF, Argentina IRCCS Neuromed, Pozzilli, Italy Toulouse Univ, Sch Med, Toulouse, France Univ Zurich, Zurich, Switzerland Univ Hosp KU Leuven, Leuven, Belgium Flemish Agcy Care & Hlth, Brussels, Belgium Univ Ghent, Ghent, Belgium FrieslandCampina, Amersfoort, Netherlands Univ Cent Venezuela, Caracas, Venezuela Bielefeld Univ, Bielefeld, Germany World Hlth Org Reg Off Europe, Moscow, Russia German Canc Res Ctr, Heidelberg, Germany Fred Hollows Fdn, Auckland, New Zealand Univ Med & Pharm Bucharest, Bucharest, Romania Swansea Univ, Swansea, W Glam, Wales Univ Coll Copenhagen, Copenhagen, Denmark Inst Publ Hlth, Tirana, Albania Munster Technol Univ, Cork, Ireland Univ La Laguna, Tenerife, Spain Univ Malta, Msida, Malta Vanderbilt Univ, 221 Kirkland Hall, Nashville, TN 37235 USA Minist Hlth, Tongatapu, Tonga Canadian Fitness & Lifestyle Res Inst, Ottawa, ON, Canada Hosp Santa Maria, Lisbon, Portugal Istanbul Univ Cerrahpasa, Istanbul, Turkey Univ Fed Juiz de Fora, Juiz De Fora, Brazil Minist Publ Hlth, Asuncion, Paraguay Natl Inst Publ Hlth, Prague, Czech Republic Gaetano Fucito Hosp, Mercato San Severino, Italy Univ Groningen, Groningen, Netherlands Karolinska Inst, Huddinge, Sweden Ctr Estudios Nutr Infantil, Buenos Aires, DF, Argentina Univ Porto, Porto, Portugal Univ Zaragoza, Zaragoza, Spain Santiago de Compostela Univ, Santiago De Compostela, Spain Minist Hlth, Ankara, Turkey Council Agr Res & Econ, Rome, Italy Fed Univ Rio Grande, Rio Grande, Brazil India Diabet Res Fdn, Chennai, Tamil Nadu, India Duke NUS Med Sch, Singapore, Singapore ICMR Natl Inst Med Stat, New Delhi, India Singapore Eye Res Inst, Singapore, Singapore Acad Sinica, Taipei, Taiwan Capital Inst Pediat, Beijing, Peoples R China Duke Univ, Durham, NH USA Kailuan Gen Hosp, Tangshan, Peoples R China Univ Oxford, Oxford, England Ahvaz Jundishapur Univ Med Sci, Ahvaz, Iran Gertner Inst Epidemiol & Hlth Policy Res, Ramat Gan, Israel Natl Ctr Publ Hlth & Anal, Sofia, Bulgaria Univ Fribourg, Fribourg, Switzerland Minist Hlth & Welf, Taipei, Taiwan CIBER Epidemiol & Salud Publ, Murcia, Spain Univ Southern Denmark, Odense, Denmark Univ Estadual Paulista, Presidente Prudente, Brazil Med Univ Silesia, Katowice, Poland Charles Univ Prague, Prague, Czech Republic Primary Hlth Care, Floriana, Malta Carol Davila Univ Med & Pharm, Bucharest, Romania Katholieke Univ Leuven, Leuven, Belgium STAT Canada, Ottawa, ON, Canada UMR CNRS MNHN 7206 Ecoanthropol, Marseille, France Agcy Prevent & Social Med, Bregenz, Austria Univ Southampton, Southampton, Hants, England Inst Pasteur, Lille, France Malawi Epidemiol & Intervent Res Unit, Lilongwe, Malawi CIBEROBN, Madrid, Spain Univ Debrecen, Debrecen, Hungary Univ Med & Pharm Carol Davila, Bucharest, Romania Univ Fed Rio Grande do Sul, Porto Alegre, RS, Brazil CNR, Reggio Di Calabria, Italy Eftimie Murgu Univ Resita, Resita, Romania Spanish Agcy Food Safety & Nutr, Madrid, Spain Indian Stat Inst, Kolkata, India Tabriz Hlth Serv Management Res Ctr, Tabriz, Iran Geneva Univ Hosp, Geneva, Switzerland Sciensano, Brussels, Belgium Univ Med Ctr Groningen, Groningen, Netherlands Natl Res Ctr Prevent Med, Moscow, Russia Innovating Hlth Int, Port Au Prince, Haiti Univ Montreal, Montreal, PQ, Canada French Natl Res Inst Sustainable Dev, Montpellier, France French Publ Hlth Agcy, St Maurice, France Mediterranea Cardioctr, Naples, Italy Univ Vale Rio dos Sinos, Sao Leopoldo, Brazil Natl Inst Hyg Epidemiol & Microbiol, Havana, Cuba Natl Council Sci & Tech Res, Buenos Aires, DF, Argentina Minist Hlth & Med Educ, Tehran, Iran Univ Novi Sad, Novi Sad, Serbia Natl Inst Nutr, Hanoi, Vietnam Univ Queensland, Brisbane, Qld, Australia Ist Super Sanita, Rome, Italy Univ Cuenca, Cuenca, Ecuador Helmholtz Zentrum Munchen, Munich, Germany Univ Tehran Med Sci, Tehran, Iran Minist Sante & Hyg Publ, Abidjan, Cote Ivoire Univ Hosp Dusseldorf, Dusseldorf, Germany Natl Inst Cardiol, Warsaw, Poland Beijing Ctr Dis Prevent & Control, Beijing, Peoples R China UMI 3189 ESS, Marseille, France Beth Israel Deaconess Med Ctr, Boston, MA 02215 USA Scuola Super Sant Anna, Pisa, Italy Univ Latvia, Riga, Latvia Minist Hlth & Med Serv, Gizo, Solomon Islands Hormozgan Univ Med Sci, Bandar Abbas, Iran Univ Benin, Benin, Nigeria Univ Skovde, Skovde, Sweden Natl Inst Nutr & Food Technol, Tunis, Tunisia Univ West Indies, Kingston, Jamaica Univ Calif Davis, Davis, CA 95616 USA Univ Stellenbosch, Cape Town, South Africa Univ Duisburg Essen, Duisburg, Germany Karadeniz Tech Univ, Trabzon, Turkey Univ Helsinki, Helsinki, Finland Mashhad Univ Med Sci, Mashhad, Razavi Khorasan, Iran Rafsanjan Univ Med Sci, Rafsanjan, Iran Queens Univ Belfast, Belfast, Antrim, North Ireland Tabriz Univ Med Sci, Tabriz, Iran Fasa Univ Med Sci, Fasa, Iran Shiraz Univ Med Sci, Shiraz, Iran Baqai Med Univ, Karachi, Pakistan Ctr Salud Villanueva Norte, Badajoz, Spain Hosp Don Benito Villanueva Serena, Badajoz, Spain Minist Hlth, Buenos Aires, DF, Argentina Univ Insubria, Varese, Italy Pontificia Univ Catolica Chile, Santiago, Chile Inst Mother & Child Hlth, Warsaw, Poland Swiss Trop & Publ Hlth Inst, Basel, Switzerland Univ Tartu, Tartu, Estonia Univ Sains Malaysia, Kota Baharu, Kelantan, Malaysia Umea Univ, Umea, Sweden Univ Fed Sao Paulo, Sao Paulo, Brazil Hosp Univ Son Espases, Palma De Mallorca, Spain Hosp Clin Porto Alegre, Porto Alegre, RS, Brazil Univ Sumatera Utara, Medan, Indonesia Kindai Univ, Osaka, Japan Kyoto Univ, Kyoto, Japan Med Univ Warsaw, Warsaw, Poland Jagiellonian Univ, Med Coll, Krakow, Poland Minist Salute DG Prevenz Sanit, Rome, Italy Univ Catania, Catania, Italy CIBER Epidemiol & Salud Publ, Alicante, Spain Africa Hlth Res Inst, Mtubatuba, South Africa Univ Geneva, Med Sch, Geneva, Switzerland CIBER Epidemiol & Salud Publ, Barcelona, Spain Univ Fed Minas Gerais, Belo Horizonte, MG, Brazil Univ Utrecht, Utrecht, Netherlands Wageningen Univ, Wageningen, Netherlands Univ Adelaide, Adelaide, SA, Australia Lund Univ, Lund, Sweden Aristotle Univ Thessaloniki, Thessaloniki, Greece Inst Natl Sante & Rech Med, Villejuif, France Minist Hlth, Jerusalem, Israel McGill Univ, Montreal, PQ, Canada Gasol Fdn, Barcelona, Spain PASs Hirszfeld Inst Immunol & Expt Therapy, Wroclaw, Poland Univ Politecn Madrid, Madrid, Spain St Annes Univ Hosp, Brno, Czech Republic Natl Inst Publ Hlth, Cuernavaca, Morelos, Mexico Ctr Estudios Diabet AC, Mexico City, DF, Mexico Univ Autonoma Santo Domingo, Santo Domingo, Dominican Rep Minist Hlth, Lisbon, Portugal Inst Clin & Expt Med, Prague, Czech Republic Childrens Mem Hlth Inst, Warsaw, Poland Natl Ctr Cardiovasc Dis, Beijing, Peoples R China Univ Ferrara, Ferrara, Italy Author Sanitaria San Marino, San Marino, CA USA Iceland Heart Assoc, Kopavogur, Iceland Univ Icesi, Cali, Colombia Univ Estadual Montes Claros, Montes Claros, MG, Brazil Kings Coll London, London, England Int Agcy Res Canc, Lyon, France Capital Med Univ, Beijing, Peoples R China Capital Med Univ, Beijing Tongren Hosp, Beijing, Peoples R China Healis Sekhsaria Inst Publ Hlth, Navi Mumbai, India Eternal Heart Care Ctr & Res Inst, Jaipur, Rajasthan, India Univ Ibadan, Ibadan, Nigeria Inst Clin Effectiveness & Hlth Policy, Buenos Aires, DF, Argentina Natl Hlth Insurance Serv, Wonju, South Korea Prevent Metab Disorders Res Ctr, Tehran, Iran Res & Educ Inst Child Hlth, Nicosia, Cyprus Danish Canc Soc Res Ctr, Copenhagen, Denmark Univ West Indies, Cave Hill, Barbados Kermanshah Univ Med Sci, Kermanshah, Iran Univ Eastern Finland, Kuopio, Finland Beijing Inst Ophthalmol, Beijing, Peoples R China Int Hellen Univ, Thessaloniki, Greece Kyushu Univ, Fukuoka, Japan Univ Bergen, Bergen, Norway Tulane Univ, New Orleans, LA USA Natl Res Inst Hlth & Family Planning, Beijing, Peoples R China Chinese Ctr Dis Control & Prevent, Beijing, Peoples R China Univ Pecs, Pecs, Hungary Danish Hlth Author, Copenhagen, Denmark Joep Lange Inst, Amsterdam, Netherlands Univ Fed Pernambuco, Recife, PE, Brazil Oulu Univ Hosp, Oulu, Finland Univ Autonoma Bucaramanga, Bucaramanga, Colombia Swiss Fed Inst Technol, Zurich, Switzerland Chron Dis Res Ctr, Tehran, Iran Univ Hong Kong, Hong Kong, Peoples R China Chinese Univ Hong Kong, Hong Kong, Peoples R China Univ Western Australia, Perth, WA, Australia Kingston Hlth Sci Ctr, Kingston, ON, Canada Fdn Oftalmol Santander, Bucaramanga, Colombia Univ Oran 1, Oran, Algeria Nay Pyi Taw, Independent Publ Hlth Specialist, Nay Pyi Taw, Myanmar Minist Hlth & Sports, Nay Pyi Taw, Myanmar Peking Univ, Beijing, Peoples R China Natl Inst Publ Hlth, Bucharest, Romania Vrije Univ Amsterdam Med Ctr, Amsterdam, Netherlands Amer Univ Beirut, Beirut, Lebanon CIBER Epidemiol & Salud Publ, San Sebastian, Spain Cairo Univ, Cairo, Egypt Erasmus MC, Rotterdam, Netherlands Med Univ Varna, Varna, Bulgaria Univ Tokyo, Tokyo, Japan Deakin Univ, Geelong, Vic, Australia Tokyo Metropolitan Inst Gerontol, Tokyo, Japan Hadassah Univ Med Ctr, Jerusalem, Israel Norwegian Univ Sci & Technol, Trondheim, Norway Univ Melbourne, Melbourne, Vic, Australia Sports Univ Tirana, Tirana, Albania Univ Hosp Ctr Zagreb, Zagreb, Croatia Univ Zagreb, Sch Med, Zagreb, Croatia Heart Fdn, Melbourne, Vic, Australia Guangzhou 12th Hosp, Guangzhou, Peoples R China Univ Eugenio Maria Hostos, Santo Domingo, Dominican Rep Simon Fraser Univ, Burnaby, BC, Canada Inst Mol & Clin Ophthalmol Basel, Basel, Switzerland Univ New South Wales, Sydney, NSW, Australia World Hlth Org Country Off, Delhi, India Guilan Univ Med Sci, Rasht, Iran Inst Publ Hlth, Belgrade, Serbia Univ Opole, Opole, Poland Finnish Inst Hlth & Welf, Helsinki, Finland Gulu Univ, Gulu, Uganda Univ Crete, Iraklion, Greece Sri Venkateswara Univ, Tirupati, Andhra Pradesh, India Sree Chitra Tirunal Inst Med Sci & Technol, Trivandrum, Kerala, India Hellen Med Assoc Obes, Athens, Greece Natl & Kapodistrian Univ Athens, Athens, Greece Maharajgunj Med Campus, Kathmandu, Nepal Aarhus Univ, Aarhus, Denmark Johns Hopkins Bloomberg Sch Publ Hlth, Baltimore, MD USA Pennington Biomed Res Ctr, Baton Rouge, LA USA Natl Inst Epidemiol, Chennai, Tamil Nadu, India Univ Munster, Munster, Germany Israel Ctr Dis Control, Ramat Gan, Israel Res Inst Primordial Prevent Noncommunicable Dis, Esfahan, Iran Amsterdam UMC Publ Hlth Res Inst, Amsterdam, Netherlands South African Med Res Council, Cape Town, South Africa Kyrgyz State Med Acad, Bishkek, Kyrgyzstan Res Inst Child Nutr, Dortmund, Germany Shahid Beheshti Univ Med Sci, Tehran, Iran Univ Cambridge, Cambridge, England Mazandaran Univ Med Sci, Sari, Iran Hypertens Res Ctr, Esfahan, Iran Med Univ Innsbruck, Innsbruck, Austria Muhimbili Univ Hlth & Allied Sci, Dar Es Salaam, Tanzania Yonsei Univ, Coll Med, Seoul, South Korea Natl Canc Ctr, Goyang Si, South Korea Univ Coll South Denmark, Haderslev, Denmark Stat Austria, Vienna, Austria BP Koirala Inst Hlth Sci, Dharan, Nepal Univ Vienna, Vienna, Austria Tartu Univ Clin, Tartu, Estonia Kansai Med Univ, Hirakata, Osaka, Japan Hungarian Sch Sport Federat, Budapest, Hungary Minist Hlth & Qual Life, Port Louis, Mauritius Univ Hosp Ulm, Ulm, Germany North West Univ, Potchefstroom, South Africa Univ Jyvaskyla, Jyvaskyla, Finland Inst Publ Hlth, Podgorica, Montenegro Amrita Inst Med Sci, Cochin, Kerala, India Inst Endocrinol, Prague, Czech Republic All India Inst Med Sci, New Delhi, India African Populat & Hlth Res Ctr, Nairobi, Kenya Hanoi Univ Publ Hlth, Hanoi, Vietnam Hassan First Univ Settat, Settat, Morocco Minist Hlth, Algiers, Algeria Harokopio Univ, Athens, Greece Sahlgrens Acad, Gothenburg, Sweden Endocrinol & Metab Res Ctr, Tehran, Iran Univ Publ Hlth, Yangon, Myanmar Int Food Policy Res Inst, Dakar, Senegal Natl Univ Singapore, Singapore, Singapore Hong Kong Polytech Univ, Hong Kong, Peoples R China Tampere Univ Hosp, Tampere, Finland Univ Douala, Douala, Cameroon Univ Cape Town, Cape Town, South Africa Natl Inst Hlth Dev, Tallinn, Estonia West Virginia Univ, Morgantown, WV 26506 USA Oswaldo Cruz Fdn Rene Rachou Res Inst, Belo Horizonte, MG, Brazil Natl Taiwan Univ, Taipei, Taiwan Univ Chinese Acad Sci, Shanghai, Peoples R China Uppsala Univ, Uppsala, Sweden Taipei Med Univ, Taipei, Taiwan Sports Med Ctr Minho, Braga, Portugal Univ San Martin Porres, Lima, Peru Consejeria Sanidad Junta Castilla & Leon, Valladolid, Spain Univ Fed Ciencias Saude Porto Alegre, Porto Alegre, RS, Brazil Norrbotten Cty Council, Lulea, Sweden Ilembula Lutheran Hosp, Ilembula, Iran Univ Fed Ouro Preto, Ouro Preto, Brazil Univ Coimbra, Coimbra, Portugal Coimbra Univ Hosp Ctr, Coimbra, Portugal Inst Neurosci Natl Res Council, Padua, Italy Baker Heart & Diabet Inst, Melbourne, Vic, Australia Agr Univ Athens, Athens, Greece Hosp Israelita Albert Einstein, Sao Paulo, Brazil SB RAS Fed Res Ctr Inst Cytol & Genet, Novosibirsk, Russia Univ Otago, Dunedin, New Zealand Univ Padua, Padua, Italy Hellen Mediterranean Univ, Siteia, Greece Loughborough Univ, Loughborough, Leics, England Minist Hlth, Nicosia, Cyprus Lausanne Univ Hosp, Lausanne, Switzerland Escola Nacl Saude Publ Sergio Arouca, Rio De Janeiro, Brazil CIBERCV, Madrid, Spain Mary Immaculate Coll, Limerick, Ireland Emory Univ, Atlanta, GA 30322 USA Hungarian Soc Sports Med, Budapest, Hungary Paracelsus Med Univ, Salzburg, Austria Univ Estadual Centro Oeste, Guarapuava, Brazil Sherikashmir Inst Med Sci, Srinagar, India UiT Arctic Univ Norway, Tromso, Norway ICMR Natl Ctr Dis Informat & Res, Bengaluru, India Cape Peninsula Univ Technol, Cape Town, South Africa Brown Univ, Providence, RI 02912 USA London Sch Hyg & Trop Med, London, England Univ Edinburgh, Edinburgh, Midlothian, Scotland Shahid Sadoughi Univ Med Sci, Tehran, Iran Inst Natl Sante & Rech Med, Lille, France Robert Koch Inst, Berlin, Germany Univ Lisbon, Lisbon, Portugal Womens Social & Hlth Studies Fdn, Trivandrum, Kerala, India Democritus Univ, Alexandroupolis, Greece Grigore T Popa Univ Med & Pharm, Iasi, Romania Univ Firenze, Florence, Italy Ain Shams Univ, Cairo, Egypt Isfahan Cardiovasc Res Ctr, Esfahan, Iran Univ Strasbourg, Strasbourg, France Mulago Hosp, Kampala, Uganda Univ San Carlos, Guatemala City, Guatemala Univ Limpopo, Sovenga, South Africa Univ Med Sci Cienfuegos, Cienfuegos, Cuba Royal Coll Surgeons Ireland, Dublin, Ireland Trobe Univ, Melbourne, Vic, Australia Int Inst Mol & Cell Biol, Warsaw, Poland Inst Conmemorat Gorgas Estudios Salud, Panama City, Panama World Hlth Org Country Off, Lilongwe, Malawi Dept Publ Hlth, Nay Pyi Taw, Myanmar Univ Brescia, Brescia, Italy Minist Hlth & Social Protect, Dushanbe, Tajikistan Univ Limerick, Limerick, Ireland Croatian Inst Publ Hlth, Zagreb, Croatia Univ Kebangsaan Malaysia, Kuala Lumpur, Malaysia Bushehr Univ Med Sci, Bushehr, Iran Ulm Univ, Ulm, Germany Dept Stat, Kuala Lumpur, Malaysia Kobe Univ, Kobe, Hyogo, Japan Suraj Eye Inst, Nagpur, Maharashtra, India UNICEF, Yaounde, Cameroon Univ Med Greifswald, Greifswald, Germany Karolinska Inst, Stockholm, Sweden Natl Inst Hyg & Epidemiol, Hanoi, Vietnam Univ Med & Pharm Ho Chi Minh City, Ho Chi Minh City, Vietnam Hanoi Med Univ, Hanoi, Vietnam LifeDoc Hlth, Memphis, TN USA Heartfile, Islamabad, Pakistan Eastern Mediterranean Publ Hlth Network, Amman, Jordan Univ Manchester, Manchester, Lancs, England State Univ Med & Pharm, Kishinev, Moldova Tachikawa Gen Hosp, Nagaoka, Niigata, Japan Korea Ctr Dis Control & Prevent, Cheongju, South Korea Japan Wildlife Res Ctr, Tokyo, Japan Istanbul Univ, Istanbul, Turkey Minist Hlth, Bandar Seri Begawan, Brunei Univ Fed Parana, Curitiba, Parana, Brazil Univ Madeira, Funchal, Portugal Univ Puerto Rico, San Juan, PR 00936 USA Ctr Clin Res & Prevent, Glostrup, Denmark MRC Lifecourse Epidemiol Unit, Southampton, Hants, England Kwame Nkrumah Univ Sci & Technol, Kumasi, Ghana UniSante, Lausanne, Switzerland Inst Canc Res, Prevent & Clin Network, Florence, Italy Univ Wisconsin, Madison, WI USA Heidelberg Univ, Heidelberg, Germany IRCCS Ente Osped Specializzato Gastroenterol S de, Bari, Italy Zayed Univ, Abu Dhabi, U Arab Emirates Catholic Univ Daegu, Daegu, South Korea Univ Med Pharm Sci & Technol Targu Mures, Targu Mures, Romania Jivandeep Hosp, Anand, Gujarat, India South African Med Res Council, Durban, South Africa Natl Dent Care Ctr Singapore, Singapore, Singapore Univ Malaga, Malaga, Spain Univ Hosp Varese, Varese, Italy Vietnam Natl Heart Inst, Hanoi, Vietnam Clin Med Avanzada Dr Abel Gonzalez, Santo Domingo, Dominican Rep Leibniz Inst Prevent Res & Epidemiol BIPS, Bremen, Germany Univ Sarajevo, Sarajevo, Bosnia & Herceg Cardiovasc Prevent Ctr Udine, Udine, Italy Pisa Univ Hosp, Pisa, Italy Minist Hlth & Med Serv, Honiara, Solomon Islands Publ Hlth Agcy Catalonia, Barcelona, Spain Inst Hosp del Mar Invest Med, Barcelona, Spain Ardabil Univ Med Sci, Ardebil, Iran Ctr Dis Prevent & Control, Riga, Latvia Alborz Univ Med Sci, Karaj, Iran Minist Hlth, Hanoi, Vietnam Pure Earth, Dhaka, Bangladesh Inst Epidemiol Dis Control & Res, Dhaka, Bangladesh Univ Turku, Turku, Finland Inst Univ Invest Atenc Primaria Jordi Gol, Girona, Spain Univ Putra Malaysia, Serdang, Malaysia Univ Malaya, Kuala Lumpur, Malaysia Sotiria Hosp, Athens, Greece Univ Valencia, Valencia, Spain Univ Philippines, Manila, Philippines Slovak Acad Sci, Bratislava, Slovakia Nutr Res Fdn, Barcelona, Spain Minas Gerais State Secretariat Hlth, Belo Horizonte, MG, Brazil CS S Agustin Ibsalut, Palma De Mallorca, Spain Amsterdam Inst Global Hlth & Dev, Amsterdam, Netherlands Natl Inst Hlth Doutor Ricardo Jorge, Lisbon, Portugal Univ Nove Julho, Sao Paulo, Brazil Publ Hlth Agcy Canada, Ottawa, ON, Canada Canarian Hlth Serv, Tenerife, Spain Univ Ind Santander, Bucaramanga, Colombia Assoc Calabrese Epatol, Reggio Di Calabria, Italy Univ Minho, Braga, Portugal Fiji Natl Univ, Suva, Fiji Spanish Nutr Fdn, Madrid, Spain Publ Hlth Agcy Sweden, Stockholm, Sweden Inst Food Sci Natl Res Council, Avellino, Italy Sitaram Bhartia Inst Sci & Res, New Delhi, India Kindergarten Avlonari, Evia, Greece Natl Inst Hlth, Lima, Peru Minist Hlth, Jakarta, Indonesia Catalan Dept Hlth, Barcelona, Spain Biodonostia Hlth Res Inst, San Sebastian, Spain Inst Saude Ambiental, Lisbon, Portugal South Karelia Social & Hlth Care Dist, Lappeenranta, Finland McMaster Univ, Hamilton, ON, Canada Natl Canc Ctr, Tokyo, Japan Univ Sao Paulo Clin Hosp, Sao Paulo, Brazil Hosp Italiano Buenos Aires, Buenos Aires, DF, Argentina Rigshosp, Copenhagen, Denmark Acad Med Ctr Univ Amsterdam, Amsterdam, Netherlands Ctr Oral Hlth Serv & Res Midnorway, Trondheim, Norway Lagos State Univ, Coll Med, Lagos, Nigeria Univ Las Palmas Gran Canaria, Las Palmas Gran Canaria, Spain Comenius Univ, Bratislava, Slovakia Natl Ctr Dis Control & Publ Hlth, Tbilisi, Georgia Nippon Med Sch, Tokyo, Japan Sungkyunkwan Univ, Seoul, South Korea Finnish Inst Occupat Hlth, Helsinki, Finland Publ Hlth Promot & Dev Org, Kathmandu, Nepal St Vincents Hosp, Sydney, NSW, Australia Nes Municipal, Aarnes, Norway Hlth Polytech Jakarta II Inst, Jakarta, Indonesia Diponegoro Univ, Semarang, Indonesia Univ Bari, Bari, Italy Inst Reg Sante Publ, Ouidah, Benin Univ Bordeaux, Bordeaux, France Oslo Metropolitan Univ, Oslo, Norway Inst Publ Hlth, Skopje, North Macedonia Univ Leuven, Leuven, Belgium Lamprecht & Stamm Sozialforsch & Beratung AG, Zurich, Switzerland Inst Natl Sante & Rech Med, Nancy, France Univ Bonn, Bonn, Germany Sotiria Hosp, Sotiria, Greece Kalina Malina Kindergarten, Pazardjik, Bulgaria Jikei Univ, Sch Med, Tokyo, Japan Fu Jen Catholic Univ, Taipei, Taiwan Natl Stat Off, Praia, Cape Verde Natl Inst Publ Hlth Natl Inst Hyg, Warsaw, Poland Deakin Univ, Sydney, NSW, Australia Sci Res Inst Maternal & Child Hlth, Ashkhabad, Turkmenistan Univ Lincoln, Lincoln, England Minist Hlth, Amman, Jordan UNICEF, Niamey, Niger Cent Univ Kerala, Kasaragod, India Amsterdam Publ Hlth Res Inst, Amsterdam, Netherlands Hlth Serv Murcia, Murcia, Spain Inst Invest Sanitaria Illes Balears, Menorca, Spain Univ Bologna, Bologna, Italy Hellen Hlth Fdn, Athens, Greece Govt Med Coll, Bhavnagar, Gujarat, India Sefako Makgatho Hlth Sci Univ, Ga Rankuwa, South Africa Addis Ababa Univ, Addis Ababa, Ethiopia Minist Hlth, Wellington, New Zealand Univ Ctr Occidental Lisandro Alvarado, Barquisimeto, Venezuela Meharry Med Coll, Nashville, TN 37208 USA Dokuz Eylul Univ, Izmir, Turkey Univ Tampere, Tays Eye Ctr, Tampere, Finland Sabiha Gokcen Ilkokulu, Ankara, Turkey Polytech Inst Porto, Porto, Portugal Icahn Sch Med Mt Sinai, New York, NY 10029 USA Univ CEU San Pablo, Madrid, Spain CNR, Pisa, Italy Univ Miguel Hernandez, Alicante, Spain Vrije Univ Amsterdam, Amsterdam, Netherlands Sunflower Nursery Sch, Craiova, Romania North Karelia Ctr Publ Hlth, Joensuu, Finland Univ Witwatersrand, Johannesburg, South Africa Med Univ Vienna, Vienna, Austria Cork Inst Technol, Cork, Ireland Inst Med Res, Kuala Lumpur, Malaysia Xinjiang Med Univ, Urumqi, Peoples R China Shanghai Educ Dev Co Ltd, Shanghai, Peoples R China Minist Hlth, Auckland, New Zealand Rebro Univ, Rebro, Sweden St Georges Univ London, London, England Univ Indonesia, Jakarta, Indonesia Minist Agr & Rural Affairs, Inst Food & Nutr Dev, Beijing, Peoples R China Fudan Univ, Childrens Hosp, Shanghai, Peoples R China Univ Cyprus, Nicosia, Cyprus Niigata Univ, Niigata, Japan Int Med Univ, Shah Alam, Selangor, Malaysia Hellen Mediterranean Univ, Iraklion, Greece Iran Univ Med Sci, Tehran, Iran Ctr Diabet & Endocrine Care, Srinagar, India Peking Univ First Hosp, Beijing, Peoples R China Jiangsu Prov Ctr Dis Control & Prevent, Nanjing, Peoples R China Sun Yat Sen Univ, Guangzhou, Peoples R China West Kazakhstan Med Univ, Aktobe, Kazakhstan Inner Mongolia Med Univ, Hohhot, Peoples R China Przedszkole 81, Warsaw, Poland Univ Estadual Paulista, Presidente Prudente, Brazil

Upon pathogen infection, receptors in plants will activate a localized immune response, the effector-triggered immunity (ETI), and a systemic immune response, the systemic acquired response (SAR). Infection also induces oscillations in the redox environment of plant cells, triggering response mechanisms involving sensitive cysteine residues that subsequently alter protein function. Arabidopsis thaliana thimet oligopeptidases TOP1 and TOP2 are required for plant defense against pathogens and the oxidative stress response. Herein, we evaluated the biochemical attributes of TOP isoforms to determine their redox sensitivity using ex vivo Escherichia coli cultures and recombinant proteins. Moreover, we explored the link between their redox regulation and plant immunity in wild-type and mutant Arabidopsis lines. These analyses revealed that redox regulation of TOPs occurs through two mechanisms: (1) oxidative dimerization of full-length TOP1 via intermolecular disulfides engaging cysteines in the N-terminal signal peptide, and (2) oxidative activation of all TOPs via cysteines that are unique and conserved. Further, we detected increased TOP activity in wild-type plants undergoing ETI or SAR following inoculation with Pseudomonas syringae strains. Mutants unable to express the chloroplast NADPH-dependent thioredoxin reductase C (NTRC) showed elevated TOP activity under unstressed conditions and were SAR-incompetent. A top1top2 knockout mutant challenged with P. syringae exhibited misregulation of ROS-induced gene expression in pathogen-inoculated and distal tissues. Furthermore, TOP1 and TOP2 could cleave a peptide derived from the immune component ROC1 with distinct efficiencies at common and specific sites. We propose that Arabidopsis TOPs are thiol-regulated peptidases active in redox-mediated signaling of local and systemic immunity.

Background The ever-increasing prevalence of diabetes and associated comorbidities serves to highlight the necessity of biologically relevant small-animal models to investigate its etiology, pathology and treatment. Although the C57BL/6 J model is amongst the most widely used mouse model due to its susceptibility to diet-induced obesity (DIO), there are a number of limitations namely [1] that unambiguous fasting hyperglycemia can only be achieved via dietary manipulation and/or chemical ablation of the pancreatic beta cells. [2] Heterogeneity in the obesogenic effects of hypercaloric feeding has been noted, together with sex-dependent differences, with males being more responsive. The KK mouse strain has been used to study aspects of the metabolic syndrome and prediabetes. We recently conducted a study which characterized the differences in male and female glucocentric parameters between the KK/HlJ and C57BL/6 J strains as well as diabetes-related behavioral differences (Inglis et al. 2019). In the present study, we further characterize these models by examining strain- and sex-dependent differences in pancreatic and adrenal gene expression using Affymetrix microarray together with endocrine-associated serum analysis. Results In addition to strain-associated differences in insulin tolerance, we found significant elevations in KK/HlJ mouse serum leptin, insulin and aldosterone. Additionally, glucagon and corticosterone were elevated in female mice of both strains. Using 2-factor ANOVA and a significance level set at 0.05, we identified 10,269 pancreatic and 10,338 adrenal genes with an intensity cut-off of ≥2.0 for all 4 experimental groups. In the pancreas, gene expression upregulated in the KK/HlJ strain related to increased insulin secretory granule biofunction and pancreatic hyperplasia, whereas ontology of upregulated adrenal differentially expressed genes (DEGs) related to cell signaling and neurotransmission. We established a network of functionally related DEGs commonly upregulated in both endocrine tissues of KK/HlJ mice which included the genes coding for endocrine secretory vesicle biogenesis and regulation: PCSK2, PCSK1N, SCG5, PTPRN, CHGB and APLP1. We also identified genes with sex-biased expression common to both strains and tissues including the paternally expressed imprint gene neuronatin. Conclusion Our novel results have further characterized the commonalities and diversities of pancreatic and adrenal gene expression between the KK/HlJ and C57BL/6 J strains as well as differences in serum markers of endocrine physiology.

The diagnosis of indolent systemic mastocytosis can be quite a challenge due to its wide spectrum of clinical manifestations. We are reporting a case of misdiagnosed indolent systemic mastocytosis in a 41-year-old male that has been wrongly treated first as a Non-Hodgkin lymphoma. The patient had generalized lymphadenopathy and eosinophilia, which are rare manifestations of indolent systemic mastocytosis. The chronic neglected pruritus and elevated tryptase levels along with histological findings were the main clues that have led us to the final diagnosis of indolent systemic mastocytosis. Later during the coronavirus disease 2019 pandemic, the patient presented with symptoms that are common between coronavirus disease 2019 and mast cell activation syndrome. Coronavirus disease 2019 was confirmed with a polymerase chain reaction test. There were no clear guidelines for managing a case like ours, so our management plan was based on the latest recommendations published at that time., Highlights • The difficulty of diagnosing indolent systemic mastocytosis comes from its exceptional diversity. • Little is known about management indolent systemic mastocytosis and coronavirus disease 2019. • Definitive evidence is lacking whether mast cells play a defending or accelerating role in coronavirus disease 2019.

Significance Phase separation of proteins is increasingly thought to play a fundamental role in biological processes. Recent studies show that tau protein phase separates, but the biological significance is unknown since artificial crowding agents are typically used and the resulting tau is not toxic. We now demonstrate that TIA1 potentiates RNA-mediated phase separation of tau, thereby enabling a process that occurs at physiological concentrations and also directs the formation of biologically active, highly neurotoxic oligomeric tau. Coordinated phase separation of functionally related proteins provides a general mechanism through which membraneless organelles can direct biological activities., Tau protein plays an important role in the biology of stress granules and in the stress response of neurons, but the nature of these biochemical interactions is not known. Here we show that the interaction of tau with RNA and the RNA binding protein TIA1 is sufficient to drive phase separation of tau at physiological concentrations, without the requirement for artificial crowding agents such as polyethylene glycol (PEG). We further show that phase separation of tau in the presence of RNA and TIA1 generates abundant tau oligomers. Prior studies indicate that recombinant tau readily forms oligomers and fibrils in vitro in the presence of polyanionic agents, including RNA, but the resulting tau aggregates are not particularly toxic. We discover that tau oligomers generated during copartitioning with TIA1 are significantly more toxic than tau aggregates generated by incubation with RNA alone or phase-separated tau complexes generated by incubation with artificial crowding agents. This pathway identifies a potentially important source for generation of toxic tau oligomers in tau-related neurodegenerative diseases. Our results also reveal a general principle that phase-separated RBP droplets provide a vehicle for coassortment of selected proteins. Tau selectively copartitions with TIA1 under physiological conditions, emphasizing the importance of TIA1 for tau biology. Other RBPs, such as G3BP1, are able to copartition with tau, but this happens only in the presence of crowding agents. This type of selective mixing might provide a basis through which membraneless organelles bring together functionally relevant proteins to promote particular biological activities.

C4b Binding Protein (C4BP) is a major fluid phase inhibitor of the classical and lectin pathways of the complement system. Complement inhibition is achieved by binding to and restricting the role of activated complement component C4b. C4BP functions as a co-factor for factor I in proteolytic inactivation of both soluble and cell surface-bound C4b, thus restricting the formation of the C3-convertase, C4b2a. C4BP also accelerates the natural decay/dissociation of the C3 convertase. This makes C4BP a prime target for exploitation by pathogens to escape complement attack, as seen in Streptococcus pyogenes or Flavivirus. Here, we examined whether C4BP can act on its own in a complement independent manner, against pathogens. C4BP bound H1N1 and H3N2 subtypes of Influenza A Virus (IAV) most likely via multiple sites in Complement Control Protein (CCP) 1-2, 4-5, and 7-8 domains of its α-chain. In addition, C4BP CCP1-2 bound H3N2 better than H1N1. C4BP bound three IAV envelope proteins: Haemagglutinin (~70 kDa), Neuraminidase (~55 kDa), and Matrix protein 1 (~25kDa). C4BP suppressed H1N1 subtype infection into the lung epithelial cell line, A549, while it promoted infection by H3N2 subtype. C4BP restricted viral entry for H1N1 but had the opposite effect on H3N2, as evident from experiments using pseudo-typed viral particles. C4BP downregulated mRNA levels of pro-inflammatory IFN-α, IL-12, and NFκB in the case of H1N1, while it promoted a pro-inflammatory immune response by upregulating IFN- α, TNF-α, RANTES, and IL-6 in the case of H3N2. We conclude that C4BP differentially modulates the efficacy of IAV entry, and hence, replication in a target cell in a strain-dependent manner, and acts as an entry inhibitor for H1N1. Thus, CCP containing complement proteins such as factor H and C4BP may have additional defense roles against IAV that do not rely on the regulation of complement activation.

Active breast cancer-associated fibroblasts (CAFs), the most influential cells in breast tumor microenvironment, express/secrete high levels of the proinvasive/metastatic interleukin-6 (IL-6). Therefore, we have tested here the effect of the IL-6 receptor (IL-6R) inhibitor tocilizumab (TCZ; Actemra) on different active breast CAFs. We have shown that TCZ potently and persistently suppresses the expression of various CAF biomarkers, namely α-SMA, SDF-1 as well as the STAT3 pathway and its downstream target AUF1. TCZ also inhibited the proliferation, migration and invasion abilities of active breast CAF cells. Additionally, TCZ repressed the ability of CAF cells in promoting epithelial-to-mesenchymal transition, and enhancing the migratory/invasive and proliferative capacities of breast cancer cells in vitro. Importantly, these findings were confirmed in orthotopic humanized breast tumors in mice. Furthermore, TCZ suppressed the expression of the pro-angiogenic factor VEGF-A and its transactivator HIF-1α in CAF cells, and consequently inhibited the angiogenic-promoting effect of active CAFs both in vitro and in orthotopic tumor xenografts. These results indicate that inhibition of the IL-6/STAT3/AUF1 pathway by TCZ can normalize active breast CAFs and suppress their paracrine pro-carcinogenic effects, which paves the way toward development of specific CAF-targeting therapy, badly needed for more efficient breast cancer treatments.

Background The WD40-repeat containing proteins, including DDB1–CUL4-associated factors (DCAFs), are abundant and conserved proteins that play important roles in different cellular processes including spermatogenesis. DCAFs are subset of WD40 family proteins that contain WDxR motif and have been proposed to function as substrate receptor for Cullin4-RING-based E3 ubiquitin ligase complexes to recruit diverse proteins for ubiquitination, a vital process in spermatogenesis. Large number of WD40 genes has been identified in different species including mouse and human. However, a systematic expression profiling of WD40 genes in different tissues of mouse and human has not been investigated. We hypothesize that large number of WD40 genes may express highly or specifically in the testis, where their expression is uniquely regulated during testis development and spermatogenesis. Therefore, the objective of this study is to mine and characterize expression patterns of WD40 genes in different tissues of mouse and human with particular emphasis on DCAF genes expressions during mouse testicular development. Results Publically available RNA sequencing (RNA seq) data mining identified 347 and 349 WD40 genes in mouse and human, respectively. Hierarchical clustering and heat map analyses of RNA seq datasets revealed differential expression patterns of WD40 genes with around 60–73% of the genes were highly or specifically expressed in testis. Similarly, around 74–83% of DCAF genes were predominantly or specifically expressed in testis. Moreover, WD40 genes showed distinct expression patterns during embryonic and postnatal testis development in mice. Finally, different germ cell populations of testis showed specific patterns of WD40 genes expression. Predicted gene ontology analyses revealed more than 80% of these proteins are implicated in cellular, metabolic, biological regulation and cell localization processes. Conclusions We have identified large number of WD40 family genes that are highly or specifically expressed in the testes of mouse and human. Moreover, WD40 genes have distinct expression patterns during embryonic and postnatal development of the testis in mice. Further, different germ cell populations within the testis showed specific patterns of WD40 genes expression. These results provide foundation for further research towards understanding the functional genomics and molecular mechanisms of mammalian testis development and spermatogenesis.

Copyright © 2020 Murugaiah, Varghese, Saleh, Tsolaki, Alrokayan, Khan, Collison, Sim, Nal, Al-Mohanna and Kishore. The complement system is an ancient innate immune defence mechanism that can recognise molecular patterns on the invading pathogens. Factor H, as an inhibitor of the alternative pathway, down-regulates complement activation on the host cell surface. Locally synthesised factor H at the site of infection/injury, including lungs, can act as a pattern recognition molecule without involving complement activation. Here, we report that factor H, a sialic acid binder, interacts with influenza A virus (IAV) and modulates IAV replication, as observed by an upregulation of matrix protein 1 (M1) expression in H3N2-infected A549 cells, while downregulating M1 in H1N1 subtype-infected cells. Far-western blot revealed that factor H binds hemagglutinin (HA, ~70kDa), neuraminidase (NA, ~60kDa), and M1 (~25kDa). IAV-induced transcriptional levels of IFN-α, TNF-α, IL-12, IL-6, IFN-α, while RANTES were reduced following factor H treatment for the H1N1 subtype at 6 h post-infection. However, for the H3N2 subtype, mRNA levels of these pro-inflammatory cytokines were enhanced. Recombinant form of vaccinia virus complement control protein (VCP), which like factor H, contains CCP modules and has complement-regulatory activity, mirrored the results obtained with factor H. Both factor H (25%) and VCP (45%) were found to reduce luciferase reporter activity in MDCK cells transduced with H1N1 pseudotyped lentiviral particles. Factor H (50%) and VCP (30%) enhanced the luciferase reporter activity for H3N2, suggesting an entry inhibitory role of factor H and VCP against H1N1, but not H3N2. Thus, factor H can modulate IAV infection and inflammatory response independent of its complement-related functions. King Saud University, Riyadh, International Scientific Partnership Programme (ISPP) ISPP-145.

The AU-rich element RNA-binding protein 1 (AUF1) is an RNA-binding protein, which can both stabilize and destabilize the transcripts of several cancer-related genes. Since epithelial-to-mesenchymal transition (EMT) and the acquisition of cancer stem cell traits are important for cancer onset and progression, we sought to determine the role of AUF1 in these two important processes. We have shown that AUF1 induces EMT and stemness in breast epithelial cells via stabilization of the SNAIL1 and TWIST1 mRNAs, and their consequent upregulation. Indeed, AUF1 binds the transcripts of these two genes at their 3′UTR and reduces their turnover. Ectopic expression of AUF1 also promoted stemness in mammary epithelial cells, and thereby increased the proportion of cancer stem cells. Importantly, breast cancer cells that ectopically express AUF1 were more efficient in forming orthotopic tumor xenografts in nude mice than their corresponding controls with limiting cell inocula. On the other hand, AUF1 downregulation with specific siRNA inhibited EMT and reduced the stemness features in breast cancer cells. Moreover, AUF1 knockdown sensitized breast cancer cells to the killing effect of cisplatin. Together, these findings provide clear evidence that AUF1 is an important inducer of the EMT process through stabilization of SNAIL1 and TWIST1 and the consequent promotion of breast cancer stem cells. Thereby, AUF1 targeted molecules could constitute efficient therapeutics for breast cancer patients.

The complexity in chemical composition alongside the genomic complexity of crop plants poses significant challenges for the characterization of their proteomes. This chapter provides specific methods that can be used for the extraction and identification of proteins from sweet potato, and a proteogenomic method for the subsequent peptide mapping on the haplotype-derived sweet potato genome assembly. We outline two basic methods for extracting proteins expressed in root and leaf tissues for the label-free quantitative proteomics-one phenol-based procedure and one polyethylene glycol (PEG) 4000-based fractionation method-and discuss strategies for the organ-specific protein extraction and increased recovery of low-abundance proteins. Next, we describe computational methods for improved proteome annotation of sweet potato based on aggregated genomics and transcriptomics resources available in our and public databases. Lastly, we describe an easily customizable proteogenomics approach for mapping sweet potato peptides back to their genome location and exemplify its use in improving genome annotations using a mass spectrometry data set.

Developmental and/or epileptic encephalopathies (DEEs) are a group of devastating genetic disorders, resulting in early-onset, therapy-resistant seizures and developmental delay. Here we report on 22 individuals from 15 families presenting with a severe form of intractable epilepsy, severe developmental delay, progressive microcephaly, visual disturbance and similar minor dysmorphisms. Whole exome sequencing identified a recurrent, homozygous variant (chr2:64083454A > G) in the essential UDP-glucose pyrophosphorylase (UGP2) gene in all probands. This rare variant results in a tolerable Met12Val missense change of the longer UGP2 protein isoform but causes a disruption of the start codon of the shorter isoform, which is predominant in brain. We show that the absence of the shorter isoform leads to a reduction of functional UGP2 enzyme in neural stem cells, leading to altered glycogen metabolism, upregulated unfolded protein response and premature neuronal differentiation, as modeled during pluripotent stem cell differentiation in vitro. In contrast, the complete lack of all UGP2 isoforms leads to differentiation defects in multiple lineages in human cells. Reduced expression of Ugp2a/Ugp2b in vivo in zebrafish mimics visual disturbance and mutant animals show a behavioral phenotype. Our study identifies a recurrent start codon mutation in UGP2 as a cause of a novel autosomal recessive DEE syndrome. Importantly, it also shows that isoform-specific start-loss mutations causing expression loss of a tissue-relevant isoform of an essential protein can cause a genetic disease, even when an organism-wide protein absence is incompatible with life. We provide additional examples where a similar disease mechanism applies. Electronic supplementary material The online version of this article (10.1007/s00401-019-02109-6) contains supplementary material, which is available to authorized users.

The expression of PD‐L1 in breast cancer is associated with estrogen receptor negativity, chemoresistance and epithelial‐to‐mesenchymal transition (EMT), all of which are common features of a highly tumorigenic subpopulation of cancer cells termed cancer stem cells (CSCs). Hitherto, the expression and intrinsic role of PD‐L1 in the dynamics of breast CSCs has not been investigated. To address this issue, we used transcriptomic datasets, proteomics and several in vitro and in vivo assays. Expression profiling of a large breast cancer dataset (530 patients) showed statistically significant correlation (p, What's new? Cancer cells that express the T‐cell inhibitory molecule programmed death‐ligand 1 (PD‐L1) readily escape immune attack. In addition, PD‐L1 expression contributes to chemoresistance and is associated with epithelial‐to‐mesenchymal transition, a process that generates cancer stem cells (CSCs). This study shows that in breast cancer, PD‐L1 expression further plays a direct part in maintaining CSC stemness. In breast cancer cells, PD‐L1 expression sustained stemness factors OCT‐4A and Nanog, via a PI3K/AKT‐dependent pathway, and promoted expression of the stemness controlling factor BMI1, independent of PI3K/AKT. Targeting PD‐L1 could help advance breast cancer therapy, owing to impacts on the pool of breast CSCs.

CYP24A1, the primary inactivating enzyme for vitamin D, is often overexpressed in human cancers, potentially neutralizing the antitumor effects of calcitriol, the active form of vitamin D. However, it is unclear whether CYP24A1 expression serves as a functional contributor versus only a biomarker for tumor progression. In this study, we investigated the role of CYP24A1 on malignant progression of a murine model of BrafV600E-induced papillary thyroid cancer (PTC). Mice harboring wild-type Cyp24a1 (BVECyp24a1-wt) developed PTC at 5 weeks of age. Mice harboring a homozygous deletion of Cyp24a1 (BVECyp24a1-null) exhibited a 4-fold reduction in tumor growth. Notably, we found the tumorigenic potential of BVECyp24a1-null-derived tumor cells to be nearly abolished in immunocompromised nude mice. This phenotype was associated with downregulation of the MAPK, PI3K/Akt, and TGFβ signaling pathways and a loss of epithelial–mesenchymal transition (EMT) in BVECyp24a1-null cells, associated with downregulation of genes involved in EMT, tumor invasion, and metastasis. While calcitriol treatment did not decrease cell proliferation in BVECyp24a1-null cells, it strengthened antitumor responses to the BRAFV600E inhibitor PLX4720 in both BVECyp24a1-null and BVECyp24a1-wt cells. Our findings offer direct evidence that Cyp24a1 functions as an oncogene in PTC, where its overexpression activates multiple signaling cascades to promote malignant progression and resistance to PLX4720 treatment. Cancer Res; 77(8); 2161–72. ©2017 AACR.

Microvascular loss may be a root cause of chronic rejection in lung transplants, which leads to the bronchiolitis obliterans syndrome. Previous research implicates T regulatory cell (Treg) as a key component of immune modulation, however, Treg has never been examined as a reparative mediator to salvage microvasculature during transplantation. Here, we reconstituted purified Tregs in to allografts, and serially monitored allografts for tissue oxygenation, microvascular perfusion for four weeks. We demonstrated that Tregs reconstitution of allografts significantly improve tissue oxygenation, microvascular flow, epithelial repair, number of CD4 + CD25 high FOXP3 + Tregs, followed by an upregulation of proinflammatory, angiogenic and regulatory genes, while prevented subepithelial deposition of CD4 + T cells at d10, and collagen at d28 post-transplantation. Altogether, these findings concluded that Treg-mediated immunotherapy has potential to preserve microvasculature and rescue allograft from sustained hypoxic/ischemic phase, limits airway tissue remodeling, and therefore may be a useful therapeutic tool to prevent chronic rejection after organ transplantation.

The data set presented here is associated with the article “Intracellular calcium and NF-kB regulate hypoxia-induced leptin, VEGF, IL-6 and adiponectin secretion in human adipocytes” (Al-Anazi et al., 2018). Data illustrate hypoxia-induced VEGF and leptin expression in human adipocytes treated with the calcium chelator BAPTA-AM (1 µM). It also shows NF-κB p65 induced expression by hypoxia. Preadipocytes were differentiated for 14 days and then subjected to 0.5–1.5% oxygen in the presence and absence of BAPTA-AM or the NF-κB inhibitor SN50 for 48 h prior to RNA isolation and PCR analysis.

Familial hypercholesterolemia (FH) is an autosomal dominant disease most often caused by mutations in the low-density lipoprotein receptor (LDLR) gene, which consists of 18 exons spanning 45 kb and codes for a precursor protein of 860 amino acids. Mutations in the LDLR gene lead to a reduced hepatic clearance of LDL as well as a high risk of coronary artery disease (CAD) and sudden cardiac death (SCD). Recently, LDLR transgenes have generated interest as potential therapeutic agents. However, LDLR packaging using a lentiviral vector (LVV) system pseudotyped with a vesicular stomatitis virus (VSV)-G envelope is not efficient. In this study, we modified the LVV system to improve transduction efficiency and investigated the LDLR regions responsible for transduction inhibition. Transduction efficiency of 293T cells with a 5&prime, LDLReGFP-3&prime, fusion construct was only 1.55% compared to 42.32% for the eGFP construct. Moreover, co-expression of LDLR affected eGFP packaging. To determine the specific region of the LDLR protein responsible for packaging inhibition, we designed constructs with mutations or sequential deletions at the 3&prime, and 5&prime, ends of LDLR cDNA. All constructs except one without the ligand-binding domain (LBD) (pWoLBD&ndash, eGFP) resulted in low transduction efficiency, despite successful packaging of viral RNA in the VSV envelope, as confirmed through RT-PCR. When we evaluated a direct interaction between LDLR and the VSV envelope glycoprotein using MD simulation and protein&ndash, protein interactions, we uncovered Val119, Thr120, Thr67, and Thr118 as exposed residues in the LDLR receptor that interact with the VSV protein. Together, our results suggest that the LBD of LDLR interacts with the VSV-G protein during viral packaging, which significantly reduces transduction efficiency.

During aging, cellular plasticity and senescence play important roles in tissue regeneration and the pathogenesis of different diseases, including cancer. We have recently shown that senescent breast luminal cells can activate their adjacent stromal fibroblasts. In the present report, we present clear evidence that these senescence-related active fibroblasts can dedifferentiate proliferating primary human luminal cells to multipotent stem cells in an interleukin-8 (IL-8)-dependent manner. This was confirmed using recombinant IL-8, while the truncated protein was not active. This IL-8-related dedifferentiation of luminal cells was mediated through the STAT3-dependent downregulation of p16(INK4A) and the microRNA miR-141. Importantly, these in vitro-generated mammary stem cells exhibited high molecular and cellular similarities to human mammary stem cells. They have also shown a long-term mammary gland-reconstituting ability and the capacity to produce milk postdelivery. Thereby, these IL-8-generated mammary stem cells could be of great value for autologous cell therapy procedures and also for biomedical research as well as drug development.

Two complementary protein extraction methodologies coupled with an automated proteomic platform were employed to analyze tissue-specific proteomes and characterize biological and metabolic processes in sweetpotato. A total of 74 255 peptides corresponding to 4321 nonredundant proteins were successfully identified. Data were compared to predicted protein accessions for Ipomoea species and mapped on the sweetpotato transcriptome and haplotype-resolved genome. The two methodologies exhibited differences in the number and class of the unique proteins extracted. Overall, 39 916 peptides mapped to 3143 unique proteins in leaves, and 34 339 peptides mapped to 2928 unique proteins in roots. Primary metabolism and protein translation processes were enriched in leaves, whereas genetic pathways associated with protein folding, transport, sorting, as well as pathways in the primary carbohydrate metabolism were enriched in storage roots. A proteogenomics analysis successfully mapped 90.4% of the total uniquely identified peptides against the sweetpotato transcriptome and genome, predicted 741 new protein-coding genes, and specified 2056 loci where gene annotations can be further improved. The proteogenomics results provide evidence for the translation of new open reading frames (ORFs), alternative ORFs, exon extensions, and intronic ORF sequences. Data are available via ProteomeXchange with identifier PXD012999.

Increasing evidence supports the critical role of active stromal adipocytes in breast cancer development and spread. However, the mediators and the mechanisms of action are still elusive. We show here that cancer-associated adipocytes (CAAs) isolated from 10 invasive breast carcinomas are proinflammatory and exhibit active phenotypes, including higher proliferative, invasive, and migratory capacities compared to their adjacent tumor-counterpart adipocytes (TCAs). Furthermore, all CAAs secreted higher level of interleukin-8 (IL-8), which is critical in mediating the paracrine procarcinogenic effects of these cells. Importantly, ectopic expression of IL-8 in TCA cells activated them and enhanced their procarcinogenic effects both in vitro, in a STAT3-dependent manner, and in vivo. In contrast, inhibition of the IL-8 signaling using specific short hairpin RNA, anti-IL-8 antibody, or reparixin suppressed the active features of CAAs, including their non-cell-autonomous tumor-promoting activities both on breast luminal cells and in orthotopic tumor xenografts in mice. IL-8 played also an important role in enhancing the proangiogenic effects of breast adipocytes. These results provide clear indication that IL-8 plays key roles in the activation of breast CAAs and acts as a major mediator for their paracrine protumorigenic effects. Thus, targeting CAAs by inhibiting the IL-8 pathway could have great therapeutic value.

An emerging dogma shows that tumors are initiated and maintained by a subpopulation of cancer cells that hijack some stem cell features and thus referred to as “cancer stem cells” (CSCs). The exact mechanism that regulates the maintenance of CSC pool remains largely unknown. Fascin is an actin-bundling protein that we have previously demonstrated to be a major regulator of breast cancer chemoresistance and metastasis, two cardinal features of CSCs. Here, we manipulated fascin expression in breast cancer cell lines and used several in vitro and in vivo approaches to examine the relationship between fascin expression and breast CSCs. Fascin knockdown significantly reduced stem cell-like phenotype (CD44hi/CD24lo and ALDH+) and reversal of epithelial to mesenchymal transition. Interestingly, expression of the embryonic stem cell transcriptional factors (Oct4, Nanog, Sox2, and Klf4) was significantly reduced when fascin expression was down-regulated. Functionally, fascin-knockdown cells were less competent in forming colonies and tumorspheres, consistent with lower basal self-renewal activity and higher susceptibility to chemotherapy. Fascin effect on CSC chemoresistance and self-renewability was associated with Notch signaling. Activation of Notch induced the relevant downstream targets predominantly in the fascin-positive cells. Limiting-dilution xenotransplantation assay showed higher frequency of tumor-initiating cells in the fascin-positive group. Collectively, our data demonstrated fascin as a critical regulator of breast CSC pool at least partially via activation of the Notch self-renewal signaling pathway and modification of the expression embryonic transcriptional factors. Targeting fascin may halt CSCs and thus presents a novel therapeutic approach for effective treatment of breast cancer. Video Highlight: https://youtu.be/GxS4fJ_Ow-o

Rationale Aspartame (L-aspartyl phenylalanine methyl ester) is a non-nutritive sweetener (NNS) approved for use in more than 6000 dietary products and pharmaceuticals consumed by the general public including adults and children, pregnant and nursing mothers. However a recent prospective study reported a doubling of the risk of being overweight amongst 1-year old children whose mothers consumed NNS-sweetened beverages daily during pregnancy. We have previously shown that chronic aspartame (ASP) exposure commencing in utero may detrimentally affect adulthood adiposity status, glucose metabolism and aspects of behavior and spatial cognition, and that this can be modulated by developmental N-methyl-D-aspartate receptor (NMDAR) blockade with the competitive antagonist CGP 39551 (CGP). Since glucose homeostasis and certain aspects of behavior and locomotion are regulated in part by the NMDAR-rich hypothalamus, which is part of the hypothalamic-pituitary-adrenal- (HPA) axis, we have elected to examine changes in hypothalamic and adrenal gene expression in response to ASP exposure in the presence or absence of developmental NMDAR antagonism with CGP, using Affymetrix microarray analysis. Results Using 2-factor ANOVA we identified 189 ASP-responsive differentially expressed genes (DEGs) in the adult male hypothalamus and 2188 in the adrenals, and a further 23 hypothalamic and 232 adrenal genes significantly regulated by developmental treatment with CGP alone. ASP exposure robustly elevated the expression of a network of genes involved in hypothalamic neurosteroidogenesis, together with cell stress and inflammatory genes, consistent with previous reports of aspartame-induced CNS stress and oxidative damage. These genes were not differentially expressed in ASP mice with CGP antagonism. In the adrenal glands of ASP-exposed mice, GABA and Glutamate receptor subunit genes were amongst those most highly upregulated. Developmental NMDAR antagonism alone had less effect on adulthood gene expression and affected mainly hypothalamic neurogenesis and adrenal steroid metabolism. Combined ASP + CGP treatment mainly upregulated genes involved in adrenal drug and cholesterol metabolism. Conclusion ASP exposure increased the expression of functional networks of genes involved in hypothalamic neurosteroidogenesis and adrenal catecholamine synthesis, patterns of expression which were not present in ASP-exposed mice with developmental NMDAR antagonism.

International audience; In the domain of wireless digital communication, floating-point arithmetic is generally used to conduct performance evaluation studies of algorithms. This is typically limited to theoretical performance evaluation in terms of communication quality and error rates. For a practical implementation perspective, using fixed-point arithmetic instead of floating-point reduces significantly implementation costs in terms of area occupation and energy consumption. However, this implies a complex conversion process, particularly if the considered algorithm includes complex arithmetic operations with high accuracy requirements and if the target system presents many configuration parameters. In this context, the purpose of the paper is to present an efficient quantization and fixed-point representation for turbo-detection and turbo-demapping. The impact of floating-to-fixed-point conversion is illustrated upon the error-rate performance of the receiver for different system configurations. Only a slight degradation in the error-rate performance of the receiver is observed when implementing the detector and demapper modules which utilize the devised quantization and fixed-point arithmetic rather than floating-point arithmetic.

The BRAF(V600E) mutation is found in approximately 40% of papillary thyroid cancers (PTC). Mice with thyroid-specific expression of Braf(V600E) (TPO-Braf(V600E)) develop PTC rapidly with high levels of serum thyroid-stimulating hormone (TSH). It is unclear to what extent the elevated TSH contributes to tumor progression. To investigate the progression of Braf(V600E)-induced PTC (BVE-PTC) under normal TSH, we transplanted BVE-PTC tumors subcutaneously into nude and TPO-Braf(WT) mice. Regression of the transplanted tumors was observed in both nude and TPO-Braf(WT) mice. They were surrounded by heavy lymphocyte infiltration and oncogene-induced senescence (OIS) was demonstrated by strong β-gal staining and absence of Ki-67 expression. In contrast, BVE-PTC transplants continued to grow when transplanted into TPO-Braf(V600E) mice. The expression of Trp53 was increased in tumor transplants undergoing OIS. Trp53 inactivation reversed OIS and enabled tumor transplants to grow in nude mice with characteristic cell morphology of anaplastic thyroid cancer (ATC). PTC-to-ATC transformation was also observed in primary BVE-PTC tumors. ATC cells derived from Trp53 knockout tumors had increased PI3K/AKT signaling and became resistant to Braf(V600E) inhibitor PLX4720, which could be overcome by combined treatment of PI3K inhibitor LY294002 and PLX4720. In conclusion, BVE-PTC progression could be contained via p53-dependent OIS and TSH is a major disruptor of this balance. Simultaneous targeting of both MAPK and PI3K/AKT pathways offer a better therapeutic outcome against ATC. The current study reinforces the importance of rigorous control of serum TSH in PTC patients.

Background Lipid metabolism dysfunction leading to excess fat deposits (obesity) may cause tumor (cancer) development. Both obesity and cancer are the epicenter of important medical issues. Lipid metabolism and cell death/proliferation are controlled by biochemical and molecular pathways involving many proteins, and organelles; alteration in these pathways leads to fat accumulation or tumor growth. Mammalian Kruppel-like factors, KLFs play key roles in both lipid metabolism and tumor development. Scope of review Substantial epidemiological and clinical studies have established strong association of obesity with a number of human cancers. However, we need more experimental verification to determine the exact role of this metabolic alteration in the context of tumor development. A clear understanding of molecules, pathways and the mechanisms involved in lipid metabolism and cell death/proliferation will have important implications in pathogenesis, and prevention of these diseases. Major conclusion The regulatory role of KLFs, in both cell death/proliferation and lipid metabolism suggests a common regulation of both processes. This provides an excellent model for delivering a precise understanding of the mechanisms linking altered expression of KLFs to obesity and tumor development. General significance Currently, mouse and rats are the models of choice for investigating disease mechanisms and pharmacological therapies but a genetic model is needed for a thorough examination of KLF function in vivo during the development of an organism. The worm Caenorhabditis elegans is an ideal model to study the connectivity between lipid metabolism and cell death/proliferation.

Aims: Phlomis cancellata is an aromatic and native plant that is a member of Labiatae. Not only a traditional medicine but also the plant is considered in modern medicine and different industries because of it s essential oil characteristics. The features of essential oils in aromatic plants are based on the part of plant from which they are extracted. In the present study the chemical composition of the essential oils of different parts of the Phlomis cancellat a were evaluated. Place and Duration of Study: Plant was collected from its natural habitat in Mazandaran province, Iran, in July 2012, its parts were separated and then dried in laboratory (Nour Branch, Islamic Azad University, Iran) and essential oils were extracted by hydrodistillation. Methodology: The essential oil of P. cancellata obtained by hydrodistillation and analyzed by GC FID and GC/MS. Results: The chemical analysis has resulted in identification of 25, 20, 11 and 2 constituents

Background There are numerous nuclear genes that cause mitochondrial disorders and clinically and genetically heterogeneous disorders whose aetiology often remains unsolved. In this study, we aim to investigate an autosomal recessive syndrome causing leukodystrophy and neuroregression. We studied six patients from five unrelated consanguineous families. Methods Patients underwent full neurological, radiological, genetic, metabolic and dysmorphological examinations. Exome sequencing coupled with autozygosity mapping, Sanger sequencing, microsatellite haplotyping, standard and molecular karyotyping and whole mitochondrial DNA sequencing were used to identify the genetic cause of the syndrome. Immunohistochemistry, transmission electron microscopy, confocal microscopy, dipstick assays, quantitative PCR, reverse transcription PCR and quantitative reverse transcription PCR were performed on different tissue samples from the patients. Results We identified a homoallelic missense founder mutation in ISCA2 leading to mitochondrial depletion and reduced complex I activity as well as decreased ISCA2 , ISCA1 and IBA57 expression in fibroblasts. MRI indicated similar white matter abnormalities in the patients. Histological examination of the skeletal muscle showed mild to moderate variation in myofibre size and the presence of many randomly distributed atrophic fibres. Conclusions Our data demonstrate that ISCA2 deficiency leads to a hereditary mitochondrial neurodegenerative white matter disease in infancy.

The activation of breast stromal fibroblasts is a crucial step toward tumor growth and spread. Therefore, it is extremely important to understand the molecular basis of this activation and determine the molecules and the mechanisms responsible for its sustainability. In the present report we have shown that the DNA methyl-transferase protein DNMT1 is critical for the activation of breast stromal fibroblasts as well as the persistence of their active status. Indeed, we have first revealed DNMT1 up-regulation in most cancer-associated fibroblasts relative to their corresponding adjacent normal fibroblasts. This effect resulted from HuR-dependent stabilization of the DNMT1 mRNA. Furthermore, ectopic expression of DNMT1 activated primary normal breast fibroblasts and promoted their pro-carcinogenic effects, both in vitro and in orthotopic tumor xenografts. By contrast, specific DNMT1 knockdown normalized breast myofibroblasts and repressed their cancer-promoting properties. These effects were sustained through inhibition of the IL-6/STAT3/NF-κB epigenetic cancer/inflammation positive feedback loop. Furthermore, we have shown that DNMT1-related activation of breast fibroblasts is mediated through upregulation of the RNA binding protein AUF1, which is also part of the loop. The present data demonstrate the critical function of DNMT1 in breast cancer-related sustained activation of breast stromal fibroblasts.

Obesity is increasingly recognized as a risk factor for breast cancer development. However, the molecular basis of obesity-related breast carcinogenesis remains elusive. In this study, we have shown that obesity reduces the level of the tumor suppressor p16INK4A protein in breast adipocytes, which showed active features and strong procarcinogenic potential both in vitro and in orthotopic tumor xenografts compared to mature adipocytes from lean women. Furthermore, obesity triggered epithelial-to-mesenchymal transition (EMT) in breast ductal epithelial cells. Interestingly, specific downregulation of p16INK4A increased the expression/secretion levels of various adipokines, including leptin, and activated breast adipocytes from lean women. Consequently, like breast adipocytes from obese women, p16-deficient adipocytes induced EMT in normal primary breast luminal cells in a leptin-dependent manner and enhanced tumor growth. Additionally, we have shown that p16INK4A negatively controls leptin at the mRNA level through microRNAs 141 and 146b-5p (miR-141 and miR-146b-5p), which bind the leptin mRNA at a specific sequence in the 3' untranslated region (UTR). These results show that obesity activates breast stromal adipocytes through p16 downregulation, which upregulates leptin and promotes procarcinogenic processes.

Mitochondrial calcium homeostasis is a tightly controlled process that is required for a variety of cellular functions. The mitochondrial calcium uniporter complex plays a critical role in this process. MICU2 is a major component of the mitochondrial calcium uniporter complex and its deficiency has been shown to impair mitochondrial calcium [Ca2+]m homeostasis although the exact mechanism remains unclear. We used exome sequencing, positional mapping, and functional characterization of MICU2 deficiency to investigate the role of MICU2 in calcium homeostasis. Using combined autozygome/exome analysis, a homozygous truncating mutation in MICU2 was found to fully segregate with a neurodevelopmental disorder in the form of severe cognitive impairment, spasticity, and white matter involvement in a multiplex consanguineous family. Patient-derived MICU2-deficient cells displayed impaired [Ca2+]m homeostasis, with associated increase in mitochondrial sensitivity to oxidative stress, and abnormal regulation of inner mitochondrial membrane potential. This is the first demonstration of MICU2 deficiency in humans, which we suggest causes a distinct neurodevelopmental phenotype secondary to impaired mitochondrial calcium uniporter-mediated regulation of intracellular calcium homeostasis.

Cardiovascular diseases are the major cause of morbidity and mortality worldwide. Our understanding of the molecular mechanisms governing these diseases has expanded exponentially over the past few decades, and a more basic understanding of disease development is necessary in order to identify molecular targets for “new” pharmaceutical therapeutics. For the heart to perform its assigned functions, it secretes many molecular moieties that synchronously act in an autocrine/paracrine and endocrine fashion, i.e., the cardiac secretome. The heart is made up of different cell types embedded in a dynamic extracellular matrix that is constantly remodeled to meet functional demands. The cardiokines are an expanding family of proteins secreted by the various cardiac cells through the classical and nonclassical secretory pathways with profound local and global effects. The ensuing chapter discusses this family and its relationship to heart functions especially those associated with disease development.

Coptotermes gestroi (Wasmann) or the Asian subterranean termite is a serious structural pest in urban settlements in Southeast Asia that has been introduced to other parts of the world through human commerce. Although mitochondrial DNA markers were previously used to shed light on the dispersal history of the Asian subterranean termite, there were limited attempts to analyze or include populations of the termite found in the wild in Southeast Asia. In this study, we analyzed the 16S ribosomal RNA (16S rRNA) and cytochrome c oxidase subunit 1 (cox1) genes of Asian subterranean termite colonies found in mangrove swamps, beach forests, plantations, and buildings in semi-urban and urban areas to determine the relationship between colonies found in the wild and the urban habitat, and to investigate the possibility of different ecotypes of the termite in Peninsular Malaysia. Our findings show that the 16S rRNA haplotypes recovered from this study clustered into eastern, western, and southern populations of the termite, while the cox1 haplotypes were often specific to an area or site. The 16S rRNA and cox1 genes or haplotypes showed that the most abundant haplotype occupied a wide range of environments or habitats. In addition, the cox1 tree showed evidence of historical biogeography where basal haplotypes inhabited a wide range of habitats, while apical haplotypes were restricted to mangrove swamps and beach forests. Information on the haplotype-habitat association of C. gestroi will enable the prediction of habitats that may harbor or be at risk of invasion in areas where they have been introduced.

Objective— Severe heatstroke is a leading cause of morbidity and mortality during heat waves. The pathogenesis of tissue injury, organ failure, and death in heatstroke is not well understood. Methods and Result— We investigated the pathways of heatstroke-induced tissue injury and cell death in anesthetized baboons ( Papio hamadyras ) subjected to environmental heat stress until core temperature attained 42.5°C (moderate heatstroke; n=3) or onset of severe heatstroke (n=4) signaled by a fall in systolic blood pressure to Conclusion— These data suggested that microvascular injury, thrombosis, inflammation, and apoptosis may play an important role in the pathogenesis of heatstroke injury.