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Aims/hypothesis: Homozygous mutations in RFX6 lead to neonatal diabetes accompanied by a hypoplastic pancreas, whereas heterozygous mutations cause MODY. Recent studies have also shown RFX6 variants to be linked with type 2 diabetes. Despite RFX6's known function in islet development, its specific role in diabetes pathogenesis remains unclear. Here, we aimed to understand the mechanisms underlying the impairment of pancreatic islet development and subsequent hypoplasia due to loss-of-function mutations in RFX6., Methods: We examined regulatory factor X6 (RFX6) expression during human embryonic stem cell (hESC) differentiation into pancreatic islets and re-analysed a single-cell RNA-seq dataset to identify RFX6-specific cell populations during islet development. Furthermore, induced pluripotent stem cell (iPSC) lines lacking RFX6 were generated using CRISPR/Cas9. Various approaches were then employed to explore the consequences of RFX6 loss across different developmental stages. Subsequently, we evaluated transcriptional changes resulting from RFX6 loss through RNA-seq of pancreatic progenitors (PPs) and endocrine progenitors (EPs)., Results: RFX6 expression was detected in PDX1 + cells in the hESC-derived posterior foregut (PF). However, in the PPs, RFX6 did not co-localise with pancreatic and duodenal homeobox 1 (PDX1) or NK homeobox 1 (NKX6.1) but instead co-localised with neurogenin 3, NK2 homeobox 2 and islet hormones in the EPs and islets. Single-cell analysis revealed high RFX6 expression levels in endocrine clusters across various hESC-derived pancreatic differentiation stages. Upon differentiating iPSCs lacking RFX6 into pancreatic islets, a significant decrease in PDX1 expression at the PF stage was observed, although this did not affect PPs co-expressing PDX1 and NKX6.1. RNA-seq analysis showed the downregulation of essential genes involved in pancreatic endocrine differentiation, insulin secretion and ion transport due to RFX6 deficiency. Furthermore, RFX6 deficiency resulted in the formation of smaller islet organoids due to increased cellular apoptosis, linked to reduced catalase expression, implying a protective role for RFX6. Overexpression of RFX6 reversed defective phenotypes in RFX6-knockout PPs, EPs and islets., Conclusions/interpretation: These findings suggest that pancreatic hypoplasia and reduced islet cell formation associated with RFX6 mutations are not due to alterations in PDX1 + /NKX6.1 + PPs but instead result from cellular apoptosis and downregulation of pancreatic endocrine genes., Data Availability: RNA-seq datasets have been deposited in the Zenodo repository with accession link (DOI: https://doi.org/10.5281/zenodo.10656891 )., Competing Interests: Acknowledgements: We would like to thank N. R. Dunn (A*STAR, Singapore) for providing the HA-RFX6 tagged H9 hESC lines (RFX6HA/HA H9-hESCs). Furthermore, we thank the Genomic Core members at QBRI for their assistance with technical support in RNA-seq. Data availability: RNA-seq datasets have been deposited in the Zenodo repository with accession link (DOI: https://doi.org/10.5281/zenodo.10656891 ). Funding: Open Access funding provided by the Qatar National Library. This work was funded by grants from Qatar Biomedical Research Institute (QBRI) (Grant no. QBRI-HSCI Project 1). NA is a PhD student with a scholarship funded by QRDI (GSRA9-L-1-0511-22008). Authors’ relationships and activities: SH is a co-founder and shareholder of Sequantrix GmbH and has research funding from Novo Nordisk and Askbio. The authors declare that there are no other relationships or activities that might bias, or be perceived to bias, their work. Contribution statement: NA performed most of the experiments and analysed the data. AKE and BM performed experiments and analysed the data. SI and SH analysed the sequencing data. EMA conceived and designed the study, supervised the project, analysed and interpreted the data, and wrote the manuscript. All authors critically reviewed the article and approved the final version of the manuscript. EMA is the guarantor of this work., (© 2024. The Author(s).)

Objectives: Chicken (Gallus gallus), as the most economically important poultry, is a classical and ideal model for studying the mechanism of vertebrate developmental biology and embryology. However, the sex determination and differentiation in chicken is still elusive, which limited the application and slowed down many basic studies in chicken., Data Description: We applied PacBio Iso-seq to multiple spatiotemporal embryo-gonad tissues in the male and female chicken, which contain the blastoderm (E0, un-differentiation stage), genital ridge (E3.5-6.5, sex-differentiation stage) and gonads (E18.5, full-sex-differentiation stage). We obtained 51,479 and 48,356 full-length transcripts in male and female chicken embryo, respectively. The comprehensive annotated and evaluated these transcripts. The 1,293 and 1,556 candidate lncRNAs, 5,766 and 4,211 AS events in male and female. Collectively, our data constitutes a grand increase in the known number of lncRNA, AS (Alternative splicing) and Poly(A) during chicken embryo sex-differentiation and plays an important role in improving current genome annotation. In the meantime, the data will be enriched the functional studies in other birds., (© 2024. The Author(s).)

The pathogenesis of diabetes involves complex changes in the expression profiles of mRNA and non-coding RNAs within pancreatic islet cells. Recent progress in induced pluripotent stem cell (iPSC) technology have allowed the modeling of diabetes-associated genes. Our recent study using FOXA2-deficient human iPSC models has highlighted an essential role for FOXA2 in the development of human pancreas. Here, we aimed to provide further insights on the role of microRNAs (miRNAs) by studying the miRNA-mRNA regulatory networks in iPSC-derived islets lacking the FOXA2 gene. Consistent with our previous findings, the absence of FOXA2 significantly downregulated the expression of islet hormones, INS, and GCG, alongside other key developmental genes in pancreatic islets. Concordantly, RNA-Seq analysis showed significant downregulation of genes related to pancreatic development and upregulation of genes associated with nervous system development and lipid metabolic pathways. Furthermore, the absence of FOXA2 in iPSC-derived pancreatic islets resulted in significant alterations in miRNA expression, with 61 miRNAs upregulated and 99 downregulated. The upregulated miRNAs targeted crucial genes involved in diabetes and pancreatic islet cell development. In contrary, the absence of FOXA2 in islets showed a network of downregulated miRNAs targeting genes related to nervous system development and lipid metabolism. These findings highlight the impact of FOXA2 absence on pancreatic islet development and suggesting intricate miRNA-mRNA regulatory networks affecting pancreatic islet cell development., (© 2024. The Author(s).)

Autism spectrum disorder (ASD) is a complex developmental disorder characterized by challenges with social interactions and restricted/repetitive behaviors. Here, we recruited nine Qatari children of Arab ethnicity (males, aged 2-4 years), including six ASD subjects (n = 3 mild-to-moderate ASD and n = 3 severe ASD) and three control subjects. We generated induced pluripotent stem cell (iPSC) lines from PBMC samples of these subjects using non-integrating Sendai viral vectors. These iPSC lines were fully characterized and exhibited pluripotency characteristics, normal karyotypes, and trilineage differentiation potential. These iPSC lines provide valuable cell models for understanding ASD pathophysiology and developing new therapeutics for ASD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Our recent studies have demonstrated the crucial involvement of FOXA2 in the development of human pancreas. Reduction of FOXA2 expression during the differentiation of induced pluripotent stem cells (iPSCs) into pancreatic islets has been found to reduce α-and β-cell masses. However, the extent to which such changes are linked to alterations in the expression profile of long non-coding RNAs (lncRNAs) remains unraveled., Methods: Here, we employed our recently established FOXA2-deficient iPSCs (FOXA2 -/- iPSCs) to investigate changes in lncRNA profiles and their correlation with dysregulated mRNAs during the pancreatic progenitor (PP) and pancreatic islet stages. Furthermore, we constructed co-expression networks linking significantly downregulated lncRNAs with differentially expressed pancreatic mRNAs., Results: Our results showed that 442 lncRNAs were downregulated, and 114 lncRNAs were upregulated in PPs lacking FOXA2 compared to controls. Similarly, 177 lncRNAs were downregulated, and 59 lncRNAs were upregulated in islet cells lacking FOXA2 compared to controls. At both stages, we observed a strong correlation between lncRNAs and several crucial pancreatic genes and TFs during pancreatic differentiation. Correlation analysis revealed 12 DE-lncRNAs that strongly correlated with key downregulated pancreatic genes in both PPs and islet cell stages. Selected DE-lncRNAs were validated using RT-qPCR., Conclusions: Our data indicate that the observed defects in pancreatic islet development due to the FOXA2 loss is associated with significant alterations in the expression profile of lncRNAs. Therefore, our findings provide novel insights into the role of lncRNA and mRNA networks in regulating pancreatic islet development, which warrants further investigations. Video Abstract., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Recently, we reported that forkhead box A2 (FOXA2) is required for the development of human pancreatic α- and β-cells. However, whether miRNAs play a role in regulating pancreatic genes during pancreatic development in the absence of FOXA2 expression is largely unknown. Here, we aimed to capture the dysregulated miRNAs and to identify their pancreatic-specific gene targets in pancreatic progenitors (PPs) derived from wild-type induced pluripotent stem cells (WT-iPSCs) and from iPSCs lacking FOXA2 (FOXA2 -/- iPSCs). To identify differentially expressed miRNAs (DEmiRs), and genes (DEGs), two different FOXA2 -/- iPSC lines were differentiated into PPs. FOXA2 -/- PPs showed a significant reduction in the expression of the main PP transcription factors (TFs) in comparison to WT-PPs. RNA sequencing analysis demonstrated significant reduction in the mRNA expression of genes involved in the development and function of exocrine and endocrine pancreas. Furthermore, miRNA profiling identified 107 downregulated and 111 upregulated DEmiRs in FOXA2 -/- PPs compared to WT-PPs. Target prediction analysis between DEmiRs and DEGs identified 92 upregulated miRNAs, predicted to target 1498 downregulated genes in FOXA2 -/- PPs. Several important pancreatic TFs essential for pancreatic development were targeted by multiple DEmiRs. Selected DEmiRs and DEGs were further validated using RT-qPCR. Our findings revealed that FOXA2 expression is crucial for pancreatic development through regulating the expression of pancreatic endocrine and exocrine genes targeted by a set of miRNAs at the pancreatic progenitor stage. These data provide novel insights of the effect of FOXA2 deficiency on miRNA-mRNA regulatory networks controlling pancreatic development and differentiation., (© 2023. The Author(s).)

Background: The genetic factors associated with insulin resistance (IR) are not well understood. Clinical studies on first-degree relatives of type 2 diabetic (T2D) patients, which have the highest genetic predisposition to T2D, have given insights into the role of IR in T2D pathogenesis. Induced pluripotent stem cells (iPSCs) are excellent tools for disease modeling as they can retain the genetic imprint of the disease. Therefore, in this study, we aimed to investigate the genetic perturbations associated with insulin resistance (IR) in the offspring of T2D parents using patient-specific iPSCs., Methods: We generated iPSCs from IR individuals (IR-iPSCs) that were offspring of T2D parents as well as from insulin-sensitive (IS-iPSCs) individuals. We then performed transcriptomics to identify key dysregulated gene networks in the IR-iPSCs in comparison to IS-iPSCs and functionally validated them., Results: Transcriptomics on IR-iPSCs revealed dysregulated gene networks and biological processes indicating that they carry the genetic defects associated with IR that may lead to T2D. The IR-iPSCs had increased lactate secretion and a higher phosphorylation of AKT upon stimulation with insulin. IR-iPSCs have increased cellular oxidative stress indicated by a high production of reactive oxygen species and higher susceptibility to H 2 O 2 -induced apoptosis., Conclusions: IR-iPSCs generated from offspring of diabetic patients confirm that oxidative stress and increased lactate secretion, associated with IR, are inherited in this population, and may place them at a high risk of T2D. Overall, our IR-iPSC model can be employed for T2D modeling and drug screening studies that target genetic perturbations associated with IR in individuals with a high risk for T2D., (© 2022. The Author(s).)

In contrast to a great diversity in insects in terrestrial and freshwater ecosystems, few known species have adapted to inhabit marine environments. In this study, we surveyed insects associated with halophytic plants of Suaeda (Amaranthaceae) distributed in intertidal zones, in northern Kyushu, Japan. On four Japanese native species of Suaeda , we found insects belonging to five orders and 18 species. Amongst them, the genus Clanoneurum (Diptera: Ephydridae) and Coleophoradeviella (Lepidoptera: Coleophoridae) were newly reported from Japan; and Orthotylus (Melanotrichus) parvulus (Hemiptera: Miridae) was newly recorded from Kyushu. The seasonal occurrence of several insects on Suaeda is reported., (Akihito Kita, Ayman Khamis Elsayed, Makoto Tokuda.)

Some herbivorous insects possess the ability to synthesize phytohormones and are considered to use them for manipulating their host plants, but how these insects acquired the ability remains unclear. We investigated endogenous levels of auxin (IAA) and cytokinins (iP and tZ), including their ribosides (iPR and tZR), in various terrestrial arthropod taxa. Surprisingly, IAA was detected in all arthropods analysed. In contrast, tZ and/or tZR was detected only in some taxa. Endogenous levels of IAA were not significantly different among groups with different feeding habits, but gall inducers possessed significantly higher levels of iPR, tZ and tZR. Ancestral state reconstruction of the ability to synthesize tZ and tZR revealed that the trait has only been acquired in taxa containing gall inducers. Our results strongly suggest critical role of the cytokinin synthetic ability in the evolution of gall-inducing habit and IAA has some function in arthropods., (© 2022. The Author(s).)

Autism spectrum disorder (ASD) is a childhood-onset neurodevelopmental disorder characterized by social interaction, behavior, and communication challenges. Here, we generated an induced pluripotent stem cell (iPSC) line, QBRIi013-A using a non-integrating Sendai virus from a 6-year-old female diagnosed with ASD and intellectual disability. The QBRIi013-A cell line was fully characterized and exhibited a pluripotency capacity and trilineage differentiation potential. Furthermore, it showed normal karyotype and genetic identity to the patient's PBMCs. Consequently, this iPSC line provides a valuable cell model in understanding the molecular mechanism underlying the complexities of ASD pathogenesis., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

p-Nitrophenol (PNP) is the main end product of organophosphorus insecticides and a derivative of diesel exhaust particles. In addition to its unfavorable impact on reproductive functions in both genders, it also has various harmful physiological effects including lung cancer and allergic rhinitis. The identification of the cellular readout that functions in metabolic pathway perpetuation is still far from clear. This research aimed to study the impact of chronic PNP exposure on the health condition of the liver in Japanese quails. Quails were exposed to different concentrations of PNP as follows: 0.0 (control), 0.01mg (PNP/0.01), 0.1mg (PNP/0.1), and 1mg (PNP/1) per kg of body weight for 2.5 months through oral administration. Liver and plasma samples were collected at 1.5, 2, and 2.5 months post-treatment for biochemical, histopathology, and immunohistochemistry assessment. The plasma aspartate aminotransferase (AST) level was assessed enzymatically. The livers were collected for histopathology, glycogen accumulation, proliferating cell nuclear antigen (PCNA), and apoptosis assessment. Our results revealed an irregularity in body weight due to the long-term exposure of PNP with a significant reduction in liver weight. PNP treatment caused histopathological alterations in the hepatic tissues which increased in severity by the long-term exposure. The low dose led to mild degeneration with lymphocytic infiltration, while the moderate dose has a congestion effect with some necrosis; meanwhile severe hepatocyte degeneration and RBCs hemolysis were noticed due to high dose of PNP. Glycogen accumulation increased in hepatocytes by prolonged exposure to p-Nitrophenol with the highest intensity in the group treated by the high dose. Moderate and high doses of PNP resulted in a significant increase in apoptosis and hepatocytes' proliferation at the different time points after treatment. This increase is markedly notable and maximized at 2.5 months post-treatment. The damage occurred in a time-dependent manner. These changes reflected on the plasma hepatic enzyme AST that was clearly increased at 2.5 months of exposure. Therefore, it could be concluded that PNP has profound toxic effects on the liver in cellular level. Taking into consideration the time and dose factors, both have a synergistic effect on the accumulation of glycogen, apoptosis, and cellular proliferation, highlighting the power of cellular investigation which will potentially open the door for earlier medical intervention to counteract this toxicity. Collectively, PNP could have critical hurtful effects on the health of human beings, wild animals as well as livestock., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Recessive mutations in the glucose transporter gene SLC2A2 (GLUT2) lead to permanent neonatal diabetes (PNDM) and Fanconi Bickel Syndrome (FBS). Here, we generated an induced pluripotent stem cell (iPSC) line, QBRIi012-A, from a 24-month-old boy with FBS and PNDM due to homozygous nonsense mutation in the SLC2A2 gene (c.901C > T). The QBRIi012-A was fully characterized using different approaches. The cell line showed normal karyotype and was able to differentiate into the three germ layers in vitro. This iPSC line provides a novel human cell model to understand the pathophysiology of FBS and diabetes associated with SLC2A2 defects., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

Autism spectrum disorder (ASD) is a multifaced neurodevelopmental disorder that becomes apparent during early childhood development. The complexity of ASD makes clinically diagnosing the condition difficult. Consequently, by identifying the biomarkers associated with ASD severity and combining them with clinical diagnosis, one may better factionalize within the spectrum and devise more targeted therapeutic strategies. Currently, there are no reliable biomarkers that can be used for precise ASD diagnosis. Consequently, our pilot experimental cohort was subdivided into three groups: healthy controls, individuals those that express severe symptoms of ASD, and individuals that exhibit mild symptoms of ASD. Using next-generation sequencing, we were able to identify several circulating non-coding RNAs (cir-ncRNAs) in plasma. To the best of our knowledge, this study is the first to show that miRNAs, piRNAs, snoRNAs, Y-RNAs, tRNAs, and lncRNAs are stably expressed in plasma. Our data identify cir-ncRNAs that are specific to ASD. Furthermore, several of the identified cir-ncRNAs were explicitly associated with either the severe or mild groups. Hence, our findings suggest that cir-ncRNAs have the potential to be utilized as objective diagnostic biomarkers and clinical targets.

Background: A gall midge species (Diptera: Cecidomyiidae) inducing leaf bud galls on Magnolia kobus DC. var. borealis Sarg. (Magnoliaceae) was found in Hokkaido and northern Honshu, Japan., New Information: Based on its morphology, the species is regarded as an undescribed species of the genus Pseudasphondylia Monzen (Cecidomyiinae, Cecidomyiidi, Asphondyliini). The species is herein described as Pseudasphondylia saohimea Matsuda, Elsayed and Tokuda sp. n. The new species is easily distinguishable from its congeners by the number of adult palpal segments and the shape of the male terminalia and larval spatula., (Hiroki Matsuda, Ayman Khamis Elsayed, Wanggyu Kim, Satoshi Yamauchi, Martin Libra, Naoto Kamata, Junichi Yukawa, Makoto Tokuda.)

MicroRNAs (miRNAs) are essential factors in the reproductive process of poultry. Here, we found miR-302d is a potential differentiation and negative factor of chicken embryonic stem cells (ESCs) into spermatogonia stem cells (SSCs). The competition mechanism was carried out for the preliminary exploration to determine the relationship among miR-302d, lncRNA-341(interacting with miR-302d), and target gene TLE4. The results showed that lncRNA-341 can competitively bind to miR-302d to decrease the targeted binding of miR-302d and TLE4 which promotes the differentiation of chicken SSCs. Moreover, it is suggested that miR-302d may participate in the Wnt signaling pathway through TLE4., Competing Interests: The authors declare no conflict of interest., (Copyright © 2021 Yani Zhang et al.)

Insulin resistance (IR) is associated with several metabolic disorders, including type 2 diabetes (T2D). The development of IR in insulin target tissues involves genetic and acquired factors. Persons at genetic risk for T2D tend to develop IR several years before glucose intolerance. Several rodent models for both IR and T2D are being used to study the disease pathogenesis; however, these models cannot recapitulate all the aspects of this complex disorder as seen in each individual. Human pluripotent stem cells (hPSCs) can overcome the hurdles faced with the classical mouse models for studying IR. Human induced pluripotent stem cells (hiPSCs) can be generated from the somatic cells of the patients without the need to destroy a human embryo. Therefore, patient-specific hiPSCs can generate cells genetically identical to IR individuals, which can help in distinguishing between genetic and acquired defects in insulin sensitivity. Combining the technologies of genome editing and hiPSCs may provide important information about the genetic factors underlying the development of different forms of IR. Further studies are required to fill the gaps in understanding the pathogenesis of IR and diabetes. In this review, we summarize the factors involved in the development of IR in the insulin-target tissues leading to diabetes. Also, we highlight the use of hPSCs to understand the mechanisms underlying the development of IR., Competing Interests: Conflict-of-interest statement: The authors declare that they have no conflict of interest, (©The Author(s) 2021. Published by Baishideng Publishing Group Inc. All rights reserved.)

Ametrodiplosis Rübsaamen (Diptera: Cecidomyiidae: Clinodiplosini) is a mostly Holarctic gall midge genus whose species are associated with a wide range of seed plant families, either as gall-inducers or inquilines. In this study, we describe three species of Ametrodiplosis from Japan: A. adetos n. sp. feeding in the flowers of Tylophora aristolochioides Miq. (Apocynaceae); A. aeroradicis n. sp. inducing aerial root galls on Trachelospermum asiaticum (Sieb. et Zucc.) Nakai and T. gracilipes var. liukiuense (Apocynaceae); and A. stellariae n. sp. forming leaf bud galls on Stellaria uliginosa Murray var. undulata (Thunb.) Ohwi (Caryophyllaceae). A molecular phylogenetic analysis using mitochondrial COI and ribosomal 16S genes and nuclear ribosomal 28S gene were conducted for the three new Ametrodiplisis species and other clinodiplosine taxa sequences available in GenBank. The analysis supported the monophyly of Ametrodiplosis despite the variable life history of the three species. In addition, it indicated very low intraspecific genetic divergence among the individuals from different localities and/or host plants. A taxonomic key to the three new Japanese species of Ametrodiplosis is provided.

FOXA2 has been identified as an essential factor for pancreas development and emerging evidence supports an association between FOXA2 and diabetes. Although the role of FOXA2 during pancreatic development is well-studied in animal models, its role during human islet cell development remains unclear. Here, we generated induced pluripotent stem cells (iPSCs) from a patient with FOXA2 haploinsufficiency (FOXA2 +/ - iPSCs) followed by beta-cell differentiation to understand the role of FOXA2 during pancreatic beta-cell development. Our results showed that FOXA2 haploinsufficiency resulted in aberrant expression of genes essential for the differentiation and proper functioning of beta cells. At pancreatic progenitor (PP2) and endocrine progenitor (EPs) stages, transcriptome analysis showed downregulation in genes associated with pancreatic development and diabetes and upregulation in genes associated with nervous system development and WNT signaling pathway. Knockout of FOXA2 in control iPSCs (FOXA2 -/- iPSCs) led to severe phenotypes in EPs and beta-cell stages. The expression of NGN3 and its downstream targets at EPs as well as INSUILIN and GLUCAGON at the beta-cell stage, were almost absent in the cells derived from FOXA2 -/- iPSCs. These findings indicate that FOXA2 is crucial for human pancreatic endocrine development and its defect may lead to diabetes based on FOXA2 dosage.

MicroRNAs play a crucial role in sperm formation, but its specific function remains unknown. Here, we found that gga-miR-218 regulates chicken sperm formation through in/ex vivo experiments. We constructed over-expression/interference carrier to overexpress and inhibit gga-miR-218 in chicken spermatogonial stem cells, separately, the detection of haploid and QRT-PCR of meiosis related genes revealed that gga-miR-218 inhibits meiosis. After injection of miR-218 in vivo, semen concentration and HE (Hematoxylin and Eosin staining) revealed that gga-miR-218 inhibits meiosis. Meanwhile, we discovered that gga-miR-218 could target Stra8 by prediction software which can inhibit the wild-type fluorescence activity by co-transfection of gga-miR-218 with the Stra8 3' untranslated regions fluorescent reporter vector (wild-type/mutant), QRT-PCR and Western blot showed that gga-miR-218 inhibits the expression level of Stra8 by targeting its 3' untranslated regions directly. Finally, we suggest that gga-miR-218 could target to srta8 directly and inhibit spermatogenesis., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Heterozygous and homozygous mutations in the glucokinase (GCK) gene leads to maturity-onset diabetes of the young type 2 (MODY2) and permanent neonatal diabetes (PNDM), respectively. Here, we report the generation of two induced pluripotent stem cell (iPSC) lines, QBRIi010-A and QBRIi011-A, from patients with MODY2 and PNDM due to mutations in the GCK gene (c.437 T > C). The generated iPSC lines displayed pluripotency characteristics, were able to differentiate into the three germ layers, and showed normal karyotypes. These iPSC lines will serve as valuable human cell models for understanding diabetes pathogenesis and developing new therpaies for diabetes., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Betula (Betulaceae), or birch, is a Holarctic genus of trees and shrubs whose species have ornamental, industrial, and medical importance. Gall midges of the genus Massalongia (Diptera: Cecidomyiidae: Cecidomyiidi) are exclusively associated with birches in the Palearctic region. In 2018, an undescribed Massalongia species was discovered forming leaf galls on the midveins of B. grossa on Mount Tara, Saga Prefecture, Kyushu, Japan. In this study the species is described as M. nakamuratetsui Elsayed & Tokuda, sp. nov. , and a DNA barcode provided for it. The other known species of Massalongia are redescribed because the original descriptions are outdated and insufficient. A lectotype is designated for M. bachmaieri . In addition, the monotypic genus Apagodiplosis , containing A. papyriferae associated with B. papyrifera in the Nearctic region, is synonymized here under Massalongia , resulting in M. papyriferae comb. nov. , rendering Massalongia a Holarctic genus with six species. Comparing the sequence data of M. nakamuratetsui with all sequences available in The Barcode of Life Data (BOLD) system supports the occurrence of Massalongia in the Nearctic region and suggest that more species could be discovered there. Massalongia species form leaf or bud galls, and their mature larvae drop to the ground in autumn and overwinter in characteristic waterproof bottle-like cocoons, which is possibly a protective adaptation for pupation in wet and snowy lands. A taxonomic key to all Massalongia species is provided., (Ayman Khamis Elsayed, Marcela Skuhravá, Kazuki Ohta, Satoshi Yoshida, Makoto Tokuda.)

Fanconi Bickel Syndrome (FBS) is an autosomal recessive disease resulting from mutations in the SLC2A2 gene, encoding the GLUT2. FBS patients develop diabetes mellitus. Using non-integrating Sendai virus, we generated an induced pluripotent stem cell (iPSC) line, QBRIi007-A, carrying the c.613-7 T>G homozygous mutation in intron 5 of the SLC2A2 gene from a 19-year-old female with FBS and diabetes. The iPSC line was characterized for pluripotency, differentiation potential, genomic integrity, and genetic identity. This iPSC line provides a useful cell model to understand the role of GLUT2 in the disease development and to discover new drug candidates., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Psoriasis is characterized by hyperproliferation and defective differentiation of keratinocytes (KCs). Patients with psoriasis are at a high risk of developing diabetes and cardiovascular diseases. The debate on the genetic origin of psoriasis pathogenesis remains unresolved due to lack of suitable in vitro human models mimicking the disease phenotypes. In this study, we provide the first human induced pluripotent stem cell (iPSC) model for psoriasis carrying the genetic signature of the patients. iPSCs were generated from patients with psoriasis (PsO-iPSCs) and healthy donors (Ctr-iPSCs) and were efficiently differentiated into mature KCs. RNA sequencing of KCs derived from Ctr-iPSCs and PsO-iPSCs identified 361 commonly upregulated and 412 commonly downregulated genes. KCs derived from PsO-iPSCs showed dysregulated transcripts associated with psoriasis and KC differentiation, such as HLA-C , KLF4 , chemokines, type I interferon-inducible genes, solute carrier family, IVL , DSG1 , and HLA-DQA1 , as well as transcripts associated with insulin resistance, such as IRS2 , GDF15 , GLUT10 , and GLUT14 . Our data suggest that the KC abnormalities are the main driver triggering psoriasis pathology and highlights the substantial contribution of genetic predisposition in the development of psoriasis and insulin resistance.

FOXA2 is a transcription factor, playing an important role during development. We established an induced pluripotent stem cell (iPSC) line, QBRIi009-A, using non-integrating Sendai virus from a 4-year-old boy, displaying a complex clinical phenotype. Molecular karyotyping and cytogenetics confirmed a de novo proximal 20p11.2 deletion with a reciprocal translocation between the short arm of chromosome 6 and 20. The deleted region (~969 kb) contains only one gene, FOXA2. The generated hiPSC line was fully characterized for its pluripotency and its genetic identity. This iPSC line provides a useful model to study FOXA2 role during human development and in disease pathogenesis., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020. Published by Elsevier B.V.)

Two new species of Schizomyia Kieffer (Diptera: Cecidomyiidae: Asphondyliini: Schizomyiina) are reported from Japan. Schizomyia broussonetiae Elsayed Tokuda n. sp. forms hairy globular leaf galls on the paper mulberry Broussonetia papyrifera (L.) Vent. (Moraceae) and S. uechiae Elsayed and Tokuda n. sp. induces red flower bud galls on the porcelain berry Ampelopsis glandulosa var. brevipedunculata (Maxim.) Momiy. (Vitaceae). Morphological features of larvae, pupae and adults of the new species are described, illustrated and compared in detail to several other Schizomyia species. An updated taxonomic key to known Japanese species of Schizomyia is provided.

Background: The genus Pseudasphondylia (Diptera: Cecidomyiidae: Asphondyliini: Asphondyliina) comprises ten Palearctic, Oriental and Australian species associated with various hosts belonging to at least ten plant families., New Information: A new species, Pseudasphondylia tominagai Elsayed & Tokuda n. sp., that induces flower bud galls on Eleutherococcus spinosus (L.f.) S.Y.Hu (Araliaceae) is described. This species is considered to alternate between host plants seasonally. A key to males of known Pseudasphondylia species is provided.

The genus Asteralobia (Diptera, Cecidomyiidae, Asphondyliini, Schizomyiina) was erected by Kovalev (1964) based on the presence of constrictions on the cylindrical male flagellomeres. In the present study, we examine the morphological features of Asteralobia and Schizomyia and found that the male flagellomeres are constricted also in Schizomyiagaliorum , the type species of Schizomyia . Because no further characters clearly separating Asteralobia from Schizomyia were observed, we synonymize Asteralobia under Schizomyia . Molecular phylogenetic analysis strongly supports our taxonomic treatment. We describe five new species of Schizomyia from Japan, S.achyranthesae Elsayed & Tokuda, sp. n. , S.diplocyclosae Elsayed & Tokuda, sp. n. , S.castanopsisae Elsayed & Tokuda, sp. n. , S.usubai Elsayed & Tokuda, sp. n. , and S.paederiae Elsayed & Tokuda, sp. n. , and redescribe three species, S.galiorum Kieffer, S.patriniae Shinji, and S.asteris Kovalev. A taxonomic key to the Japanese Schizomyia species is provided.

We describe a gall midge Macarangamyia itiokai Elsayed Tokuda gen. n., sp. n. belonging to the subtribe Schizomyiina (Diptera: Cecidomyiidae: Asphondyliini) inducing petiole galls on Macaranga bancana (Miq.) in Lambir Hills National Park, Borneo, Malaysia. The new genus is distinguishable from all known genera of Schizomyiina by the unique dorsally-placed aedeagus slit, the short, membranous, protrusible ovipositor, with scattered strong setae ventrally and dorsally, and the presence of spiracles on all larval thoracic segments. It is compared and separated from its closely related Oriental genera of Schizomyiina.

Background: The expression of a specific combination of transcription factors (TFs) in the multipotent progenitor cells (MPCs) is critical for determining pancreatic cell fate. NKX6.1 expression in PDX1 + MPCs is required for functional β cell generation. We have recently demonstrated the generation of a novel population of human pluripotent stem cell (hPSC)-derived MPCs that exclusively express NKX6.1, independently of PDX1 (PDX1 - /NKX6.1 + ). Therefore, the aim of this study was to characterize this novel population to elucidate its role in pancreatic development., Methods: The hPSCs were exposed to two differentiation protocols to generate MPCs that were analyzed using different techniques., Results: Based on the expression of PDX1 and NKX6.1, we generated three different populations of MPCs, two of them were NKX6.1 + . One of these NKX6.1 populations coexpressed PDX1 (PDX1 + /NKX6.1 + ) which is known to mature into functional β cells, and an additional novel population did not express PDX1 (PDX1 - /NKX6.1 + ) with an undefined role in pancreatic cell fate. This novel population was enriched using our recently established protocol, allowing their reorganization in three-dimensional (3D) structures. Since NKX6.1 induction in MPCs can direct them to endocrine and/or ductal cells in humans, we examined the coexpression of endocrine and ductal markers. We found that the expression of the pancreatic endocrine progenitor markers chromogranin A (CHGA) and neurogenin 3 (NGN3) was not detected in the NKX6.1 + 3D structures, while few structures were positive for NKX2.2, another endocrine progenitor marker, thereby shedding light on the origin of this novel population and its role in pancreatic endocrine development. Furthermore, SOX9 was highly expressed in the 3D structures, but cytokeratin 19, a main ductal marker, was not detected in these structures., Conclusions: These data support the existence of two independent NKX6.1 + MPC populations during human pancreatic development and the novel PDX1 - /NKX6.1 + population may be involved in a unique trajectory to generate β cells in humans.

Male germ cell differentiation is a subtle and complex regulatory process. Currently, its regulatory mechanism is still not fully understood. In our experiment, we performed the first comprehensive genome and transcriptome-wide analyses of the crucial genes and signaling pathways in three kinds of crucial cells (embryonic stem cells, primordial germ cell, and spermatogonial stem cells) that are associated with the male germ cell differentiation. We identified thousands of differentially expressed genes in this process, and from these we chose 173 candidate genes, of which 98 genes were involved in cell differentiation, 19 were involved in the metabolic process, and 56 were involved in the differentiation and metabolic processes, like GAL9, AMH, PLK1, and PSMD7 and so on. In addition, we found that 18 key signaling pathways were involved mainly in cell proliferation, differentiation, and signal transduction processes like TGF-β, Notch, and Jak-STAT. Further exploration found that the candidate gene expression patterns were the same between in vitro induction experiments and transcriptome results. Our results yield clues to the mechanistic basis of male germ cell differentiation and provide an important reference for further studies., (© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.)

Here, we explore the regulatory mechanism of lipid metabolic signaling pathways and related genes during differentiation of male germ cells in chickens, with the hope that better understanding of these pathways may improve in vitro induction. Fluorescence-activated cell sorting was used to obtain highly purified cultures of embryonic stem cells (ESCs), primitive germ cells (PGCs), and spermatogonial stem cells (SSCs). The total RNA was then extracted from each type of cell. High-throughput analysis methods (RNA-seq) were used to sequence the transcriptome of these cells. Gene Ontology (GO) analysis and the KEGG database were used to identify lipid metabolism pathways and related genes. Retinoic acid (RA), the end-product of the retinol metabolism pathway, induced in vitro differentiation of ESC into male germ cells. Quantitative real-time PCR (qRT-PCR) was used to detect changes in the expression of the genes involved in the retinol metabolic pathways. From the results of RNA-seq and the database analyses, we concluded that there are 328 genes in 27 lipid metabolic pathways continuously involved in lipid metabolism during the differentiation of ESC into SSC in vivo, including retinol metabolism. Alcohol dehydrogenase 5 (ADH5) and aldehyde dehydrogenase 1 family member A1 (ALDH1A1) are involved in RA synthesis in the cell. ADH5 was specifically expressed in PGC in our experiments and aldehyde dehydrogenase 1 family member A1 (ALDH1A1) persistently increased throughout development. CYP26b1, a member of the cytochrome P450 superfamily, is involved in the degradation of RA. Expression of CYP26b1, in contrast, decreased throughout development. Exogenous RA in the culture medium induced differentiation of ESC to SSC-like cells. The expression patterns of ADH5, ALDH1A1, and CYP26b1 were consistent with RNA-seq results. We conclude that the retinol metabolism pathway plays an important role in the process of chicken male germ cell differentiation.

The Cecidomyiidae (Diptera: Bibionomorpha) fauna of Egypt is poorly known. Investigations in northern Egypt in 2013 revealed the presence of seven species of gall midges on three host plant species: Atriplex halimus L., Arthrocnemum macrostachyum (Moric.) and Suaeda pruniosa Lange (all Chenopodiaceae). Among the gall midges, Baldratia salicorniae  Kieffer and Stefaniella trinacriae De Stefani are reconfirmed records in Egypt; Houardiella gracilis Dorchin & Freidberg and Asphondylia punica Marchal are new records; and Baldratia karamae Elsayed & Skuhraván. sp. , Primofavilla aegyptiaca Elsayed n. sp. and Stefaniella skuhravae Elsayed n. sp. are new to science. Adult morphology of the latter three new species is described and illustrated, and their biology and geographic distribution are given.

Differentiation of germ cells from embryonic stem cells in vitro could have great application for treating infertility and provide an excellent model for uncovering molecular mechanisms of germline generation. In this study, we aim to screen the suitable inducers that may prove the efficiency of driving chicken embryonic stem cells (ES cells) toward germ cells. The male ES cells were separeted into different groups: single retinoic acid (RA) treatment, co-cultured with sertoli cell feeder with RA induction, cultured on matrix proteins (fibronectin, laminin and collagen) with RA treatment, cultured on fibronectin with sertoli cell feeder and RA induction, and single bone morphogenetic protein 4 (BMP4) treatment. Quantitative RT-PCR and immunoourescence were performed to characterize the ES cells differentiation process. The results showed that spermatogonial stem cells (SSCs)-like were not detected in single RA and RA with collagen groups, but were observed in the other groups. The expression of ES specific genes (Nanog and Sox2) was decreased while SSCs marker genes (Dazl, Stra8, integrin α6, integrinβ1 and C-kit) was remarkably increased. The multiple comparsion results showed that the expression of SSCs marker genes in RA with sertoli cells group was significantly higher than the other groups(P<0.05). Collectively, our results suggested that chicken ES cells possess the potency to differentiate into SSCs-like cells in vitro through RA, matrix proteins, sertoli cells and BMP4 induction, of which co-cultured with sertoli cell feeder with RA induction was proved to be the best., (Copyright © 2014 Elsevier B.V. All rights reserved.)

This experiment was conducted to explore the biological functions of myogenin (MyoG) gene. MyoG gene was cloned from genome of Hu sheep by overlap extension PCR. Then, pEGFP-C1-MyoG and pcDNA3.0-MyoG fusion expression vectors was constructed and pEGFP-C1-MyoG vector had been transfected into NIH-3T3 cells by liposomes-mediated method, and MyoG was detected in vitro by RT-PCR,western blotting and its subcellular localization by EGFP marker. pcDNA3.0-MyoG was transfected into goat embryonic fibroblasts (GEF) cells in order to detect the myogenic function of MyoG in vitro. Then pEGFP-C1-MyoG plasmid was injected into the testes of sheep and goat, respectively, to produce the transgenic generation. The results showed that the length of MyoG coding region of Hu sheep was 675 bp, encoding 224 amino acids. Compared with goat, cattle, pig and rat, the sequence homology of sheep MyoG cDNA was 99.26, 97.04, 92.00, and 87.70 %, respectively. The bioinformatics prediction showed that MyoG protein contained a typical bHLH structure, but without a short signal peptide, revealing that MyoG protein might be a non-secretory protein. The result of RT-PCR and western blotting demonstrated that MyoG could be expressed successfully in the transfected cells in vitro and the MyoG protein was located in nucleus. The positive transfected GEF cells with pcDNA3.0-MyoG were found to express desmin protein. The positive rates of transgenic sheep and transgenic goat were 7.1 and 7.4 % in F1 generation, respectively. Conclusively, MyoG cDNA from Hu sheep had been cloned successfully. The subcellular localization and myogenic activity of MyoG were exactly detected on the basis of multiple biological analyses, which expanded our understanding of the biological function of MyoG.

As an attempt to increase the resistance to Newcastle Disease Virus (NDV) and so further reduction of its risk on the poultry industry. This work aimed to build the eukaryotic gene co-expression plasmid of neuraminidase (NA) gene and myxo-virus resistance (Mx) and detect the gene expression in transfected mouse fibroblasts (NIH-3T3) cells, it is most important to investigate the influence of the recombinant plasmid on the chicken embryonic fibroblasts (CEF) cells. cDNA fragment of NA and mutant Mx gene were derived from pcDNA3.0-NA and pcDNA3.0-Mx plasmid via PCR, respectively, then NA and Mx cDNA fragment were inserted into the multiple cloning sites of pVITRO2 to generate the eukaryotic co-expression plasmid pVITRO2-Mx-NA. The recombinant plasmid was confirmed by restriction endonuclease treatment and sequencing, and it was transfected into the mouse fibroblasts (NIH-3T3) cells. The expression of genes in pVITRO2-Mx-NA were measured by RT-PCR and indirect immunofluorescence assay (IFA). The recombinant plasmid was transfected into CEF cells then RT-PCR and the micro-cell inhibition tests were used to test the antiviral activity for NDV. Our results showed that co-expression vector pVITRO2-Mx-NA was constructed successfully; the expression of Mx and NA could be detected in both NIH-3T3 and CEF cells. The recombinant proteins of Mx and NA protect CEF cells from NDV infection until after 72 h of incubation but the individually mutagenic Mx protein or NA protein protects CEF cells from NDV infection till 48 h post-infection, and co-transfection group decreased significantly NDV infection compared with single-gene transfection group (P<0. 05), indicating that Mx-NA jointing contributed to delaying the infection of NDV in single-cell level and the co-transfection of the jointed genes was more powerful than single one due to their synergistic effects.

Unlabelled: With 14 figures and 3 tables, Summary: Each adrenal gland consisted of cortex and medulla that developed from different embryological origins and presented different cellular organization. One hundred male or female camel embryos or fetuses with crown vertebral rump lengths (CVRL) that ranged from 0.8 to 117 cm were examined. The adrenal cortex, which is derived from intermediate mesoderm, was first observed in the 0.8-cm CVRL camel embryo. The adrenal cortex initially was combined with the gonad as a thickened region of proliferating cells derived from splanchnic intermediate mesoderm. Adrenocortical tissue was first separated from the gonadal tissue in the 2-cm CVRL camel fetus and was observed as a separate dorso-medial mass of cells. At 2.5-cm CVRL, the adrenocortical tissue was surrounded by a capsule of undifferentiated mesenchymal cells, except at its proximal pole, where an invagination was located through which chromaffinoblast cells entered the cortex. The chromaffinoblast cells migrated from the neural crest to form the medulla of the developing adrenal gland. In the 3.5-cm CVRL camel fetus, the adrenocortical cells differentiated into two layers: the inner fetal cortex and the outer definitive cortex. As development proceeded, the fetal cortex degenerated and the definitive cortex formed the zona glomerulosa and zona fasciculata. The zona reticularis did not form until the end of gestation. During prenatal life, the adrenal medulla was much thicker than the cortex., (© 2010 Blackwell Verlag GmbH.)