Normal Numbers Research Articles

Murine uterine gland branching is necessary for gland function in implantation.

Uterine glands are branched, tubular structures whose secretions are essential for pregnancy success. It is known that pre-implantation glandular expression of leukemia inhibitory factor (LIF) is crucial for embryo implantation; however, the contribution of uterine gland structure to gland secretions, such as LIF, is not known. Here, we use mice deficient in estrogen receptor 1 (ESR1) signaling to uncover the role of ESR1 signaling in gland branching and the role of a branched structure in LIF secretion and embryo implantation. We observed that deletion of ESR1 in neonatal uterine epithelium, stroma, and muscle using the progesterone receptor PgrCre causes a block in uterine gland development at the gland bud stage. Embryonic epithelial deletion of ESR1 using a Müllerian duct Cre line, Pax2Cre, displays gland bud elongation but a failure in gland branching. Reduction of ESR1 in adult uterine epithelium using the lactoferrin-Cre (LtfCre) displays normally branched uterine glands. Unbranched glands from Pax2Cre Esr1flox/flox uteri fail to express glandular pre-implantation Lif, preventing implantation chamber formation and embryo alignment along the uterine mesometrial-antimesometrial axis. In contrast, branched glands from LtfCre Esr1flox/flox uteri display reduced expression of ESR1 and glandular Lif resulting in delayed implantation chamber formation and embryo-uterine axes alignment but mice deliver a normal number of pups. Finally, pre-pubertal unbranched glands in control mice express Lif in the luminal epithelium but fail to express Lif in the glandular epithelium, even in the presence of estrogen. These data strongly suggest that branched glands are necessary for pre-implantation glandular Lif expression for implantation success. Our study is the first to identify a relationship between the branched structure and secretory function of uterine glands and provides a framework for understanding how uterine gland structure-function contributes to pregnancy success.

Penerapan Rom (Range Of Motion) Dan Massage Punggung Untuk Mencegah Luka Tekan Pada Lansia Tirah Baring

Immobilization is an extrinsic variable that can cause problems related to skin integrity. The inability to move freely due to situations that interfere with or limit movement may be directly related to an internal problem such as a chronic illness or health condition. Pressure ulcers or also known as decubitus ulcers, are local injuries caused by pressure or a combination of friction and pressure on the skin and/or underlying tissue, usually over a bony prominence. This research method uses a descriptive case study. The research population consisted of 1 person, namely a patient with heart failure with a nursing diagnosis of Activity Intolerance. The evaluation results from providing back massage (efflurage) and range of motion (ROM) to the client showed a reduction in complaints of soreness while in bed. After being taught range of motion (ROM) the client can do it themselves every time the client wakes up. Apart from that, clients also say they feel fresher after doing efflurage massage. The conclusion from applying ROM to these patients is that there is an increase in the Braden scale score which shows that there is a reduced risk of decubitus or pressure sores in the elderly. Meanwhile, when giving effluerage back massage, results were obtained in the form of client validation which said that the client was fresher, and there was a change in blood pressure results to normal numbers.

Cytological Studies in the Genus <i>Amaryllis</i>

One species and eight horticultural varieties of Amaryllis show a basic number of x=11, which is the predominant basic number of the family. In both Amaryllis belladonna and Amaryllis-Mrs. Garfield the normal somatic number is 2n=22. All the seven colour varieties of Amaryllis-Giant Dutch have 2n=44 chromosomes. These varieties differ from one another in the minor alterations of the chromosome types suggesting the role of minor structural alterations in evolution. In addition to the normal somatic complement a number of variant nuclei with altered karyotypes have been recorded.

Concentration of fatty acid binding protein as a new indicator of ejaculate fertility

Objective. To determine the marker function of fatty acid binding protein (FABP) concentration in seminal plasma (SP) in order to assess ejaculate fertility used in in vitro fertilization (IVF) treatment of infertility in couples. Materials and methods. The study involved semen samples of 96 men of reproductive age: the study group (n=63) – patients with a decrease in concentration and/or total content of spermatozoa in ejaculate, the comparison group (n=33) – men with normal concentration and number of spermatozoa in ejaculate. The content of FABP in SP was determined by enzyme-linked immunosorbent assay using the test system “FABP – ELISA – BEST” (A-9102, Vector-Best, Russia). In order to determine the informative value of using the concentration of FABP in SP as a criterion of ejaculate fertility of men of the study group, the predictive value of positive and negative IVF results was determined by calculating the diagnostic sensitivity, specificity and efficiency. Results. The content of FABP in SP accounted for 1.29 ± 0.24 ng/mL, the median and interquartile range comprised 1.23 [1.13–1.35] ng/mL, ranging from 1.08 to 2.79 ng/mL. The study revealed statistically significant intergroup differences (Mann-Whitney test U=79.00; p=0.000016), a weak correlation between the level of FABP and the concentration of spermatozoa (R=0.578008) and their number in ejaculate (R=0.583599). The diagnostic sensitivity of the test for FABP in SP of the men of the study group accounted for 81.82%, specificity – 78.95%, efficiency – 80.95%. Conclusion. Seminal plasma FABP can act as a marker of spermatogenesis disorders. The study of this protein in ejaculate provides the accuracy of predicting the outcome of in vitro fertilization.

Complex Pythagorean Normal Interval-Valued Fuzzy Aggregation Operators for Solving Medical Diagnosis Problem

This paper presents a new methodology for solving multiple-attribute decision-making problems (MADMs) using a complex Pythagorean normal interval-valued fuzzy set (CPNIVFS), which is an extended concept of a complex Pythagorean fuzzy set. Four types of different aggregating operations (AOs), including CPNIVF weighted averaging (CPNIVFWA), CPNIVF weighted geometric (CPNIVFWG), generalized CPNIVFWA (CGPNIVFWA), and generalized CPNIVFWG (CGPNIVFWG), are discussed. The scoring function, accuracy function, and operational laws of the CPNIVFS are defined. Algebraic structures, such as associative, distributive, idempotent, bounded, commutativity, and monotonicity properties, are also shown to be satisfied by complex Pythagorean normal interval-valued fuzzy numbers. Furthermore, an algorithm is proposed to solve the MADM problems based on the defined AOs. The proposed approach is then used for a medical diagnosis problem about brain tumors because computer science and machine tool technology are among the most important components of brain tumor research. The five types of brain tumors diagnosed in these patients are gliomas, meningiomas, metastases, embryonal tumors, and ependymomas. Several types of treatments are available, which are often combined as part of an overall treatment plan. Brain tumors can be treated in various ways, including surgery, radiation therapy, chemotherapy, immunotherapy, and clinical trials. Based on the comparisons and options gathered, the most suitable treatment can be chosen. In this regard, it is evident that the value of the integer ⅁\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\Game $$\\end{document} plays a significant role in determining the model. The candidate models under consideration can be validated by comparing them with the previously proposed ones. The proposed technique is compared with the existing method to demonstrate its superiority and validity, and the results conclude that the former is more reliable and effective than the latter. Finally, the criteria are evaluated by expert assessments to determine the most appropriate options.

A Spinocerebellar Ataxia Type 6 Patient Caused by <i>De Novo</i> Expansion of Normal Range CAG Repeats

Spinocerebellar ataxia type 6 (SCA6) is classified as a CAG repeat disorder, where the number of expanded CAG repeats often undergoes meiotic instability, when transmitted from one generation to the next. However, in SCA6, both normal and expanded CAG repeats tend to remain stable during transmission due to its relatively small repeat numbers. We herein report <i>de-novo</i> expansion of CAG repeats in SCA6 gene in a 41-year-old female patient, whose parents had normal repeat numbers.

The ion channel TRPV5 regulates B-cell signaling and activation.

B-cell activation triggers the release of endoplasmic reticulum calcium stores through the store-operated calcium entry (SOCE) pathway resulting in calcium influx by calcium release-activated calcium (CRAC) channels on the plasma membrane. B-cell-specific murine knockouts of SOCE do not impact humoral immunity suggesting that alternative channels may be important. We identified a member of the calcium-permeable transient receptor potential (TRP) ion channel family, TRPV5, as a candidate channel expressed in B cells by a quantitative polymerase chain reaction (qPCR) screen. To further investigate the role of TRPV5 in B-cell responses, we generated a murine TRPV5 knockout (KO) by CRISPR-Cas9. We found TRPV5 polarized to B-cell receptor (BCR) clusters upon stimulation in a PI3K-RhoA-dependent manner. TRPV5 KO mice have normal B-cell development and mature B-cell numbers. Surprisingly, calcium influx upon BCR stimulation in primary TRPV5 KO B cells was not impaired; however, differential expression of other calcium-regulating proteins, such as ORAI1, may contribute to a compensatory mechanism for calcium signaling in these cells. We demonstrate that TRPV5 KO B cells have impaired spreading and contraction in response to membrane-bound antigen. Consistent with this, TRPV5 KO B cells have reduced BCR signaling measured through phospho-tyrosine residues. Lastly, we also found that TRPV5 is important for early T-dependent antigen specific responses post-immunization. Thus, our findings identify a role for TRPV5 in BCR signaling and B-cell activation.

Random walks on tori and normal numbers in self-similar sets

abstract: We study random walks on a $d$-dimensional torus by affine expanding maps whose linear parts commute. Assuming an irrationality condition on their translation parts, we prove that the Haar measure is the unique stationary measure. We deduce that if $K\subset\mathbb{R}^d$ is an attractor of a finite iterated function system of $n\geq 2$ maps of the form $x\mapsto D^{-1}x+t_i$ ($i=1,\dotsc,n$), where $D$ is an expanding $d\times d$ integer matrix, and is the same for all the maps, under an irrationality condition on the translation parts $t_i$, almost every point in $K$ (w.r.t. any Bernoulli measure) has an equidistributed orbit under the map $x\mapsto Dx$ (multiplication mod $\mathbb{Z}^d$). In the one-dimensional case, this conclusion amounts to normality to base $D$. Thus for example, almost every point in an irrational dilation of the middle-thirds Cantor set is normal to base $3$.

Abstract 2268: CREDAC: Copy number-based reference-free expression deconvolution analysis of cancers

Abstract Introduction: Analyses of bulk RNA sequencing data are central to most large-scale tumor sequencing studies. Bulk expression data represent population averages and its interpretation is confounded by both normal cell contamination and somatic copy number alterations. Several computational methods to deconvolve have been developed recently. However, these methods often rely on a set of cell-type-specific reference signatures and ignore the effect of copy number changes. Methods: To address these issues, we have developed a method that formalizes the relationship between allele-specific copy number, expression and sample purity to deconvolve the expression profiles of tumor and normal cells from bulk RNA-seq in an unbiased manner. Our method was applied to sequencing data produced by the TRACERx consortium, a longitudinal study with multi-region whole-exome and RNA-seq of non-small-cell lung cancers. A total of 414 primary tumor regions and 140 adjacent normal tissue samples from 140 patients with matched DNA and RNA-seq data were processed. Results: We were able to directly deconvolve a median of ~2,000 genes per sample and indirectly infer tumor and normal expression profiles of ~10,000 genes using an scaled and weighted mean approach. The accuracy of the deconvolution was validated using (1) in-silico mixtures of patient-derived tumor and normal cells, (2) pseudo-bulk scRNAseq and (3) in regions with loss of heterogeneity (LOH) directly on the bulk sequencing data, where the total fraction of expression attributed to tumor cells can be computed directly using somatic mutations. CREDAC allows for single sample bulk-level tumor-normal differential expression analysis and revealed a strong and constitutive genome-wide overexpression in cancer compared to admixed normal cells, with a more pronounced overexpression in lung squamous cell carcinoma than lung adenocarcinoma (p<0.001). CREDAC adjusts for variations in copy number, thus facilitating the investigation of cancer-specific dosage compensatory responses triggered by aneuploidy. We revealed that a majority of genes (~65%) exhibit proportionality to copy number alterations, while a substantial proportion (~35%) shows anti-scaling or dosage compensation strategies. We show an enrichment of genes involved in cell cycle and expression processes within the dosage compensated group and revealed methylation as a key mechanism driving this compensatory process. Conclusion: Overall, our results suggest that CREDAC is able to accurately disentangle the expression of tumor and normal cells from bulk RNA-seq without any previous knowledge. It has potential applications in many studies that include matched RNA-seq and copy number data and can provide new insights functional characterization, the taxonomy of cancer, and tumor evolution. Citation Format: Carla Castignani, Jonas Demeulemeester, Oriol Pich, Tom Lesluyes, Robert E. Hynds, David R. Pearce, Elizabeth Larose Cadieux, Stefan C. Dentro, TRACERx Consortium, Nnenna Kanu, Charles Swanton, Peter Van Loo. CREDAC: Copy number-based reference-free expression deconvolution analysis of cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2268.

Abstract 6602: Targeting the SWI/SNF complex in POU2F-POU2AF transcription factor-driven malignancies

Abstract Background: Small cell lung cancer (SCLC) is a rapidly progressing subtype of lung cancer with a high growth rate and early metastasis propensity, often resulting in a more advanced disease stage at diagnosis. A recent transcriptome analysis of human SCLC tumors revealed that SCLC could be characterized by the expression pattern of certain transcription factors (TFs), including ASCL1 (achaete-scute family bHLH transcription factor 1) and POU2F3 (POU domain class 2 transcription factor 3), exemplifying SCLC as a TF-driven malignancy. ASCL1-driven SCLC (SCLC-A) manifests a neuroendocrine phenotype, while POU2F3-driven SCLC (SCLC-P) is characterized as a tuft-cell-like variant. Recent studies revealed that POU domain class 2 transcription factors uniquely rely on coactivators to achieve their lineage-defining functions in the tuft cell and B cell lineages. In tuft-cell-like SCLC cells, the coactivators of POU2F3 (POU2AF2 and POU2AF3) endow POU2F3 with a critical transactivation domain by forming a master regulator complex, which supports enhancer-mediated cancer-promoting gene activation in SCLC-P cells. This indicates a potential therapeutic vulnerability of tuft-cell-like SCLC whereby strategies aimed at blocking POU2F3-POU2AF2/3 function may lead to clinical benefit. Methods: We conducted a domain-targeted CRISPR screen to identify druggable targets for SCLC-P, followed by pharmacological validation in preclinical SCLC models. Multi-omics techniques, including ATAC-seq, ChIP-seq, RNA-seq, and RIME, were employed to elucidate the SWI/SNF complex's regulatory influence on the POU2F-POU2AF axis. Results: Here we identified that the POU2F3 molecular subtype of SCLC (SCLC-P) exhibits an exquisite dependence on the activity of the SWI/SNF epigenetic complex. SCLC-P cell lines were sensitive to low nanomolar levels of a SWI/SNF ATPase degrader when compared to other molecular subtypes of SCLC. Co-factors of POU2F were found to interact with components of the SWI/SNF complex. The POU2F3 transcription factor complex is evicted from chromatin upon SWI/SNF ATPase degradation. A novel, orally bioavailable SWI/SNF ATPase PROTAC degrader demonstrated preferential efficacy in SCLC-P relative to the SCLC-A subtype. The SWI/SNF ATPase PROTAC degrader did not alter normal tuft cell numbers in lung or colon, nor did it exhibit toxicity in mice. B cell malignancies which displayed a dependency on the POU2F co-factor, POU2AF1, were also remarkably sensitive to SWI/SNF ATPase degradation. In a POU2AF1-dependent, disseminated murine model of multiple myeloma, AU-24118 had an enhanced survival benefit as compared to pomalidomide, an approved treatment for multiple myeloma. Conclusions: Taken together, our studies suggest that POU2F-POU2AF driven malignancies have an intrinsic dependence on the SWI/SNF complex representing a striking therapeutic vulnerability. Citation Format: Lanbo Xiao, Tongchen He, Olaf Klingbeil, Young Eleanor, Xiaoli Wu, Yuanyuan Qiao, Abhijit Parolia, Sanjana Eyunni, Rahul Mannan, Somnath Mahapatra, NamHoon Kim, Heng Zheng, Fengyun Su, Xuhong Cao, Susanta Samajdar, Murali Ramachandra, Christopher R. Vakoc, Arul M. Chinnaiyan. Targeting the SWI/SNF complex in POU2F-POU2AF transcription factor-driven malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6602.

Sensorineural correlates of failed functional recovery after natural regeneration of vestibular hair cells in adult mice.

Vestibular hair cells (HCs) are mechanoreceptors that sense head motions by modulating the firing rate of vestibular ganglion neurons (VGNs), whose central processes project to vestibular nucleus neurons (VNNs) and cerebellar neurons. We explored vestibular function after HC destruction in adult Pou4f3+/DTR (DTR) mice, in which injections of high-dose (50 ng/g) diphtheria toxin (DT) destroyed most vestibular HCs within 2 weeks. At that time, DTR mice had lost the horizontal vestibulo-ocular reflex (aVORH), and their VNNs failed to upregulate nuclear cFos expression in response to a vestibular stimulus (centrifugation). Five months later, 21 and 14% of HCs were regenerated in utricles and horizontal ampullae, respectively. The vast majority of HCs present were type II. This degree of HC regeneration did not restore the aVORH or centrifugation-evoked cFos expression in VNNs. The failure to regain vestibular pathway function was not due to degeneration of VGNs or VNNs because normal neuron numbers were maintained after HC destruction. Furthermore, sinusoidal galvanic stimulation at the mastoid process evoked cFos protein expression in VNNs, indicating that VGNs were able to regulate VNN activity after HC loss. aVORH and cFos responses in VNNs were robust after low-dose (25 ng/g) DT, which compared to high-dose DT resulted in a similar degree of type II HC death and regeneration but spared more type I HCs in both organs. These findings demonstrate that having more type I HCs is correlated with stronger responses to vestibular stimulation and suggest that regenerating type I HCs may improve vestibular function after HC loss.

The transcription factor RhMYB17 regulates the homeotic transformation of floral organs in rose (Rosa hybrida) under cold stress.

Low temperatures affect flower development in rose (Rosa hybrida), increasing petaloid stamen number and reducing normal stamen number. We identified the low-temperature-responsive R2R3-MYB transcription factor RhMYB17, which is homologous to Arabidopsis MYB17 by similarity of protein sequences. RhMYB17 was up-regulated at low temperatures, and RhMYB17 transcripts accumulated in floral buds. Transient silencing of RhMYB17 by virus-induced gene silencing decreased petaloid stamen number and increased normal stamen number. According to the ABCDE model of floral organ identity, class A genes APETALA 1 (AP1) and AP2 contribute to sepal and petal formation. Transcription factor binding analysis identified RhMYB17 binding sites in the promoters of rose APETALA 2 (RhAP2) and APETALA 2-LIKE (RhAP2L). Yeast one-hybrid assays, dual-luciferase reporter assays, and electrophoretic mobility shift assays confirmed that RhMYB17 directly binds to the promoters of RhAP2 and RhAP2L, thereby activating their expression. RNA sequencing further demonstrated that RhMYB17 plays a pivotal role in regulating the expression of class A genes, and indirectly influences the expression of the class C gene. This study reveals a novel mechanism for the homeotic transformation of floral organs in response to low temperatures.

Proband-independent haplotyping based on NGS-based long-read sequencing for detecting pathogenic variant carrier status in preimplantation genetic testing for monogenic diseases.

Preimplantation genetic testing for monogenic diseases (PGT-M) can be used to select embryos that do not develop disease phenotypes or carry disease-causing genes for implantation into the mother's uterus, to block disease transmission to the offspring, and to increase the birth rate of healthy newborns. However, the traditional PGT-M technique has some limitations, such as its time consumption, experimental procedural complexity, and the need for a complete family or reference embryo to construct the haplotype. In this study, proband-independent haplotyping based on NGS-based long-read sequencing (Phbol-seq) was used to effectively construct haplotypes. By targeting the mutation sites of single gene disease point mutations and small fragment deletion carriers, embryos carrying parental disease-causing mutations were successfully identified by linkage analysis. The efficiency of embryo resolution was then verified by classical Sanger sequencing, and it was confirmed that the construction of haplotype and SNP linkage analysis by Phbol-seq could accurately and effectively detect whether embryos carried parental pathogenic mutations. After the embryos confirmed to be nonpathogenic by Phbol-seq-based PGT-M and confirmed to have normal copy number variation by Phbol-seq-based PGT-A were transplanted into the uterus, gene detection in amniotic fluid of the implanted embryos was performed, and the results confirmed that Phbol-seq technology could accurately distinguish normal genotype embryos from genetically modified carrier embryos. Our results suggest that Phbol-seq is an effective strategy for accurately locating mutation sites and accurately distinguishing between embryos that inherit disease-causing genes and normal embryos that do not. This is critical for Phbol-seq-based PGT-M and could help more single-gene disease carriers with incomplete families, de novo mutations or suspected germline mosaicism to have healthy babies with normal phenotypes. It also helps to reduce the transmission of monogenic genetic diseases in the population.

The exact order of discrepancy for Levin’s normal number in base 2

The exact order of discrepancy for Levin’s normal number in base 2

LIGHTWEIGHT ENVIRONMENTS FOR TESTING SPEED AND RELIABILITY OF SOFTWARE BASED ON OPERATING SYSTEM LEVEL VIRTUALIZATION

An environment for performance software testing has been described, based on the use of containerization technology through the use of appropriate software (Docker or others) on the one hand, and a tool for measuring performance and carrying out load testing such as ApacheBench or similar. The advantage of containerization technology for achieving the goal was evaluated. It is shown that in this case, the test environment can be deployed on different computers separately, rather than being deployed centrally. This provides following advantages: less cost in terms of computing resources compared to other ways of deploying environments on local computers. Certain types of environment configurations can be easily distributed and replicated between individual computers and workstations through the distribution and application of prepared configuration files. Compared to cloud environments, this approach does not require additional material costs for support. The scheme of operation of such an environment for non-functional testing has been analyzed.. The environment also allows you to simulate more complex configurations, for example, to study the operation of various load balancing algorithms/systems. The created testing environment was tested on the example of performance of testing web applications. It showed that the created environment allows you to calculate the approximate response time of the application and the presence of failures depending on the number of users who use the application at the same time. Also, load testing allows you to calculate the normal and critical number of application users and predict and prevent potential failures. The mechanism for creating test environments considered in this work can be used to build models that are necessary for studying and/or modeling certain types of software testing based on loads, load balancing algorithms between workstations, and others. Since such an approach can be used on almost any personal workstation with relatively small requirements for computing resources and the deployment and distribution of environments is quite simple, such an approach can be implemented not only in business, but also in universities for courses that involve the use similar environments.

Characterization and establishment of an immortalized rabbit ovary granulosa cell line for reproductive biology experiments.

Granulosa cells (GCs) are the key components of ovarian follicles and regulate the maturation, communication, growth, and development of oocytes. GCs have great potential as human therapeutic models and in livestock breeding. In this study, we established an immortalized cell line (Im-RGCs) by transforming primary rabbit granulosa cells (Pri-RGCs) with lentivirus-mediated simian virus 40 Large T (SV40LT). Morphologically, Im-RGCs were indistinguishable from Pri-RGCs and maintained intact cell structure as observed by H&E staining. Also, Im-RGCs exhibited no significant change in cell proliferation, viability, and growth. Furthermore, GC-specific markers, such as FSHR, StAR, CYP11A1, and CYP19A1, were examined by PCR, immunofluorescence, and Western blotting. ELISA and karyotype analysis showed that Im-RGCs can synthesize steroid hormones and maintain the normal number of chromosomes during the infinite passage. Furthermore, soft-agar cloning and nude mice tumorigenic experiments indicated the absence of any malignancy transformation in Im-RGCs. In conclusion, we successfully established the immortalized granulosa cell line of rabbit follicles that can be used for biological, animal husbandry, and female reproductive research.

Overcoming the egress block of Plasmodium sporozoites expressing fluorescently tagged circumsporozoite protein.

Plasmodium sporozoites are the highly motile and invasive forms of the malaria parasite transmitted by mosquitoes. Sporozoites form within oocysts at the midgut wall of the mosquito, egress from oocysts and enter salivary glands prior to transmission. The GPI-anchored major surface protein, the circumsporozoite protein (CSP) is important for Plasmodium sporozoite formation, egress, migration and invasion. To visualize CSP, we previously generated full-length versions of CSP internally tagged with the green fluorescent protein, GFP. However, while these allowed for imaging of sporogony in oocysts, sporozoites failed to egress. Here, we explore different strategies to overcome this block in egress and obtain salivary gland resident sporozoites that express CSP-GFP. Replacing the N-terminal and repeat region with GFP did not allow sporozoite formation. Lowering expression of CSP-GFP at the endogenous locus allowed sporozoite formation but did not overcome egress block. Crossing of CSP-GFP expressing parasites that are blocked in egress with wild-type parasites yielded a small fraction of parasites that entered salivary glands and expressed various levels of CSP-GFP. Expressing CSP-GFP constructs from a silent chromosome region from promoters that are active only post salivary gland invasion yielded normal numbers of fluorescent salivary gland sporozoites, albeit with low levels of fluorescence. We also show that lowering CSP expression by 50% allowed egress from oocysts but not salivary gland entry. In conclusion, Plasmodium berghei parasites with normal CSP expression tolerate a certain level of CSP-GFP without disruption of oocyst egress and salivary gland invasion.

Whole-Genome Resequencing of Ujimqin Sheep Identifies Genes Associated with Vertebral Number.

The number of vertebrae is a crucial economic trait that can significantly impact the carcass length and meat production in animals. However, our understanding of the quantitative trait loci (QTLs) and candidate genes associated with the vertebral number in sheep (Ovis aries) remains limited. To identify these candidate genes and QTLs, we collected 73 Ujimqin sheep with increased numbers of vertebrae (T13L7, T14L6, and T14L7) and 23 sheep with normal numbers of vertebrae (T13L6). Through high-throughput genome resequencing, we obtained a total of 24,130,801 effective single-nucleotide polymorphisms (SNPs). By conducting a selective-sweep analysis, we discovered that the most significantly selective region was located on chromosome 7. Within this region, we identified several genes, including VRTN, SYNDIG1L, LTBP2, and ABCD4, known to regulate the spinal development and morphology. Further, a genome-wide association study (GWAS) performed on sheep with increased and normal vertebral numbers confirmed that ABCD4 is a candidate gene for determining the number of vertebrae in sheep. Additionally, the most significant SNP on chromosome 7 was identified as a candidate QTL. Moreover, we detected two missense mutations in the ABCD4 gene; one of these mutations (Chr7: 89393414, C > T) at position 22 leads to the conversion of arginine (Arg) to glutamine (Gln), which is expected to negatively affect the protein's function. Notably, a transcriptome expression profile in mouse embryonic development revealed that ABCD4 is highly expressed during the critical period of vertebral formation (4.5-7.5 days). Our study highlights ABCD4 as a potential major gene influencing the number of vertebrae in Ujimqin sheep, with promising prospects for future genome-assisted breeding improvements in sheep.

The clinical significance of low levels of papp-a during first trimester pregnancies in fetuses with normal chromosome number

The clinical significance of low levels of papp-a during first trimester pregnancies in fetuses with normal chromosome number

Phosphorylation of hnRNP A1-Serine 199 Is Not Required for T Cell Differentiation and Function.

hnRNP A1 is an important RNA-binding protein that influences many stages of RNA processing, including transcription, alternative splicing, mRNA nuclear export, and RNA stability. However, the role of hnRNP A1 in immune cells, specifically CD4+ T cells, remains unclear. We previously showed that Akt phosphorylation of hnRNP A1 was dependent on TCR signal strength and was associated with Treg differentiation. To explore the impact of hnRNP A1 phosphorylation by Akt on CD4+ T cell differentiation, our laboratory generated a mutant mouse model, hnRNP A1-S199A (A1-MUT) in which the major Akt phosphorylation site on hnRNP A1 was mutated to alanine using CRISPR Cas9 technology. Immune profiling of A1-MUT mice revealed changes in the numbers of Tregs in the mesenteric lymph node. We found no significant differences in naive CD4+ T cell differentiation into Th1, Th2, Th17, or T regulatory cells (Tregs) invitro. In vivo, Treg differentiation assays using OTII-A1-Mut CD4+ T cells exposed to OVA food revealed migration and homing defects in the A1-MUT but no change in Treg induction. A1-MUT mice were immunized with NP- keyhole limpet hemocyanin, and normal germinal center development, normal numbers of NP-specific B cells, and no change in Tfh numbers were observed. In conclusion, Akt phosphorylation of hnRNP A1 S199 does not play a role in CD4+ T cell fate or function in the models tested. This hnRNP A1-S199A mouse model should be a valuable tool to study the role of Akt phosphorylation of hnRNP A1-S199 in different cell types or other mouse models of human disease.