Effect of potassium simplex optimization medium (KSOM) and embryo screening on the production of human lactoferrin transgenic cloned dairy goats

In this study, we produced cloned transgenic dairy goat based on dairy goat ear skin fibroblast as donor cells for nuclear transfer (NT), which were modified by human lactoferrin (hLF) gene. The developmental competence of NT embryos was compared with either between different embryo culture medium, potassium simplex optimization medium (KSOM) and tissue culture medium (TCM 199), or different classification of NT embryos (48 h after fusion). First we cultured NT embryos to cleavage stage (48 h after fusion) by TCM 199 supplemented with 1 mg/ml bovine serum albumin BSA and KSOM, then used TCM 199 supplemented with 10% FBS to culture them to blastula stage. The results show that the NT embryos in KSOM (19.5%) were superior to TCM 199 (10.6%) in blastulation. In the second experiment, we found that the growth rate of NT embryos (48 h after fusion) was different, then we divided them into four groups: 2-cell, 3- to 4-cell, 5- to 8-cell and >8-cell in stereo microscope and cultured them in vitro respectively. The results show day-2 embryos at 3-4cell and 5-8cell stage (31.9 and 28.2%, P < 0.05) had higher blastocyst formation rates than those at both 2-cell (9.1%) and >8-cell (8.3%) stage, and finally three healthy cloned transgenic goat were successfully produced using 3-8 cell embryos at Day-2 (82%). Using Hoechst 33342 staining, we also found that the >8 cells embryos at Day- 2 demonstrated higher frequency of fragmentation, which may be the one cause of the low blastocyst formation rate. This study therefore demonstrates that KSOM medium could be selected as the early embryo culture medium, and 3-8 cell embryos at day-2 (48 h after fusion) may be the suitable embryos for transplantation, which could reduce the nuclei fragmentation and result in good quality blastocysts that may also enhance the efficiency of transgenic cloned dairy goats production, as well as decrease the economic loss due to embryonic mortality when embryos are transferred to synchronized recipients.Key words: Nuclear transfer, KSOM, transgenic, human lactoferrin, dairy goat

KASP-IEva: an intelligent typing evaluation model for KASP primers

KASP marker technology has been used in molecular marker-assisted breeding because of its high efficiency and flexibility, and an intelligent evaluation model of KASP marker primer typing results is essential to improve the efficiency of marker development on a large scale. To this end, this paper proposes a gene population delineation method based on NTC identification module and data distribution judgment module to improve the accuracy of K-Means clustering, and introduces a decision tree to construct the KASP-IEva primer typing evaluation model. The model firstly designs the NTC identification module and data distribution judgment module to extract four types of data, grouping and categorizing to achieve the improvement of the distinguishability of amplification product signals; secondly, the K-Means algorithm is used to aggregate and classify the data, to visualize the five aggregated clusters and to obtain the morphology location eigenvalues; lastly, the evaluation criteria for the typing effect level are constructed, and the logical decision tree is used to make conditional discrimination on the eigenvalues in order to realize the score prediction. The performance of the model was tested by the KASP marker typing test results of 2519 groups of cotton varieties, and the following conclusions were obtained: the model is able to visualize the aggregation and classification effects of the amplification products of NTC, pure genotypes, heterozygous genotypes, and untyped genotypes, enabling rapid and accurate KASP marker typing evaluation. Comparing and analyzing the model evaluation results with the expert evaluation results, the average accuracy rate of the four grades evaluated by the model was 87%, and the overall evaluation results showed an uneven distribution of the grades with significant differential characteristics. When evaluating 2519 KASP fractal maps, the expert evaluation consumes 15 hours, and the model evaluation only uses 8min27.45s, which makes the model intelligent evaluation significantly better than the expert evaluation from the perspective of time. The establishment of the model will further enhance the application of KASP markers in molecular marker-assisted breeding and provide technical support for the large-scale screening and identification of excellent genotypes

A cross-sectional study of predictive factors of health literacy among rheumatoid arthritis patients in China

ObjectiveTo investigate the factors that influence health literacy (HL) among Chinese patients with rheumatoid arthritis (RA) and furnish theoretical underpinnings for the development of intervention strategies aimed at enhancing patients’ quality of life.MethodsFrom May 2022 to December 2022, a comprehensive survey was conducted among both outpatients and inpatients diagnosed with (RA) in a tertiary hospital in China. The survey utilized various instruments, including a general information questionnaire, a chronic disease patient health literacy scale, the Health Assessment Questionnaire-Disability Index (HAQ-DI), the Chinese-translated Rheumatoid Arthritis Self-Efficacy Scale, the Chinese-translated Rheumatoid Arthritis Stigma Scale, and the Chinese-translated Compliance Questionnaire for Rheumatology Treatments.ResultsThe average scores of HL, self-efficacy, medication adherence, and disability index were 83.54 ± 17.43, 84.91 ± 14.37, 70.16 ± 11.24, and 0.26 ± 0.44, respectively. HL in Chinese RA patients was negatively correlated with age, erythrocyte sedimentation rate (ESR), number of tender joints, number of swollen joints, and disease activity, while positively correlated with self-efficacy and medication adherence. Age, disease activity, disability index, self-efficacy, and medication adherence are predictive factors of HL, and a predictive model has been initially constructed.ConclusionIn the management of RA, healthcare professionals should develop and implement effective intervention measures by focusing on improving medication adherence, enhancing patients’ self-efficacy, improving patients’ physical function, and reducing disease activity. This will help enhance the health literacy and promote clinical outcomes in RA patients

Multimodal mechanisms of pathogenic variants in the signal peptide of FIX leading to hemophilia B

Signal peptide (SP) is essential for protein secretion, and pathogenic variants in the SP of factor IX (FIX) have been identified in hemophilia B (HB). However, the underlying mechanism for the genotype-phenotype correlation of these variants has not been well studied. Here, we systematically examined the effects of 13 pathogenic point variants in the SP of FIX using different approaches. Our results showed that these point variants lead to HB by missense variants and/or aberrant premessenger RNA (pre-mRNA) splicing. The missense variants in a hydrophobic core (h-region) mainly affected the cotranslational translocation function of the SP, and those in C-terminal containing cleavage site (c-region) caused FIX deficiency mainly by disturbing the cotranslational translocation and/or cleavage of the SP. Almost absolute aberrant pre-mRNA splicing was only observed in variants of c.82T\u3eG, but a slight change of splicing patterns was found in variants of c.53G\u3eT, c.77C\u3eA, c.82T\u3eC, and c.83G\u3eA, indicating that these variants might have different degrees of impact on pre-mRNA splicing. Although two 6-nt deletion aberrant pre-mRNA splicing products caused FIX deficiency by disturbing the SP cleavage, they could produce some functional mature FIX, and vitamin K could increase the secretion of functional FIX. Taken together, our data indicated that pathogenic variants in the SP of FIX caused HB through diverse molecular mechanisms or even a mixture of several mechanisms, and vitamin K availability could be partially attributed to varying bleeding tendencies in patients carrying the same variant in the SP

Investigation of the prevalence and clinical implications of ERBB2 exon 16 skipping mutations in Chinese pan-cancer patients

BackgroundAlthough rare, ERBB2 exon 16 skipping mutations (ERBB2ΔEx16) have been implicated in resistance to anti-HER2 and anti-EGFR targeted agents. Our study investigated the prevalence and clinical significance of ERBB2ΔEx16 in Chinese pan-cancer patients.MethodsWe retrospectively screened 40996 patients, spanning 19 cancer types, who had available genomic profiles acquired with DNA-based next-generation sequencing (NGS). We characterized the clinical and molecular features of the ERBB2ΔEx16-positive patients. Furthermore, we also analyzed a pan-cancer dataset from the Cancer Genome Atlas (TCGA; n=8705).ResultsA total of 22 patients were detected with ERBB2ΔEx16, resulting in an overall prevalence rate of 0.054% (22/40996). Of them, 16 patients had lung cancer (LC; 0.05%, 16/30890), five patients had gastric cancer (GC; 0.35%, 5/1448), and one patient had ovarian cancer (0.12%, 1/826). Among the 16 LC patients, ERBB2ΔEx16 was detected in four treatment-naïve EGFR/ALK-negative patients and 12 EGFR-positive patients after the onset of resistance to EGFR tyrosine kinase inhibitors (TKIs). The treatment-naïve patients harbored no LC-associated oncogenic drivers except ERBB2 amplification, suggesting a potential oncogenic role for ERBB2ΔEx16. Consistently, ERBB2ΔEx16+ patients from TCGA data also carried no known drivers despite various concurrent alterations. In the 12 EGFR TKI-resistant LC patients, relative variant frequencies for ERBB2ΔEx16 were lower than in untreated patients, suggesting ERBB2ΔEx16 as secondary alterations following TKI treatment and thereby implicating ERBB2ΔEx16 in mediating therapeutic resistance.ConclusionsOur study identified an overall ERBB2ΔEx16 prevalence rate of 0.054% and provided insights into the clinical implications of ERBB2ΔEx16 in Chinese pan-cancer patients

Progress of nanoparticle drug delivery system for the treatment of glioma

Gliomas are typical malignant brain tumours affecting a wide population worldwide. Operation, as the common treatment for gliomas, is always accompanied by postoperative drug chemotherapy, but cannot cure patients. The main challenges are chemotherapeutic drugs have low blood-brain barrier passage rate and a lot of serious adverse effects, meanwhile, they have difficulty targeting glioma issues. Nowadays, the emergence of nanoparticles (NPs) drug delivery systems (NDDS) has provided a new promising approach for the treatment of gliomas owing to their excellent biodegradability, high stability, good biocompatibility, low toxicity, and minimal adverse effects. Herein, we reviewed the types and delivery mechanisms of NPs currently used in gliomas, including passive and active brain targeting drug delivery. In particular, we primarily focused on various hopeful types of NPs (such as liposome, chitosan, ferritin, graphene oxide, silica nanoparticle, nanogel, neutrophil, and adeno-associated virus), and discussed their advantages, disadvantages, and progress in preclinical trials. Moreover, we outlined the clinical trials of NPs applied in gliomas. According to this review, we provide an outlook of the prospects of NDDS for treating gliomas and summarise some methods that can enhance the targeting specificity and safety of NPs, like surface modification and conjugating ligands and peptides. Although there are still some limitations of these NPs, NDDS will offer the potential for curing glioma patients

Abnormal Topology of the Structural Connectome in the Limbic Cortico-Basal-Ganglia Circuit and Default-Mode Network Among Primary Insomnia Patients

Purpose: Primary insomnia (PI) is the second most common mental disorder. However, the topologic alterations in structural brain connectome in patients with PI remain largely unknown.Methods: A total of 44 PI patients and 46 age-, gender-, and education level matched healthy control (HC) participants were recruited in this study. Diffusion tensor imaging (DTI) and resting state MRI were used to construct structural connectome for each participant, and the network parameters were employed by non-parametric permutations to evaluate the significant differences between the two groups. Relationships between abnormal network metrics and clinical characteristics, including the disease duration, the Pittsburgh Sleep Quality Index (PSQI), the Insomnia Severity Index (ISI), the Self-Rating Anxiety Scale (SAS), and the Self-Rating Depression Scale (SDS), were investigated with Spearman’s correlation analysis in PI patients.Results: PI patients demonstrated small-world architecture with lower global (P = 0.005) and local (P = 0.035) efficiencies compared with the HC group. The unique hub nodal properties in PI patients were mainly in the right limbic cortico-basal-ganglia circuit. Five disrupted subnetworks in PI patients were observed in the limbic cortico-basal-ganglia circuit and left default-mode networks (DMN) (P < 0.05, NBS corrected). Moreover, most unique hub nodal properties in the right limbic cortico-basal-ganglia circuit were significantly correlated with disease duration, and clinical characteristics (SAS, SDS, ISI scores) in PI processing.Conclusion: These findings suggested the abnormal anatomical network architecture may be closely linked to clinical characteristics in PI. The study provided novel insights into the neural substrates underlying symptoms and neurophysiologic mechanisms of PI

Difficulties and measures of driving super long piles in Bohai Gulf

Long piles of the ocean oil platform are usually manufactured as the integration of several segments, which have to be assembled one by one during installation. During pile driving, excessive pore pressure will build up in such a high level that hydraulic fracturing in the soil round the pile may take place, which will cause the soil to consolidate much faster during pile extension period. Consequently, after pile extension, the soil strength will recover to some extent and the driving resistance will increase considerably, which makes restarting driving the pile very difficult and even causes refusal. A finite element (FE) analysis procedure is presented for judging the risk of refusal by estimating the blow counts after pile extension, in which the regain of soil strength is considered. A case analysis in Bohai Gulf is performed using the proposed procedure to explain the pile refusal phenomenon