Efficient generation of cloned cats with altered coat colour by editing of the KIT gene.

Coat colour largely determines the market demand for several cat breeds. The KIT proto-oncogene (KIT) gene is a key gene controlling melanoblast differentiation and melanogenesis. KIT mutations usually cause varied changes in coat colour in mammalian species. In this study, we used a pair of single-guide RNAs (sgRNAs) to delete exon 17 of KIT in somatic cells isolated from two different Chinese Li Hua feline foetuses. Edited cells were used as donor nuclei for somatic cell nuclear transfer (SCNT) to generate cloned embryos presenting an average cleavage rate exceeding 85%, and an average blastocyst formation rate exceeding 9.5%. 131 cloned embryos were transplanted into four surrogates, and all surrogates carried their pregnancies to term, and delivered 4.58% (6/131) alive cloned kittens, with 1.53% (2/131) being KIT-edited heterozygotes (KITD17/+). The KITD17/+ cats presented an obvious darkness reduction in the mackerel tabby coat. Immunohistochemical analysis (IHC) of skin tissues indicated impaired proliferation and differentiation of melanoblasts caused by the lack of exon17 in feline KIT. To our knowledge, this is the first report on coat colour modification of cats through gene editing. The findings could facilitate further understanding of the regulatory role of KIT on feline coat colour and provide a basis for the breeding of cats with commercially desired coat colour.

Classification of superficial suspected lymph nodes: non-invasive radiomic model based on multiphase contrast-enhanced ultrasound for therapeutic options of lymphadenopathy.

Background: Accurate determination of the types of lymphadenopathy is of great importance in disease diagnosis and treatment and is usually confirmed by pathological findings. Radiomics is a non-invasive tool that can extract quantitative information from medical images. Our study was designed to develop a non-invasive radiomic approach based on multiphase contrast-enhanced ultrasound (CEUS) images for the classification of different types of lymphadenopathy. Methods: A total of 426 patients with superficial suspected lymph nodes (LNs) from three centres were grouped into a training cohort (n=190), an internal testing cohort (n=127), and an external testing cohort (n=109). The radiomic features were extracted from the prevascular phase, vascular phase, and postvascular phase of the CEUS images. Model 1 (the conventional feature model), model 2 (the multiphase radiomics model), and model 3 (the combined feature model) were established for lymphadenopathy classification. The area under the curve (AUC) and confusion matrix were used to evaluate the performance of the three models. The usefulness of the models was assessed in different threshold probabilities by decision curve analysis. Results: There were 139 patients (32.6%) with benign LNs, 110 patients (25.8%) with lymphoma, and 177 patients (41.5%) with metastatic LNs in our population. Finally, twenty features were selected to construct the radiomics models for these three types of lymphadenopathy. Model 2 integrating multiphase images of the CEUS yielded the AUCs of 0.838, 0.739, and 0.733 in the training cohort, internal testing cohort, and external testing cohort, respectively. After the combination of conventional features and radiomic features, the AUCs of model 3 improved to 0.943, 0.823 and 0.785 in the training cohort, internal testing cohort, and external testing cohort. Besides, model 3 had an accuracy of 81.05%, sensitivity of 80%, and specificity of 90.43% in the training cohort. Model performance was further confirmed in the internal testing cohort and external testing cohort. Conclusions: We constructed a combined feature model using a series of CEUS images for the classification of the lymphadenopathies. For patients with superficial suspected LNs, this model can help clinicians make a decision on the LN type noninvasively and choose appropriate treatments.

Mussel- and nacre-inspired dual-bionic alginate-based hydrogel coating with multi-matrix applicability, high separation stability and antifouling performance for oil/water separation.

Natural hydrogel-modified porous matrices with superwetting interfaces are ideal for oil/water separation. In this study, inspired by two marine organisms, a novel hydrogel coating with multi-matrix suitability, high oil/water separation capability and antifouling properties was developed. Specifically, inspired by mussel byssus, hydrogel coating was successfully deposited on porous matrix surface based on the introduction of tannic acid (TA). Moreover, inspired by the "brick and mortar" microstructure of Pinctada nacre, silica particles were in-situ synthesized in the sodium alginate (SA)/Ca2+ hydrogel to provide the filling effect and to increase strength. Furthermore, Sodium alginate-tannic acid-tetraethyl orthosilicate (SA-TA-TEOS) hydrogel coating-modified membrane exhibited super-hydrophilic and underwater super-oleophobic performance (underwater oil contact angle >150°), and achieved efficient oil/water separation for four oil/water emulsions (flux = 493-584 L·m-2·h-1 and rejection = 97.3-99.5 %). The modified membrane also demonstrated good anti-fouling performance and flux recovery. Notably, hydrogel coating-modified non-woven fabric also had high oil/water separation capacity (rejection >98 %) and cyclic stability, which proved the universal applicability of this hydrogel coating. In short, this work provides new insights into the fabrication of hydrogel coating-modified porous materials based upon a marine organism biomimetic strategy, which has potential applications in separating oil/water emulsions in industrial scenarios.

The role of m6A methylation in prenatal maternal psychological distress and birth outcome.

BACKGROUND: Prenatal maternal psychological distress (PMPD) is a known risk factor for adverse birth outcomes. N6-methyladenosine RNA (m6A) methylation is crucial in moderating RNA biology. This study aimed to evaluate the relationships between PMPD, birth outcomes, and placental m6A methylation. METHODS: This was a prospective cohort study. PMPD exposure was assessed by questionnaires about prenatal stress, depression, and anxiety. Placental m6A methylation was measured using a colorimetric assay. The relationships between PMPD, m6A methylation, gestational age (GA), and birth weight (BW) were analyzed using structural equation models (SEMs). Maternal weight gain during pregnancy and infant sex were included as covariables. RESULTS: The study included 209 mother-infant dyads. In an adjusted SEM, PMPD was associated with BW (B = -26.034; 95 % CI: -47.123, -4.868) and GA (B = -0.603; 95 % CI: -1.102, -0.154). M6A methylation was associated with PMPD (B = 0.055; 95 % CI: 0.040,0.073) and BW (B = -305.799; 95 % CI: -520.164, -86.460) but not GA. The effect of PMPD on BW was partially mediated by m6A methylation (B = -16.817; 95 % CI: -31.348, -4.638) and GA (B = -12.280; 95 % CI: -23.612, -3.079). Maternal weight gain was associated with BW (B = 5.113; 95 % CI: 0.229,10.438). LIMITATIONS: The study sample size was small, and the specific mechanism of m6A methylation on birth outcomes needs to be further explored. CONCLUSIONS: In this study, PMPD exposure negatively affected BW and GA. Placental m6A methylation was associated with PMPD and BW and partially mediated the effect of PMPD on BW. Our findings highlight the importance of perinatal psychological evaluation and intervention.

Prolactin and glucocorticoid receptors in the prefrontal cortex are associated with anxiety-like behavior in prenatally stressed adolescent offspring rats.

Prenatal stress (PS) causes anxiety in mothers and their offspring and chewing is a commonly observed behavior during maternal stress. Prolactin (PRL) is an anti-anxiety factor that suppresses the hypothalamic-pituitary-adrenal axis. Here, we studied the roles of PRL, corticosterone (CORT), and their receptors in PS-induced anxiety-like behavior in dams and their offspring. We further investigated whether chewing during maternal stress could prevent PS-induced harmful consequences. Pregnant rats were randomly divided into PS, PS + chewing, and control groups. Anxiety-like behaviors of dams and their adolescent offspring were assessed using the open field test and elevated plus maze. Serum levels of PRL and CORT were measured by ELISA. Expression of mRNA and protein of PRLR and glucocorticoid receptor (GR) in the prefrontal cortex (PFC) were evaluated by qRT-PCR and western blotting, respectively. Compared to the control rats, dams and their female offspring, but not male offspring, in the PS group showed increased anxiety-like behaviors. The PS-affected rats had a lower serum PRL level and increased PRLR expression in the PFC. In contrast, these rats had a higher serum CORT level and decreased GR expression in the PFC. Chewing ameliorated anxiety-like behaviors and counteracted stress-induced changes in serum PRL and CORT, as well as the expression of their receptors in the PFC. Conclusion: PS-induced anxiety-like behavior is associated with changes in the serum levels of PRL and CORT and expression of their receptors in the PFC. Moreover, chewing blunts the hormonal and receptor changes and may serve as an effective stress-coping method for preventing PS-induced anxiety-like behavior.

Impact of KIT Editing on Coat Pigmentation and Fresh Meat Color in Yorkshire Pigs.

The white coat color of Yorkshire pigs is caused by the dominant white I allele, which has been associated with at least one copy of the 450-kb duplication encompassing the entire KIT gene and a splice mutation (G > A) at the first base of intron 17. The splice mutation in KIT has an adverse effect on pigmentation in mice. Therefore, removing the 450 kb duplications harboring the KIT copy with splice mutations is expected to affect Yorkshire pig pigmentation. In this study, we describe the use of a Yorkshire pig kidney cell strain with the I?/IBe-ed genotype, previously created by CRISPR-Cas9, as donor cells for somatic cell nuclear transfer to generate gene-edited Yorkshire pigs. The removal of the 450 kb duplications harboring the KIT copy with splice mutation did not alter the white coat color of Yorkshire pigs, which was confirmed by the absence of fully mature melanocytes and melanin accumulation in the hair follicles. Except for the improved transcription of tyrosinase, and slight increase in microphthalmia transcription factor and tyrosinase-related protein 1 protein expression, there was no significant impact of the removal of splice mutations on genes and signaling pathways (PI3K/AKT) involved in melanogenesis. However, the removal of the 450 kb duplications harboring the KIT copy with splice mutation substantially improved fresh meat color accompanied by significantly increased red blood cell number, which merits further investigation. Our study provides new insights into the role of structural mutations of the KIT gene in the formation of white coat color and erythropoiesis in Yorkshire pigs.

Engineering a superwetting membrane with spider-web structured carboxymethyl cellulose gel layer for efficient oil-water separation based on biomimetic concept.

Superhydrophilic and underwater superoleophobic membranes have recently attracted significant interest as materials for effective oil-water emulsion separation. In this work, a superwetting membrane with a spider web structured gel layer was designed for efficient oil-water separation. Biomaterial, carboxymethyl cellulose (CMC), was used as the raw material, a spider web structured gel layer was constructed on the PVDF membrane surface by heat-treatment and chemical cross-linking. The hydrophilic gel layer imparted excellent superhydrophilic and underwater superoleophobic properties to the membrane, while the special spider web structure improved the membrane mechanical stability. The fabricated membrane exhibited superhydrophilicity and underwater superoleophobicity. Among different CMC concentration-modified membranes, the M-0.5 membrane containing 0.5 wt% CMC exhibited a flux of 612 L·m-2 h-1 during dichloromethane oil-water emulsion separation, which was 4.2-fold higher than that of the pristine PVDF membrane, while the membrane showed efficient oil-water separation capacity. Additionally, the water flux recovery reached as high as 93.3 %, and oil rejection attained 99.1 %. Meanwhile, the spiderweb-structured gel layer on the membrane surface displayed good mechanical stability. In summary, this novel membrane-modification method, inspired by the spider web structure, was simple, cost effective and environmentally friendly, thereby making it promising for future preparation of highly efficient oil-water emulsion separation membranes.

Independent risk factors and feasibility of ultrasound diagnosis of ultrasound-guided non-cytologically diagnostic thyroid nodules.

BACKGROUND: To explore the independent risk factors and feasibility of ultrasound diagnosis of ultrasound-guided non-cytologically diagnostic thyroid nodules. METHODS: This study included 200 patients with thyroid nodules that were diagnosed or suspected of being Thyroid Imaging Reporting and Data System (TI-RADS) 4 nodules between January 2017 and January 2019. All patients received surgical treatment and pathological diagnosis, and were divided into a cytologically diagnostic group and a non-cytologically diagnostic group based on whether they could be diagnosed by ultrasound-guided fine needle aspiration cytology (UG-FNAC). Patients were further divided into benign and malignant groups according to the results of surgical pathology. Logistic regression analysis was used to clarify the risk factors that could not be cytologically diagnosed in TI-RADS 4 nodules. For the diagnosis of benign and malignant nodules, we combined contrast-enhanced ultrasound (CEUS) and ultrasound elastography (UE) to establish a joint scoring strategy. The diagnostic value of the joint scoring strategy was evaluated by receiver operating characteristic (ROC) curve. RESULTS: A total of 216 TI-RADS type 4 nodules were detected in 200 patients. Among them, 40 nodules were included in the non-cytologically diagnostic group, and 176 nodules were included in the cytologically diagnostic group. The multi-parameter logistic regression showed that: aspect ratio <1, irregular edge, scattered coarse calcification, middle layer, and lower layer were independent influencing factors leading to undiagnosed puncture cytology. Among the 216 nodules in 200 patients, 168 were pathologically diagnosed as malignant nodules (malignant group), and 48 nodules were diagnosed as benign nodules (benign group). According to the joint scoring strategy, the distribution of comprehensive scores changes significantly at 5-6 scores while in malignant nodules, the distribution of comprehensive scores changes significantly at 6-7 scores. ROC curve analysis showed that the diagnostic value was the highest when '6 scores' was used as the critical point for diagnosis [area under curve (AUC) =0.893, P<0.05]. CONCLUSIONS: The combined scoring strategy of CEUS and UE was an effective method to diagnose TI-RADS4 nodules could not be diagnosed by UG-FNAC.

Modern climate and vegetation variability recorded in organic compounds and carbon isotopic compositions in the Dianchi watershed.

The aliphatic hydrocarbons distribution and compound-specific characteristics of carbon isotopic compositions in the sediments from the small catchment (197 km(2)) of the Dianchi watershed were investigated for identification of modern climate and vegetation variations in the study area. Results show that a regular bimodal n-alkanes distribution ranged from nC16 to nC33, with strong dominance at nC17 for short-chain n-alkanes and nC31 for long-chain n-alkanes. Mass chromatogram of total fatty acids also indicates corresponding mixed contribution of algae, hydrophilous non-emergent (C4 plants) and terrestrial plants (C3 plants) to sedimentary organic matter (OM). At the depth of -24 to -25 cm (early 1970s), nC31/nC17 and terrestrial to aquatic ratio of hydrocarbons (TAR) values decrease, suggesting a shift of OM origins from C3 terrestrial plants to algae-derived C4 plants. The highest water stage in 1971 was found to be recorded in the particle size (<4 µm). For long-chain alkanes, the values of δ (13)Corg and δ (13)Cn-alkanes varied from -26.9 to -22.4 and -33.4 to -27.9 ‰, respectively. Population growth and economic development led to a demand for abundant habitable and cultivable land. Due to unreasonable land expansion, the primordial forest sporadically distributed. A mixture of C3 and C4 plants probably replaced C3 plants as the sources of OM in the past 10 years. The changes of land-use types and severe drought resulted in the excessive OM inputs to the watershed.

Biomarker and stable carbon isotopic signatures for 100-200 year sediment record in the Chaihe catchment in southwest China.

Natural inputs and anthropogenic influences on lakes and their catchments are reflected in the sediment record. In the present study, the extractable organic compounds from sediments in the Chaihe catchment of the Dianchi watershed were analyzed to characterize source inputs. Results show that the sediments are dominated by odd numbered n-alkanes (n-C16-n-C33), maximizing at n-C17, n-C29 and n-C31. Aliphatic hydrocarbon may be composed of terrestrial plants and bacteria. The values of δ(13)C27, δ(13)C29 and δ(13)C31 of n-alkanes exhibit a range from -33.27‰ to -25.46‰, from -35.76‰ to -28.47‰ and from -33.67‰ to -27.42‰, respectively and three records strongly covary with depth, falling within the range of C3 plants in the study area. An isotopic model revealed C3 plant contribution to sedimentary organic matter (OM) ranging from 40.75% to 97.22%. The values of ACL27-33, CPI27-33, OEP, Paq, Pr/Ph, (C27+C29)/2C31, (C21+C23+C25)/3C17 and nC26(-)/nC27(+) are consistent with the C3 plant predominance. A constant CRS model gave the accumulation rates ranging from 2.69 to 8.46mma(-1) spanning 1885-2010. It was concluded that OM transport in the Chaihe catchment was influenced strongly by human activities resulting in enhanced eutrophication.

Effects of plant species on soil microbial processes and CH4 emission from constructed wetlands.

Methane (CH(4)) emission from constructed wetland has raised environmental concern. This study evaluated the influence of mono and polyculture constructed wetland and seasonal variation on CH(4) fluxes. Methane emission data showed large temporal variation ranging from 0 to 249.29 mg CH(4) m(-2) h(-1). Results indicated that the highest CH(4) flux was obtained in the polyculture system, planted with Phragmites australis, Zizania latifolia and Typha latifolia, reflecting polyculture system could stimulate CH(4) emission. FISH analysis showed the higher amount of methanotrophs in the profile of Z. latifolia in both mono and polyculture systems. The highest methanogens amount and relatively lower methanotrophs amount in the profile of polyculture system were obtained. The results support the characteristics of CH(4) fluxes. The polyculture constructed wetland has the higher potential of global warming.