Integration of the Ligase Chain Reaction with the CRISPR-Cas12a System for Homogeneous, Ultrasensitive, and Visual Detection of microRNA.

The ligase chain reaction (LCR), as a classic nucleic acid amplification technique, is popular in the detection of DNA and RNA due to its simplicity, powerfulness, and high specificity. However, homogeneous and ultrasensitive LCR detection is still quite challenging. Herein, we integrate the LCR with a CRISPR-Cas12a system to greatly promote the application of the LCR in a homogeneous fashion. By employing microRNA as the model target, we design LCR probes with specific protospacer adjacent motif sequences and the guide RNA. Then, the LCR is initiated by target microRNA, and the LCR products specifically bind to the guide RNA to activate the Cas12a system, triggering secondary signal amplification to achieve ultrasensitive detection of microRNA without separation steps. Moreover, by virtue of a cationic conjugated polymer, microRNA can not only be visually detected by naked eyes but also be accurately quantified based on RGB ratio analysis of images with no need of sophisticated instruments. The method can quantify microRNA up to 4 orders of magnitude, and the determination limit is 0.4 aM, which is better than those of other reported studies using CRISPR-Cas12a and can be compared with that of the reverse-transcription polymerase chain reaction. This study demonstrates that the CRISPR-Cas12a system can greatly expand the application of the LCR for the homogeneous, ultrasensitive, and visual detection of microRNA, showing great potential in efficient nucleic acid detection and in vitro diagnosis.

Identification of a hippocampal lncRNA-regulating network in a natural aging rat model.

BACKGROUND: Dysregulation of long noncoding RNA (lncRNA) expression is related to aging and age-associated neurodegenerative diseases, and the lncRNA expression profile in the aging hippocampus is not well characterized. In the present investigation, the changed mRNAs and lncRNAs were confirmed via deep RNA sequencing. GO and KEGG pathway analyses were conducted to investigate the principal roles of the clearly dysregulated mRNAs and lncRNAs. Subsequently, through the prediction of miRNAs via which mRNAs and lncRNAs bind together, a competitive endogenous RNA network was constructed. RESULTS: A total of 447 lncRNAs and 182 mRNAs were upregulated, and 385 lncRNAs and 144 mRNAs were downregulated. Real-time reverse transcription-polymerase chain reaction validated the reliability of mRNA and lncRNA sequencing. KEGG pathway and GO analyses revealed that differentially expressed (DE) mRNAs were associated with cell adhesion molecules (CAMs), the p53 signaling pathway (SP), phagosomes, PPAR SP and ECM-receptor interactions. KEGG pathway and GO analyses showed that the target genes of the DE lncRNAs were related to cellular senescence, the p53 signaling pathway, leukocyte transendothelial migration and tyrosine metabolism. Coexpression analyses showed that 561 DE lncRNAs were associated with DE mRNAs. A total of 58 lncRNA-miRNA-mRNA target pairs were confirmed in this lncRNA‒miRNA‒mRNA network, comprising 10 mRNAs, 13 miRNAs and 38 lncRNAs. CONCLUSIONS: We found specific lncRNAs and mRNAs in the hippocampus of natural aging model rats, as well as abnormal regulatory ceRNA networks. Our outcomes help explain the pathogenesis of brain aging and provide direction for further research.

[Mechanism of ferroptosis in regulating testicular spermatogenic dysfunction: Progress in research].

Ferroptosis is a new type of cell death found in recent years, which is characterized by iron overload, excessive accumulation of lipid peroxide and destruction of mitochondrial structure. Testicular spermatogenic dysfunction (TSD) is an important factor affecting the health and quality of life of middle-aged and elderly men. Studies show that ferroptosis plays an important role in the development and progression of TSD, mainly involved in the mechanisms of abnormal iron metabolism, lipid peroxidation, oxidative stress response and mitochondrial damage. It is believed that ferroptosis can induce a series of symptoms, such as sperm quality and quantity reduction and spermatogenesis disorder. Therefore, inhibiting the development of ferroptosis can provide a new basis for the study of the mechanisms of the disease. This review focuses on the concept and characteristics of ferroptosis and the advances in the studies of the its mechanism in regulating TSD, with the purpose of introducing the clinical application prospect of targeted inhibition of ferroptosis in the prevention and treatment of TSD and expanding the ideas for the treatment of TSD and other male reproductive diseases.

Transcriptome Profiles of the Liver in Two Cold-Exposed Sheep Breeds Revealed Different Mechanisms and Candidate Genes for Thermogenesis.

Cold-induced thermogenesis plays an important role in the survival of lambs exposed to low air temperatures. The liver produces and mediates heat production in mammals; however, to date, little is known about the role of liver genes in cold-induced thermogenesis in lambs. In this study, the difference in the liver transcriptome between Altay and Hu ewe lambs was compared. Because of different backgrounds of the two breeds, we hypothesized that the transcriptome profiles of the liver would differ between breeds when exposed to cold. Cold-exposed Altay lambs activated 8 candidate genes (ACTA1, MYH1, MYH2, MYL1, MYL2, TNNC1, TNNC2, and TNNT3) involved in muscle shivering thermogenesis; 3 candidate genes (ATP2A1, SLN, and CKM) involved in muscle nonshivering thermogenesis related to the Ca2+ signal and creatine cycle; and 6 candidate genes (PFKM, ALDOC, PGAM2, ENO2, ENO3, and ENO4) involved in enhancing liver metabolism. In contrast, the liver may not act as the main tissue for thermogenesis in cold-exposed Hu lambs. We concluded that Altay lambs rely on liver-mediated shivering and nonshivering thermogenesis by muscle tissue to a greater extent than Hu lambs. Results from this study could provide a theoretical foundation for the breeding and production of cold-resistant lambs.

Ischemia-Modified Albumin Is Associated with Arterial Stiffness in Hemodialysis Patients.

Increased arterial stiffness is strongly associated with cardiovascular morbidity and mortality in dialysis patients. Ischemia-modified albumin (IMA) is a useful biomarker of cardiac ischemia. This study was aimed to explore the association between IMA and arterial stiffness in hemodialysis patients. An observational study was conducted with 120 hemodialysis patients. Clinical data and laboratory characteristics were collected. Arterial stiffness was evaluated by brachial-ankle pulse wave velocity (baPWV). Hemodialysis patients had extensive arterial stiffness and high levels of IMA. Comparing to hemodialysis patients with normal baPWV, those with high baPWV had significantly higher levels of IMA (93.7 ± 8.6 versus 73.1 ± 10.7 Ku/L, P = 0.027). The multiple linear regression analysis showed that IMA was significantly associated with arterial stiffness in hemodialysis patients (ß = 0.43, P < 0.001). Moreover, IMA, with a threshold value of 90.4 Ku/L, provided 77.4% sensitivity and 86.6% specificity for predicting arterial stiffness. Hemodialysis patients with arterial stiffness had high levels of IMA. IMA was a good predictive marker of arterial stiffness for hemodialysis patients.

Effect of air temperature on growth performance, apparent digestibilities, rumen fermentation and serum metabolites in Altay and Hu lambs.

Fat-tailed Altay sheep are indigenous to the Altay prefecture and well adapted to severe cold and sparse pasture of poor quality. Hu sheep were introduced to this region in the 1970s and are raised mainly in feedlots. We hypothesized that the dietary energy utilization would differ between breeds and predicted that Altay sheep would be more efficient than Hu sheep, in particular at a low air temperature. To test this prediction, we examined growth performance, apparent digestibilities, rumen fermentation and serum metabolites in Altay (32 ± 2.6 kg) and Hu sheep (31 ± 2.7 kg) at air temperatures of -5 and 20°C. Average daily gain (ADG), feed and metabolizable energy intakes were greater in Altay than in Hu sheep (p < .01), and all were greater at -5°C than 20°C (p < .01). Body mass gain-to-feed intake ratio was higher in Altay than in Hu sheep (p < .001), but was not affected by air temperature (p > .10). Rumen total volatile fatty acid (VFA) concentration was greater in Hu than in Altay sheep (p < .05) and was greater at 20°C than at -5°C (p < .05), while rumen microbial protein concentration was greater in Altay than in Hu sheep (p < .05). Rectal temperature was higher at -5°C than 20°C (p < .05) and was similar between breeds (p > .05). Serum glucose, non-esterified fatty acid (NEFA) and urea-N concentrations were higher in Hu than in Altay sheep (p < .05). It was concluded that Altay sheep are better able to cope with poor quality diets as they had higher digestibility of nutrients, ADG and body mass gain-to-feed intake ratio than Hu sheep.

miR-375-3p/YWHAZ/ß-catenin axis regulates migration, invasion, EMT in gastric cancer cells.

YWHAZ (14-3-3ζ) plays crucial roles in regulating proliferation, apoptosis, migration, and invasion of gastric cancer (GC) cells. However, its extensive roles and potential mechanisms in GC cells remain unknown, and need to be researched deeply. In this study, we focus on the role of miR-375/YWHAZ axis in migration, invasion and epithelial-to-mesenchymal transition (EMT) of GC cells. YWHAZ level was assessed by western blot and qPCR assays in GC cells. Scratch and transwell assays were used to determine the migration and invasion of GC cells. The protein levels of correlative molecules were detected by western blot. The regulation of miR-375 on the expression of its target gene YWHAZ was verified by dual-luciferase report system. According to the results, knockdown of YWHAZ inhibited the migration, invasion and EMT of GC cells. Moreover, silencing of YWHAZ restrained the activation of wnt/ß-catenin signalling pathway. YWHAZ was confirmed to be a target gene of miR-375, and its expression was regulated by miR-375 in GC cells. Transfection of miR-375 inhibitor promoted the migration, invasion, EMT and activation of wnt/ß-catenin pathway in GC cells, which was suppressed by inhibition of YWHAZ. Taken together, this study suggests that miR-375/YWHAZ axis may be served as a novel therapeutic target for GC patients.

Molecular characterization and expressing analysis of the c-type and g-type lysozymes in Qihe crucian carp Carassius auratus.

Lysozyme as an important nonspecific immune factor, can kill bacteria by hydrolyzing ß-1,4-glycosidic linkages of peptidoglycan layer, and plays an important role in innate immune response against pathogen infection. In the present study, we report molecular cloning, tissue distribution and functional characterization of the c-type and g-type lysozymes in Qihe crucian carp Carassius auratus (designated as Ca-clys and Ca-glys, respectively). The full-length of Ca-clys and Ca-glys cDNA were cloned using RT-PCR and RACE methods. Catalytic and other conserved residues, required for functionality, were identified by multiple sequence alignment and structure predicted. The findings indicating the Ca-clys with signal peptide sequence, while the Ca-glys without, imply that the two isozymes function in different sites of cell. Phylogenetic analysis revealed that Ca-clys and Ca-glys genes evolve at different rates. Moreover, spatial expression analysis showed that Ca-clys transcript was most abundant in kidney and least in gill. However, the expression level of Ca-glys was significantly lower compared with Ca-clys in various tissues, which was the most abundant in spleen and least in brain. After intraperitoneal injection with A. hydrophila and lipopolysaccharide (LPS), the mRNA levels of Ca-clys and Ca-glys were generally up-regulated in liver and gill, but indicated the different expression changes in spleen, kidney and head kidney. With regard to the lysozyme activity, it was showed that the total enzyme activities generally increased in liver, gill, spleen, and head kidney after stimulation. These results confirmed that both Ca-clys and Ca-glys play an important role in non-specific immunity after A. hydrophila invasion. In this study, it was speculated that expressions of Ca-clys and Ca-glys were regulated in different patterns against pathogens.

Rickettsia raoultii, the predominant Rickettsia found in Dermacentor silvarum ticks in China-Russia border areas.

Since the year 2000, clinical patterns resembling tick-borne rickettsioses have been noticed in China-Russia border areas. Epidemiological data regarding species of the aetiological agent, tick vector prevalence and distribution as well as incidence of human cases in the areas are still sparse to date. In order to identify Rickettsia species occurring in the areas, we investigated Dermacentor silvarum collected in the selected areas. Rickettsia raoultii was the predominant Rickettsia found in D. silvarum evident with ompA, ompB, gltA and 17 kDa protein genes. The Rickettsia prevalence in D. silvarum appeared to be 32.25 % with no sex difference. The results extend the common knowledge about the geographic distribution of R. raoultii and its candidate vector tick species, which suggest an emerged potential threat of human health in the areas.

Pedestrian walking speed monitoring at street scale by an in-flight drone.

The walking speed of pedestrians is not only a reflection of one's physiological condition and health status but also a key parameter in the evaluation of the service level of urban facilities and traffic engineering applications, which is important for urban design and planning. Currently, the three main ways to obtain walking speed are based on trails, wearable devices, and images. The first two cannot be popularized in larger open areas, while the image-based approach requires multiple cameras to cooperate in order to extract the walking speed of an entire street, which is costly. In this study, a method for extracting the pedestrian walking speed at a street scale from in-flight drone video is proposed. Pedestrians are detected and tracked by You Only Look Once version 5 (YOLOv5) and Simple Online and Realtime Tracking with a Deep Association Metric (DeepSORT) algorithms in the video taken from a flying unmanned aerial vehicle (UAV). The distance that pedestrians traveled related to the ground per fixed time interval is calculated using a combined algorithm of Scale-Invariant Feature Transform (SIFT) and random sample consensus (RANSAC) followed by a geometric correction algorithm. Compared to ground truth values, it shows that 90.5% of the corrected walking speed predictions have an absolute error of less than 0.1 m/s. Overall, the method we have proposed is accurate and feasible. A particular advantage of this method is the ability to accurately predict the walking speed of pedestrians without keeping the flight speed of the UAV constant, facilitating accurate measurements by non-specialist technicians. In addition, because of the unrestricted flight range of the UAV, the method can be applied to the entire scale of the street, which assists in a better understanding of how the settings and layouts of urban affect people's behavior.

[Retracted] MicroRNA­663 inhibits the proliferation and invasion of clear cell renal cell carcinoma cells by directly targeting PAK4.

Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that certain of the data shown for the Transwell cell migration and invasion assays in Figs. 2C, 5D and 6D were strikingly similar to data appearing in different form in other articles by different authors, several of which have been retracted. Owing to the fact that the contentious data in the above article had already been published elsewhere, or were already under consideration for publication, prior to its submission to Molecular Medicine Reports, the Editor has decided that this paper should be retracted from the Journal. After having been in contact with the authors, they agreed with the decision to retract the paper. The Editor apologizes to the readership for any inconvenience caused. [Molecular Medicine Reports 19: 711­718, 2019; DOI: 10.3892/mmr.2018.9652].

Observation of nonvanishing optical helicity in thermal radiation from symmetry-broken metasurfaces.

Spinning thermal radiation is a unique phenomenon observed in condensed astronomical objects, including the Wolf-Rayet star EZ-CMa and the red degenerate star G99-47, due to the existence of strong magnetic fields. Here, by designing symmetry-broken metasurfaces, we demonstrate that spinning thermal radiation with a nonvanishing optical helicity can be realized even without applying a magnetic field. We design nonvanishing optical helicity by engineering a dispersionless band that emits omnidirectional spinning thermal radiation, where our design reaches 39% of the fundamental limit. Our results firmly suggest that metasurfaces can impart spin coherence in the incoherent radiation excited by thermal fluctuations. The symmetry-based design strategy also provides a general pathway for controlling thermal radiation in its temporal and spin coherence.

Multifunctional Phase-Transition Nanoparticles for Effective Targeted Sonodynamic-Gene Therapy Against Thyroid Papillary Carcinoma.

Introduction: In order to diagnose and treat papillary thyroid carcinoma (PTC) accurately, phase-transition nanoparticles, P@IP-miRNA (PFP@IR780/PLGA-bPEI-miRNA338-3p), was engineered. The nanoparticles (NPs) can target the tumor cells, realize the multimodal imaging, and provide sonodynamic-gene therapy for PTC. Methods: P@IP-miRNA NPs were synthesized through double emulsification method, and miRNA338-3p was attached to the surface of the NPs by electrostatic adsorption. The characterization of NPs was detected to screen out qualified nanoparticles. In vitro, laser confocal microscopy and flow cytometry were used to detect the targeting and subcellular localization of NPs. Western blot, qRT-PCR, and immunofluorescence were used to detect the ability to transfect miRNA. CCK8 kit, laser confocal microscopy and flow cytometry were used to detect the inhibition on TPC-1 cells. In vivo experiments were performed based on tumor-bearing nude mice. The efficacy of combined treatment by NPs was comprehensively evaluated, and the multimodal imaging ability of NPs in vivo and in vitro was detected. Results: P@IP-miRNA NPs were successfully synthesized which have spherical shape, uniform size, good dispersion and positive potential. The encapsulation rate of IR780 was (82.58±3.92) %, the drug loading rate was (6.60±0.32) %, and the adsorption capacity of miRNA338-3p was 41.78 µg/mg. NPs have excellent tumor targeting ability, miRNA transfection ability, ROS production ability and multimodal imaging ability in vivo and in vitro. The antitumor effect of combined treatment group was the best, and the efficacy was better than that of single factor treatment group, and the difference was statistically significant. Conclusion: P@IP-miRNA NPs can realize multimodal imaging and sonodynamic-gene therapy, providing a new idea for accurate diagnosis and treatment of PTC.

Guanidinoacetic Acid Attenuates Adipogenesis through Regulation of miR-133a in Sheep.

Guanidinoacetic acid (GAA) is an amino acid derivative, previously described in the skeletal muscle of vertebrates, that serves as an important regulator of cellular bioenergetics and has been widely used as a feed additive. Nevertheless, the effect of GAA on adipose tissue growth remains unclear. Here, we hypothesized that dietary GAA negatively affected adipose tissue development in lambs. Lambs were individually fed diets with (0.09%) or without GAA for 70 d ad libitum, and the subcutaneous adipose tissues were sampled for analysis. The results showed that dietary GAA supplementation decreased the girth rib (GR) value (p < 0.01) of lamb carcasses. Both real-time PCR and Western blot analysis suggested that dietary GAA inhibited the expression of adipogenic markers, including peroxisome proliferator-activated receptor γ (PPARγ, p < 0.05), CCAAT/enhancer-binding protein α (C/EBPα, p < 0.01) and sterol-regulatory-element-binding protein 1c (SREBP1C, p < 0.01) in subcutaneous adipose tissue. In vitro, GAA inhibited sheep stromal vascular fraction (SVF) cell proliferation, which was associated with downregulation of proliferating cell nuclear antigen (PCNA, p < 0.05), cyclin-dependent kinase 4 (CDK 4, p < 0.05) and cyclin D1 (p < 0.01). GAA suppressed adipogenesis of SVF cells. Furthermore, miRNA sequencing revealed that GAA affected the miRNA expression profile, and real-time PCR analysis confirmed that miR-133a expression in both subcutaneous adipose tissue and SVF cell was downregulated by GAA. Meanwhile, miR-133a promoted adipogenic differentiation of SVF cells by targeting Sirt1. miR-133a mimics alleviated the inhibitory effect of GAA on SVF cells' adipogenic differentiation. In summary, GAA attenuated adipogenesis of sheep SVF cells, which might occur through miR-133a-modulated Sirt1 expression.

Effect of dietary Agriophyllum squarrosum on average daily gain, meat quality and muscle fatty acids in growing Tan lambs.

The herb A. squarrosum is reputed to possess medicinal properties for humans, and has the potential to be a feed resource for livestock. We hypothesized that this herb would improve the meat quality of lambs. To test this hypothesis, 24 Tan ewe-lambs (27.7 ± 0.45 kg) were offered diets containing 0 (CON), 100 (AS100), 200 (AS200) and 300 (AS300) g A. squarrosum/kg DM, and average daily gain, carcass traits, blood metabolites, meat quality and fatty acid profiles were determined. Drip loss % and cooking loss % decreased with the AS100 and AS200 diets (P < 0.05). Dietary A. squarrosum reduced muscle fiber area and diameter and increased density of the meat (P < 0.05), which indicated that the meat was more tender. The concentrations of C10:0 and C18:1n-9 t were 1ower and of C17:0 and C18:3n-3 were greater in the AS200 and AS300 treatments than CON (P < 0.05). Our results suggest that feeding lambs up to 200 g/kg DM of A. squarrosum can increase the water-holding capacity and L* value of meat without compromising growth. Further research is needed to determine the optimal level.

[Establishment of a Mouse Model of Acquired Aplastic Anemia Mediated by Cyclophosphamide Combined with Cyclosporine].

OBJECTIVE: To establish a stable mouse model of acquired aplastic anemia. METHODS: Female BALB/C mice aged 6 months were intraperitoneally injected with cyclophosphamide and cyclosporine for 14 days. The number of peripheral blood cells, the concentration of hemoglobin, the number of bone marrow nucleated cells, bone marrow smear, bone marrow pathological sections and other indexes were observed. RESULTS: In BALB/C mice injected intraperitoneally with cyclophosphamide and cyclosporine, the number of peripheral blood cells and the concentration of hemoglobin were significantly decreased, especially the white blood cells and platelets. Bone marrow smear showed a significant decrease in the number of nucleated cells and bone marrow hyperplasia. Bone marrow pathology showed decreased hematopoietic cells and increased non-hematopoietic cells such as adipocytes. CONCLUSION: The mouse model with intraperitoneal injection of cyclophosphamide and cyclosporine can meet the diagnostic criteria of acquired aplastic anemia, which can be used as a mouse model for the study of the pathogenesis and treatment of acquired aplastic anemia.

Transcriptome analysis revealed potential genes involved in thermogenesis in muscle tissue in cold-exposed lambs.

Cold tolerance is an important trait for sheep raised at high altitudes. Muscle tissue, comprising 30-40% of the total body mass, produces heat during cold exposure. However, little is known about the genetic mechanisms of this tissue and its role in thermogenesis in lambs. We examined genes in skeletal muscle tissue in a cold-adapted sheep breed, Altay, and a cold-intolerant sheep breed, Hu, when exposed to low air temperature. Three ewe-lambs of each breed were maintained at -5°C and three ewe-lambs of each breed were maintained at 20°C. After cold exposure for 25 days, the longissimus dorsi of each lamb was collected, and transcriptome profiles were sequenced and analyzed. The results of RNA-seq showed that the average reads among the four groups were 11.0 Gbase. The genome mapping rate averaged 88.1% and the gene mapping rate averaged 82.5%. The analysis of differentially expressed genes (DEGs) indicated that the peroxisome proliferator-activated receptors (PPAR), cAMP, and calcium signaling pathways and muscle contraction in muscle tissue were linked to thermogenesis in cold-exposed lambs. Furthermore, PCK1 (phosphoenolpyruvate carboxykinase1) increased glyceroneogenesis in cold-exposed Altay lambs, and APOC3 (apolipoprotein C3), LPL (lipoprotein lipase), and FABP4 (fatty acid binding protein 4, adipocyte) were involved in the intake and transport of free fatty acids. In Hu sheep, cAMP biosynthesis from ATP hydrolysis was regulated by ADCY10 (adenylate cyclase) and ADORA2a (adenosine A2a receptor). Skeletal muscle contraction was regulated by MYL2 (myosin light chain 2). In conclusion, cold exposure altered the expression level of genes involved in heat production in muscle tissue. Some potential mechanisms were revealed, including calcium ion transport in the calcium signaling pathway, fatty acid metabolism in the PPAR signaling pathway, and cAMP biosynthesis in the cAMP signaling pathway. This study implied that skeletal muscle plays an important role in thermoregulation in lambs.

Dietary Supplementation of Fruit from Nitraria tangutorum Improved Immunity and Abundance of Beneficial Ruminal Bacteria in Hu Sheep.

The fruit of Nitraria tangutorum (FNT) is reputed to possess medicinal properties; however, its effect on sheep (Ovis aries) is unknown. The aim of this study was to fill this gap. In a 3 × 3 Latin square design, six 12-month-old rumen-fistulated Hu rams (56.2 ± 8.26 kg; mean ± SD) were penned individually and offered one of three levels of FNT, namely, 0 g/d (control; CON), 16 g/d (N16), and 48 g/d (N48). The concentration of serum immunoglobulin G increased linearly (p = 0.03) with an increasing intake of FNT. The serum concentration of ß-hydroxybutyrate in the N48 group was lower than in the CON group (p = 0.01) and decreased linearly with increasing FNT (p = 0.001). The concentration of serum lactate dehydrogenase tended to decrease (p = 0.07) linearly with an increase in FNT intake, while the concentration of glucose did not differ among groups (p = 0.14) but displayed a quadratic curve with an increase in FNT (p = 0.05). The rumen concentration of lipase decreased linearly with increasing FNT (p = 0.04). The rumen fermentation variables were not affected by FNT. The FNT intake increased the abundance of beneficial ruminal bacteria, such as Lachnoclostridium, Rhodocyclaceae, and Candidatus Arthromitus. Prevotella, Rikenellaceae_RC9_gut_group, Ruminococcus, Olsenella, Lachnospiraceae_NK3A20_group, and Quinella were the dominant bacterial genera in all treatments. We conclude that FNT can improve immunity and increase the relative abundance of beneficial ruminal bacteria in sheep.

Supplementing Diets with Agriophyllum squarrosum Reduced Blood Lipids, Enhanced Immunity and Anti-Inflammatory Capacities, and Mediated Lipid Metabolism in Tan Lambs.

Agriophyllum squarrosum (sand rice), a widespread desert plant, possesses anti-hyperglycemic and anti-inflammatory properties, and has been used in traditional Chinese medicine for many years. However, its effects on ruminants are unknown. To fill this gap, we examined the effects of A. squarrosum on the immune and anti-inflammatory responses of lambs. A total of 23, 6-month-old Tan ewe-lambs (27.6 ± 0.47 kg) were divided into four groups and offered a basic diet (C­control), or a diet that contained 10%, 20%, or 30% A. squarrosum, on a dry matter basis, for 128 days. Serum concentrations of total cholesterol were lower (p = 0.004) in the 30% supplemented lambs than controls, while concentrations of high-density lipoprotein cholesterol were lower (p = 0.006) in the 10% and 20%, but not in 30% supplemented lambs than controls. Serum-cortisol concentrations were lower (p = 0.012) in the 30% supplemented lambs and free fatty acid concentrations were higher in the 10% and 20% supplemented lambs than in control lambs (p < 0.001). Supplementation with A. squarrosum decreased (p < 0.05) the area of adipocytes in subcutaneous adipose tissue, but there was no difference between the 20% and 30% diets. Conversely, the area in visceral adipose tissue (VAT) increased (p < 0.05), especially for the 10% and 20% supplemented diets. Supplementation with A. squarrosum also enriched immune and anti-inflammatory related and lipid and glucose-metabolic pathways and associated differentially expressed gene expressions in adipose tissue. A total of 10 differential triacylglycerol, 34 differential phosphatidylcholines and seven differential phosphatidylethanolamines decreased in the diet with 30% supplementation, when compared to the other diets. Finally, adipocyte-differentiation genes, and immune and inflammatory response-related gene expression levels decreased in lamb adipocytes cultured with an aqueous A. squarrosum extract. In conclusion, supplementing lamb diets with A. squarrosum reduced blood lipids, enhanced immunity and anti-inflammatory capacities, and mediated lipid metabolism in adipose tissue and adipocytes of Tan lambs. A level of approximately 10% is recommended, but further research is required to determine the precise optimal level.

Subtyping of Internalizing and Externalizing Behaviors in Japanese Community-Based Children: A Latent Class Analysis and Association with Family Activities.

Externalizing and internalizing behavioral problems occurs at a high rate among children. However, this has rarely been examined among Japanese children using a person-oriented method. Hence, this study aims to explore its subtypes and clarify their association with family-based group activities. We conducted a cross-sectional survey in a typical community-based suburban area for all families with primary school children in Japan. We investigated children's internalizing and externalizing behaviors based on the Japanese version of the Strengths and Difficulties Questionnaire (SDQ), and different types of activities that family members frequently engaged in. Data from 206 families were collected and used for the analysis. The subtypes were explored using latent class analysis (LCA). The relationship between family activities and latent class membership was analyzed using a logistic regression model. Moreover, three latent class models and their probabilities were identified, namely, risk group (31.3%), moderate group (44.9%), and normal group (23.8%). Frequent family activities including play sports, traveling or hiking, watching TV and communicating, cooking or making a dessert, and doing housework, which were significantly related to the normal group. These results would add evidence to potential types of children's behavioral problems and preventive childcare practices needed in the primary gate of families.