STAT transcription factor is indispensable for oogenesis in silkworm.

Signal Transducer and Activator of Transcription (STAT) proteins represent a critical transcription factor family with multifaceted roles in diverse fundamental eukaryotic processes. In Drosophila, STAT exerts a pivotal regulatory influence on oogenesis, governing the early differentiation of follicular cells and ensuring proper encapsulation of germ-line cells. However, the role of STAT in egg development in silkworms remains unknown. In the present study, using CRISPR/Cas9 technology, we successfully generated a strain of silkworms with targeted deletion of the STAT-L gene, which resulted in significant reproductive abnormalities observed in female moths, including shortened fallopian tubes and reduced egg production. The ovaries dissected from STAT-L knockout silkworms during the pupal stage of silkworm exhibited varying degrees of fusion among egg chambers. Additionally, paraffin sections of prepupal ovaries also revealed evidence of egg chambers fusion. To elucidate the molecular mechanism underlying the role of the STAT-L gene regulation on egg development in silkworm, we performed ovarian transcriptomic analysis following STAT-L knockout. Our findings indicated that STAT-L gene can modulate Notch signaling pathway by down-regulating APH-1 gene expression. These results suggest that STAT-L gene plays a crucial role in normal egg chamber formation in silkworms, potentially through its influence on Notch signaling pathway expression.

Dissociation protocols influence the phenotypes of lymphocyte and myeloid cell populations isolated from the neonatal lymph node.

Frequencies and phenotypes of immune cells differ between neonates and adults in association with age-specific immune responses. Lymph nodes (LN) are critical tissue sites to quantify and define these differences. Advances in flow cytometry have enabled more multifaceted measurements of complex immune responses. Tissue processing can affect the immune cells under investigation that influence key findings. To understand the impact on immune cells in the LN after processing for single-cell suspension, we compared three dissociation protocols: enzymatic digestion, mechanical dissociation with DNase I treatment, and mechanical dissociation with density gradient separation. We analyzed cell yields, viability, phenotypic and maturation markers of immune cells from the lung-draining LN of neonatal and adult mice two days after intranasal respiratory syncytial virus (RSV) infection. While viability was consistent across age groups, the protocols influenced the yield of subsets defined by important phenotypic and activation markers. Moreover, enzymatic digestion did not show higher overall yields of conventional dendritic cells and macrophages from the LN. Together, our findings show that the three dissociation protocols have similar impacts on the number and viability of cells isolated from the neonatal and adult LN. However, enzymatic digestion impacts the mean fluorescence intensity of key lineage and activation markers that may influence experimental findings.

Transcription factor STAT enhanced antimicrobial activities in Bombyx mori.

STAT, a transcription factor in the JAK/STAT signaling pathway, regulates immune response to pathogens. In the silkworm (Bombyx mori), STAT exists as two split-forms, STAT-S and STAT-L. However, the role of STAT in silkworm immunity remains unclear. Our purpose was to investigate the effect of STAT on the expression of antimicrobial peptide (AMP) genes and resistance against pathogens. The expression levels of STAT-S and STAT-L were significantly up-regulated after induction by pathogenic microorganisms. In BmE cells, lipopolysaccharide (LPS), peptidoglycan (PGN) and ß-glucan stimulated STAT-S and STAT-L to transfer from the cytoplasm to the nucleus. We found that overexpression of STAT-S and STAT-L in cells could promote the expression of AMPs. We generated transgenic silkworm lines overexpressing STAT-L or STAT-S (OE-STAT-S; OE-STAT-L) or interfering with STAT (A4-dsSTAT). Overexpression of STAT-S and STAT-L upregulated the expression of AMP genes in the OE-STAT-S and OE-STAT-L, increased the survival rates of the OE-STAT-S silkworms and lowered the mortality of OE-STAT-L silkworms infected with S. aureus or Beauveria bassiana. By contrast, the death rate of A4-dsSTAT silkworms was higher after infection with these pathogenic microorganisms. These findings may provide insights into the role of STAT in the antimicrobial immune response of silkworms.

Acid tolerance of lactate-utilizing bacteria of the order Bacteroidales contributes to prevention of ruminal acidosis in goats adapted to a high-concentrate diet.

The rapid accumulation of organic acids, particularly lactate, has been suggested as the main cause of ruminal acidosis (RA) for ruminants fed high-concentrate diets. Previous research has shown that a gradual shift from low-to high-concentrate diets within 4 to 5 weeks effectively reduces the risk for RA. However, the mechanisms remain unknown. In this study, 20 goats were randomly allocated into four groups (n = 5) and fed with a diet containing a weekly increasing concentrate portion of 20%, 40%, 60%, and 80% over 28 d. At d 7, 14, 21, and 28, one group (named C20, C40, C60, and C80 according to the last concentrate level that they received) was killed and the ruminal microbiome was collected. Ruminal acidosis was not detected in any of the goats during the experiment. Nonetheless, ruminal pH dropped sharply from 6.2 to 5.7 (P < 0.05) when dietary concentrate increased from 40% to 60%. A combined metagenome and metatranscriptome sequencing approach identified that this was linked to a sharp decrease in the abundance and expression of genes encoding nicotinamide adenine dinucleotide (NAD)-dependent lactate dehydrogenase (nLDH), catalyzing the enzymatic conversion of pyruvate to lactate (P < 0.01), whereas the expression of two genes encoding NAD-independent lactate dehydrogenase (iLDH), catalyzing lactate oxidation to pyruvate, showed no significant concomitant change. Abundance and expression alterations for nLDH- and iLDH-encoding genes were attributable to bacteria from Clostridiales and Bacteroidales, respectively. By analyzing the gene profiles of 9 metagenome bins (MAG) with nLDH-encoding genes and 5 MAG with iLDH-encoding genes, we identified primary and secondary active transporters as being the major types of sugar transporter for lactate-producing bacteria (LPB) and lactate-utilizing bacteria (LUB), respectively. Furthermore, more adenosine triphosphate was required for the phosphorylation of sugars to initiate their catabolic pathways in LPB compared to LUB. Thus, the low dependence of sugar transport systems and catabolic pathways on primary energy sources supports the acid tolerance of LUB from Bacteroidales. It favors ruminal lactate utilization during the adaptation of goats to a high-concentrate diet. This finding has valuable implications for the development of measures to prevent RA.

Increased innate immune activation induces protective RSV-specific lung-resident memory T cells in neonatal mice.

Young infants frequently experience respiratory tract infections, yet vaccines designed to provide mucosal protection are lacking. Localizing pathogen-specific cellular and humoral immune responses to the lung could provide improved immune protection. We used a well-characterized murine model of respiratory syncytial virus (RSV) to study the development of lung-resident memory T cells (TRM) in neonatal compared to adult mice. We demonstrated that priming with RSV during the neonatal period failed to retain RSV-specific clusters of differentiation (CD8) TRM 6 weeks post infection, in contrast to priming during adulthood. The reduced development of RSV-specific TRM was associated with poor acquisition of two key markers of tissue residence: CD69 and CD103. However, by augmenting both innate immune activation and antigen exposure, neonatal RSV-specific CD8 T cells increased expression of tissue-residence markers and were maintained in the lung at memory time points. Establishment of TRM correlated with more rapid control of the virus in the lungs upon reinfection. This is the first strategy to effectively establish RSV-specific TRM in neonates providing new insight into neonatal memory T cell development and vaccine strategies.

SARS-CoV-2 spike-ferritin-nanoparticle adjuvanted with ALFQ induces long-lived plasma cells and cross-neutralizing antibodies.

This study demonstrates the impact of adjuvant on the development of T follicular helper (Tfh) and B cells, and their influence on antibody responses in mice vaccinated with SARS-CoV-2-spike-ferritin-nanoparticle (SpFN) adjuvanted with either Army Liposome Formulation containing QS-21 (SpFN + ALFQ) or Alhydrogel® (SpFN + AH). SpFN + ALFQ increased the size and frequency of germinal center (GC) B cells in the vaccine-draining lymph nodes and increased the frequency of antigen-specific naive B cells. A single vaccination with SpFN + ALFQ resulted in a higher frequency of IL-21-producing-spike-specific Tfh and GC B cells in the draining lymph nodes and spleen, S-2P protein-specific IgM and IgG antibodies, and elicitation of robust cross-neutralizing antibodies against SARS-CoV-2 variants as early as day 7, which was enhanced by a second vaccination. This was associated with the generation of high titer, high avidity binding antibodies. The third vaccination with SpFN + ALFQ elicited high levels of neutralizing antibodies against the Omicron variant. No cross-neutralizing antibodies against Omicron were induced with SpFN + AH. These findings highlight the importance of ALFQ in orchestrating early induction of antigen-specific Tfh and GC B cell responses and long-lived plasma cells in the bone marrow. The early engagement of S-2P specific naive B cells and high titer IgM antibodies shape the development of long-term neutralization breadth.

Cytokine receptor DOME controls wing disc development in Bombyx mori.

In multicellular organisms, the JAK/STAT signaling pathway is involved in cell proliferation, differentiation, apoptosis, and immune regulation. Through activation of the Stat92E transcription factor, JAK/STAT signaling induced proper wing development in Drosophila. Domeless (DOME) was the first identified invertebrate JAK/STAT receptor. However, the function of DOME in Bombyx mori development remains unclear, especially in wing morphogenesis. In this study, we isolated the cytokine receptor DOME gene in B. mori and evaluated its function in DOME-knockout models. We found that overexpression of DOME at the cellular level upregulated the expression of JAK/STAT pathway-related genes, promoted proliferation, and inhibited apoptosis. The results of the interference with DOME had the opposite effects with those of overexpression at the cellular level. Using CRISPR/Cas9 technology, we constructed a DOME-knockout transgenic silkworm strain (KO-DOME) and found that the wings of the pupa and moth stages were vesicle-shaped and smaller than those of the wild-type silkworm. Some KO-DOME silkworms were unable to extend their wings from the pupal case after eclosion. We detected the expression of cyclin and apoptosis-related genes in the wing disc of the moth stage and found that some cyclin genes, such as CyclinA, CyclinB, and CyclinD, were downregulated, whereas apoptotic genes, such as Caspase1, Caspase3, and Caspase8, were upregulated. We propose that DOME regulates cell proliferation and apoptosis by affecting the JAK/STAT signaling pathway, ultimately influencing the development of wing discs. Our study provides empirical evidence for the biological function of the silkworm DOME gene, which is essential for the normal development of wings.

Direct effect of lipopolysaccharide and histamine on permeability of the rumen epithelium of steers ex vivo.

Disruption of the ruminal epithelium barrier occurs during subacute ruminal acidosis due to low pH, hyper-osmolality, and increased concentrations of lipopolysaccharide and histamine in ruminal fluid. However, the individual roles of lipopolysaccharide and histamine in the process of ruminal epithelium barriers disruption are not clear. The objective of the present investigation was to evaluate the direct effect of lipopolysaccharide and histamine on the barrier function of the ruminal epithelium. Compared with control (CON), histamine (HIS, 20 µM) increased the short-circuit current (Isc; 88.2%, P < 0.01), transepithelial conductance (Gt; 29.7%, P = 0.056), and the permeability of fluorescein 5(6)-isothiocyanate (FITC) (1.04-fold, P < 0.01) of ruminal epithelium. The apparent permeability of LPS was 1.81-fold higher than HIS (P < 0.01). The mRNA abundance of OCLN in ruminal epithelium was decreased by HIS (1.1-fold, P = 0.047). The results of the present study suggested that mucosal histamine plays a direct role in the disruption of ruminal epithelium barrier function, whereas lipopolysaccharide (at a pH of 7.4) has no effect on the permeability of rumen tissues ex vivo.

Lipopolysaccharide and histamine are common chemicals in rumen when the ruminant animal takes too much concentrate. We wandered whether these two chemicals have direct effects on the rumen tissues. Using Ussing chamber, we found that histamine could directly improve the permeability of rumen barrier.

SARS-CoV-2 ferritin nanoparticle vaccine induces robust innate immune activity driving polyfunctional spike-specific T cell responses.

The emergence of variants of concern, some with reduced susceptibility to COVID-19 vaccines underscores consideration for the understanding of vaccine design that optimizes induction of effective cellular and humoral immune responses. We assessed a SARS-CoV-2 spike-ferritin nanoparticle (SpFN) immunogen paired with two distinct adjuvants, Alhydrogel® or Army Liposome Formulation containing QS-21 (ALFQ) for unique vaccine evoked immune signatures. Recruitment of highly activated multifaceted antigen-presenting cells to the lymph nodes of SpFN+ALFQ vaccinated mice was associated with an increased frequency of polyfunctional spike-specific memory CD4+ T cells and Kb spike-(539-546)-specific long-lived memory CD8+ T cells with effective cytolytic function and distribution to the lungs. The presence of this epitope in SARS-CoV, suggests that generation of cross-reactive T cells may be induced against other coronavirus strains. Our study reveals that a nanoparticle vaccine, combined with a potent adjuvant that effectively engages innate immune cells, enhances SARS-CoV-2-specific durable adaptive immune T cell responses.

Corrigendum: The Regulation of Ruminal Short-Chain Fatty Acids on the Functions of Rumen Barriers.

[This corrects the article DOI: 10.3389/fphys.2019.01305.].

Corrigendum: Dietary Energy Level Promotes Rumen Microbial Protein Synthesis by Improving the Energy Productivity of the Ruminal Microbiome.

[This corrects the article DOI: 10.3389/fmicb.2019.00847.].

Durability of SARS-CoV-2-Specific T-Cell Responses at 12 Months Postinfection.

BACKGROUND: Characterizing the longevity and quality of cellular immune responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) enhances understanding of coronavirus disease 2019 (COVID-19) immunity that influences clinical outcomes. Prior studies suggest SARS-CoV-2-specific T cells are present in peripheral blood 10 months after infection. Analysis of the function, durability, and diversity of cellular response long after natural infection, over a range of ages and disease phenotypes, is needed to identify preventative and therapeutic interventions. METHODS: We identified participants in our multisite longitudinal, prospective cohort study 12 months after SARS-CoV-2 infection representing a range of disease severity. We investigated function, phenotypes, and frequency of T cells specific for SARS-CoV-2 using intracellular cytokine staining and spectral flow cytometry, and compared magnitude of SARS-CoV-2-specific antibodies. RESULTS: SARS-CoV-2-specific antibodies and T cells were detected 12 months postinfection. Severe acute illness was associated with higher frequencies of SARS-CoV-2-specific CD4 T cells and antibodies at 12 months. In contrast, polyfunctional and cytotoxic T cells responsive to SARS-CoV-2 were identified in participants over a wide spectrum of disease severity. CONCLUSIONS: SARS-CoV-2 infection induces polyfunctional memory T cells detectable at 12 months postinfection, with higher frequency noted in those who experienced severe disease.

Bombyx mori nucleopolyhedrovirus downregulates transcription factor BmFoxO to elevate virus infection.

Forkhead-box O (FoxO) is the primary transcriptional effector of the insulin-like signaling pathway that enhances gluconeogenesis through transcriptional activation of PEPCK and G6Pase in mammals. We have previously demonstrated the involvement of phosphoenolpyruvate carboxykinase (BmPEPCK-2) in antiviral immunity against the multiplication of Bombyx mori nuclearpolyhedrosisvirus (BmNPV) in silkworm. Therefore, we speculated that BmFoxO might suppress BmNPV by regulating the expression of PEPCK in silkworm. In the present study, we found that the expression of BmFoxO decreased after BmNPV infection in Bombyx mori; this finding was consistent with BmPEPCK-2 expression. In addition, the expression of BmFoxO was altered, and it was found that reduced expression of BmFoxO (dsBmFoxO) downregulated the expression of BmPEPCK-2 and increased the viral fluorescence and content in silkworm embryonic cell line BmE cells, and vice versa. BmFoxO could upregulate the expression of BmPEPCK-2 by binding to the BmPEPCK-2 promoter. Moreover, overexpression of BmFoxO significantly increased the expression of autophagy genes ATG6/7/8 after infection with BmNPV, consistent with BmPEPCK-2. These results indicate that BmNPV downregulates transcription factor BmFoxO to elevate virus infection, and BmFoxO overexpression upregulates BmPEPCK-2 expression and enhances silkworm antiviral resistance.

Co-immunization of DNA and Protein in the Same Anatomical Sites Induces Superior Protective Immune Responses against SHIV Challenge.

We compare immunogenicity and protective efficacy of an HIV vaccine comprised of env and gag DNA and Env (Envelope) proteins by co-administration of the vaccine components in the same muscles or by separate administration of DNA + protein in contralateral sites in female rhesus macaques. The 6-valent vaccine includes gp145 Env DNAs, representing six sequentially isolated Envs from the HIV-infected individual CH505, and matching GLA-SE-adjuvanted gp120 Env proteins. Interestingly, only macaques in the co-administration vaccine group are protected against SHIV CH505 acquisition after repeated low-dose intravaginal challenge and show 67% risk reduction per exposure. Macaques in the co-administration group develop higher Env-specific humoral and cellular immune responses. Non-neutralizing Env antibodies, ADCC, and antibodies binding to FcγRIIIa are associated with decreased transmission risk. These data suggest that simultaneous recognition, processing, and presentation of DNA + Env protein in the same draining lymph nodes play a critical role in the development of protective immunity.

Functional Changes of the Community of Microbes With Ni-Dependent Enzyme Genes Accompany Adaptation of the Ruminal Microbiome to Urea-Supplemented Diets.

Urea is an inexpensive non-protein nitrogen source commonly supplemented to the diets of ruminants. It is cleaved to ammonia by bacterial ureases, which require Ni as a catalyst for ureolysis. The key event in the changes of the ruminal microbiome after urea supplementation remains unknown. We have therefore investigated changes in the ruminal microbiome and its community with Ni-dependent enzyme genes following urea supplementation and analyzed the associations of rumen environmental factors, including fermentation variables and Ni concentrations, with the compositional and functional changes of these communities. We found that urea supplementation increased urease activity and the concentrations of ammonia and Ni, and tended to increase concentrations of short chain fatty acids and acetate, whereas it decreased rumen pH and the L-/D-lactate ratio. With standards for genome completeness >60% and strain heterogeneity <10%, 20 bacterial species containing five Ni-dependent enzyme genes were detected in the metagenome sequences. For the five Ni-dependent enzyme genes, urea supplementation increased the relative abundances of genes of urease and acetyl-CoA synthase, whereas it decreased the relative abundances of genes of glyoxalase I, [NiFe]-hydrogenase, and lactate racemase. For the 20 microbes with Ni-dependent enzyme genes, urea supplementation increased the relative abundances of five bacteria exhibiting high capacities for the utilization of hemicellulose and pectin for butyrate and fatty acid biosynthesis. For the ruminal microbiome, urea supplementation increased the metagenomic capacities for hemicellulose and pectin degradation, butyrate generation, fatty acid biosynthesis, and carbon fixation, whereas it decreased the metagenomic capacities for starch degradation, propionate generation, and sulfur and nitrogen metabolism. Constrained correspondence analysis identified rumen ammonia and Ni concentrations as likely driving factors in the reshaping of the ruminal microbiome and, together with pH, of the community of microbes with Ni-dependent enzyme genes. Thus, the functional change of the latter community is probably an important event in the adaptation of the ruminal microbiome to urea-supplemented diets. This result provides a new perspective for the understanding of the effects of urea supplementation on rumen fermentation.

Transcriptomic analyses suggest a dominant role of insulin in the coordinated control of energy metabolism and ureagenesis in goat liver.

BACKGROUND: The ureagenesis plays a central role in the homeostatic control of nitrogen metabolism. This process occurs in the liver, the key metabolic organ in the maintenance of energy homeostasis in the body. To date, the understanding of the influencing factors and regulators of ureagenesis in ruminants is still poor. The aim of this study was to investigate the relationship between energy metabolism and ureagenesis and detect the direct regulators of ureagenesis in the liver by using RNA-seq technology. RESULTS: Eighteen four-month-old male goats were divided into two groups randomly and received a diet containing 10% (LNFC group, n = 9) or 30% non-fiber carbohydrate (MNFC group, n = 9), respectively, for four weeks. The global gene expression analysis of liver samples showed that, compared with a LNFC diet, the MNFC diet promoted the expression of genes required for synthesis of fatty acid and glycerol, whereas it suppressed those related to fatty acid oxidation, gluconeogenesis from amino acids and ureagenesis. Additionally, gene expression for rate-limiting enzymes of ureagenesis were highly correlated to the gene expression of key enzymes of both fatty acid synthesis and glycerol synthesis (Spearman correlation coefficient > 0.8 and p < 0.05). In the differentially expressed signaling pathways related to the endocrine system, the MNFC diet activated the insulin and PPAR signaling pathway, whereas it suppressed the leptin-JAK/STAT signaling pathway, compared with the LNFC diet. Reverse transcription quantitative PCR analyses of 40 differentially expressed genes confirmed the RNA-seq results (R2 = 0.78). CONCLUSION: Our study indicated that a dietary NFC-induced increase of energy supply promoted lipid anabolism and decreased ureagenesis in the caprine liver. By combining our results with previously published reports, insulin signaling can be suggested to play the dominant role in the coordinated control of hepatic energy metabolism and ureagenesis.

The Regulation of Ruminal Short-Chain Fatty Acids on the Functions of Rumen Barriers.

The rumen barriers, constituted by the microbial, physical and immune barrier, prevent the transmission of pathogens and toxins to the host tissue in the maintenance of host-microbe homeostasis. Ruminal short-chain fatty acids (SCFAs), which are the important signaling molecules derived from the rumen microbiota, regulate a variety of physiological functions of the rumen. So far, how the ruminal SCFAs regulate the function of rumen barriers is unclear. By the combined methods of transcriptome sequencing, 16S rRNA gene sequencing, and metagenome shotgun sequencing, we have investigated the regulatory effects of ruminal SCFAs on the functions of rumen barriers, by determining the composition and functions of epimural microbiota and on the structure and immunity of the rumen epithelium in goats receiving a 10% (LC group), 35% (MC group), or 65% concentrate diet (HC group). We found that, when the dietary concentrate shifted from 10 to 35%, the increase of total SCFA is associated with the diversification of epimural microbiota and the diversity of its gene pool. Within the microbial community, the relative abundance of genera Sphingobium, Acinetobacter, and Streptococcus increase mostly. Meantime, the signals on pathways concerning the mechanical connections and growth homeostasis in the rumen epithelium were upregulated. Under these conditions, the responses of immune components in the rumen epithelium decrease. However, when the dietary concentrate shifted from 35 to 65%, the increase of acetate and reduction of pH decrease the diversity of epimural microbiota and the diversity of its gene pool. Within the microbial community, the relative abundance of genera Sphingobium, Acinetobacter, and Streptococcus significantly decrease. Concomitantly, the signals on pathways concerning the cell growth and tight junction disruption were upregulated, while the signals on pathways concerning paracellular permeability were downregulated. Under these conditions, the signals on the pathways relating to the immune components increase. Our data thus indicates that diet-SCFA axis maintains the host-microbe homeostasis via promoting the diversification of epimural microbiota and maintaining the integrity of rumen epithelium in healthy animals, while via enhancing the activities of immune barrier in animal with lower rumen pH.

Effects of Dietary-SCFA on Microbial Protein Synthesis and Urinal Urea-N Excretion Are Related to Microbiota Diversity in Rumen.

Two experiments were performed in this study. In Experiment 1, twenty goats were fed with an isonitrogenous diet, containing 28% Non-Fiber Carbohydrate (MNFC group, n = 10) or 14% NFC (LNFC group, n = 10). In the MNFC group, the ruminal concentration of Short Chain Fatty Acids (SCFA) increased, and pH declined. Compared with those in the LNFC group, the microbial protein synthesis in rumen and mRNA abundance of urea transporter B (UT-B) in rumen epithelium increased in the MNFC group, although serum urea-N (SUN) did not differ significantly between groups. Simultaneously, urinal urea-N excretion was reduced in the MNFC group. Significant correlations were found between rumen SCFA and UT-B and between UT-B and urinal urea-N excretion. Furthermore, the abundances of SCFA receptor of GPR41 and GPR43 increased in the rumen epithelium of the MNFC group. These results suggest that increases of SUN transported into the rumen and incorporated into microbial protein and decreases of urinal urea-N excretion are related to ruminal SCFA. This is supported by data from our previous study in which added SCFA on the mucosal side caused increases of urea transport rate (flux Jsm urea) from the blood to the ruminal lumen side. In Experiment 2, we used 16S rRNA Amplicon Sequencing to analyze the structure of the ruminal microbiota community in relation to SCFA. An additional eight goats were assigned into the MNFC (n = 4) and LNFC (n = 4) groups. The dietary ingredients, chemical composition, and feeding regimes were the same as those in Experiment 1. Constrained correspondence analysis (CCA analysis) revealed NFC promoted the expansion of microbiota diversity, particularly of SCFA-producing microbes. The function prediction of 19 upregulated Kyoto Encyclopedia of Genes and Genomes (KEGG) ortholog groups showed an NFC-induced increase of the types and abundances of genes coding for enzymes catalyzing N and fatty acid metabolism. Based on our present and previous investigations, our results indicate that, in goats consuming a MNFC diet, the facilitated urea transport in the rumen and improved urea N salvage are triggered by an expansion of ruminal microbiota diversity and are signaled by ruminal SCFA. This study thus provides new insights into the microbiota involved in the dietary modulation of urea-N salvage in ruminant animals.

Effects of glucagon-like peptides 1 and 2 and epidermal growth factor on the epithelial barrier of the rumen of adult sheep.

Epidermal growth factor (EGF) and glucagon-like peptides (GLP) modulate the tight junctions (TJ) of the intestinal epithelial barrier (EB) of monogastric animals. This work tried to elucidate whether GLP-1, GLP-2 and EGF can affect the EB of the rumen. Ovine ruminal epithelia were incubated in Ussing chambers for 7 hr with 25 or 250 nM of either GLP-1 or GLP-2 on the serosal side, with 2.5 nM of EGF on the serosal side or with 0.25 or 2.5 nM EGF on the mucosal side. No treatment affected tissue conductance. Short-circuit current (Isc ) was affected by time and treatment and their interactions. Only 250 nM of either GLP-1 or GLP-2 decreased Isc in certain periods compared with 25 nM GLP-1 or 0.25 nM mucosally applied EGF; however, not when compared to control epithelia. Fluorescein flux rates (Jfluor ) of ruminal epithelia were affected by treatment, time and time × treatment interaction. The time × treatment interaction was based on an increase in Jfluor between the first and last hour in epithelia incubated with 25 nM GLP-1 or GLP-2 and in epithelia incubated with EGF. After 7 hr incubation, claudin-7 mRNA expression was downregulated in all treatments. Claudin-1 mRNA was upregulated after incubation with 2.5 nM EGF on the serosal side, claudin-4 mRNA was downregulated by 2.5 nM EGF on the mucosal side, and occludin mRNA was increased after incubation with 250 nM GLP-2. The protein abundance of all tested TJ proteins was not influenced by treatment. We conclude that GLP-1, GLP-2, and EGF have no obvious acute effects on the EB of ruminal epithelia under simulated physiological conditions ex vivo. However, by decreasing the mRNA expression of claudin-7 and partly affecting other TJ proteins, they may modulate EB in the longer term or under certain conditions.

Overexpression of BmFoxO inhibited larval growth and promoted glucose synthesis and lipolysis in silkworm.

Forkhead box O (FoxO) is a downstream transcription factor of the insulin-signaling pathway, which plays vital roles in the growth and metabolism of organisms. In this study, BmFoxO was overexpressed in BmE cells, in which proliferation was inhibited and apoptosis was increased. The transgenic vector overexpressing BmFoxO was constructed, and the transgenic silkworm line A4FoxO was generated via embryonic microinjection. The body size of A4FoxO silkworm was smaller than that of non-transgenic silkworm (WT). The quantitative polymerase chain reaction results revealed that the insulin pathway was enhanced and the growth-related TOR pathway was suppressed. Furthermore, the translation of proteins in the fat body of A4FoxO silkworm was inhibited. The expression level of genes involved in the glucose synthesis and lipolysis pathways was increased, whereas that of genes involved in fat synthesis was decreased. Oil red O staining revealed that the amount of lipid droplets was reduced in A4FoxO silkworms compared with WT. Further analysis showed that the content of triglyceride and glycogen was significantly decreased in fat body, but the content of glucose and trehalose was increased in the hemolymph of A4FoxO silkworms. These results suggest that the enhanced expression of BmFoxO disturbs glycolipid metabolism and affects silkworm growth.