A bi-allelic REC114 loss-of-function variant causes meiotic arrest and nonobstructive azoospermia.

Nonobstructive azoospermia (NOA), the most severe manifestation of male infertility, lacks a comprehensive understanding of its genetic etiology. Here, a bi-allelic loss-of-function variant in REC114 (c.568C > T: p.Gln190*) were identified through whole exome sequencing (WES) in a Chinese NOA patient. Testicular histopathological analysis and meiotic chromosomal spread analysis were conducted to assess the stage of spermatogenesis arrested. Co-immunoprecipitation (Co-IP) and Western blot (WB) were used to investigate the influence of variant in vitro. In addition, our results revealed that the variant resulted in truncated REC114 protein and impaired interaction with MEI4, which was essential for meiotic DNA double-strand break (DSB) formation. As far as we know, this study presents the first report that identifies REC114 as the causative gene for male infertility. Furthermore, our study demonstrated indispensability of the REC114-MEI4 complex in maintaining DSB homoeostasis, and highlighted that the disruption of the complex due to the REC114 variant may underline the mechanism of NOA.

Bi-allelic MEI1 variants cause meiosis arrest and non-obstructive azoospermia.

Non-obstructive azoospermia (NOA) is characterized by the failure of sperm production due to testicular disorders and represents the most severe form of male infertility. Growing evidences have indicated that gene defects could be the potential cause of NOA via genome-wide sequencing approaches. Here, bi-allelic deleterious variants in meiosis inhibitor protein 1 (MEI1) were identified by whole-exome sequencing in four Chinese patients with NOA. Testicular pathologic analysis and immunohistochemical staining revealed that spermatogenesis is arrested at spermatocyte stage, with defective programmed DNA double-strand breaks (DSBs) homoeostasis and meiotic chromosome synapsis in patients carrying the variants. In addition, our results showed that one missense variant (c.G186C) reduced the expression of MEI1 and one frameshift variant (c.251delT) led to truncated proteins of MEI1 in in vitro. Furthermore, the missense variant (c.T1585A) was assumed to affect the interaction between MEI1 and its partners via bioinformatic analysis. Collectively, our findings provide direct genetic and functional evidences that bi-allelic variants in MEI1 could cause defective DSBs homoeostasis and meiotic chromosome synapsis, which subsequently lead to meiosis arrest and male infertility. Thus, our study deepens our knowledge of the role of MEI1 in male fertility and provides a novel insight to understand the genetic aetiology of NOA.

Identification of a missense variant of MND1 in meiotic arrest and non-obstructive azoospermia.

Meiotic arrest is a common pathologic phenotype of non-obstructive azoospermia (NOA), yet its genetic causes require further investigation. Meiotic nuclear divisions 1 (MND1) has been proved to be indispensable for meiotic recombination in many species. To date, only one variant of MND1 has been reported associated with primary ovarian insufficiency (POI), yet there has been no report of variants in MND1 associated with NOA. Herein, we identified a rare homozygous missense variant (NM_032117:c.G507C:p.W169C) of MND1 in two NOA-affected patients from one Chinese family. Histological analysis and immunohistochemistry demonstrated meiotic arrest at zygotene-like stage in prophase I and lack of spermatozoa in the proband's seminiferous tubules. In silico modeling demonstrated that this variant might cause possible conformational change in the leucine zippers 3 with capping helices (LZ3wCH) domain of MND1-HOP2 complex. Altogether, our study demonstrated that the MND1 variant (c.G507C) is likely responsible for human meiotic arrest and NOA. And our study provides new insights into the genetic etiology of NOA and mechanisms of homologous recombination repair in male meiosis.

A homozygous frameshift variant in SYCP2 caused meiotic arrest and non-obstructive azoospermia.

Genetic causation for the majority of non-obstructive azoospermia (NOA) remains unclear. Mutations in synaptonemal complex (SC)-associated genes could cause meiotic arrest and NOA. Previous studies showed that heterozygous truncating variants in SYCP2 encoding a protein essential for SC formation, are associated with non-obstructive azoospermia and severe oligozoospermia. Herein, we showed a homozygous loss-of-function variant in SYCP2 (c.2689_2690insT) in an NOA-affected patient. And this variant was inherited from heterozygous parental carriers by natural reproduction. HE, IF, and meiotic chromosomal spread analyses demonstrated that spermatogenesis was arrested at the zygotene stage in the proband with NOA. Thus, this study revealed that SYCP2 associated with NOA segregates in an autosomal recessive inheritance pattern, rather than an autosomal dominant pattern. Furthermore, our study expanded the knowledge of variants in SYCP2 and provided new insight into understanding the genetic etiology of NOA.

Hydrogen Sulfide Regulates Glucose Uptake in Skeletal Muscles via S-Sulfhydration of AMPK in Muscle Fiber Type-Dependent Way.

BACKGROUND: The effect of hydrogen sulfide (H2S) on glucose homeostasis remains to be elucidated, especially in the state of insulin resistance. OBJECTIVES: In the present study, we aimed to investigate H2S-regulated glucose uptake in the M. pectoralis major (PM) muscle (which mainly consists of fast-twitch glycolytic fibers) and M. biceps femoris (BF) muscle (which mainly consists of slow-twitch oxidative fibers) of the chicken, a potential model of insulin resistance. METHODS: Chicks were subjected to intraperitoneal injection of sodium hydrosulfide (NaHS, 50 µmol/kg body mass/day) twice a day to explore glucose homeostasis. In vitro, myoblasts from PM and BF muscles were used to detect glucose uptake and utilization. Effects of AMP-activated protein kinase (AMPK) phosphorylation, AMPK S-sulfhydration, and mitogen-activated protein kinase (MAPK) pathway induction by NaHS were detected. RESULTS: NaHS enhanced glucose uptake and utilization in chicks (P < 0.05). In myoblasts from PM muscle, NaHS (100 µM) increased glucose uptake by activating AMPK S-sulfhydration, AMPK phosphorylation, and the AMPK/p38 MAPK pathway (P < 0.05). However, NaHS decreased glucose uptake in myoblasts from BF muscle by suppressing the p38 MAPK pathway (P < 0.05). Moreover, NaHS increased S-sulfhydration and, in turn, the phosphorylation of AMPK (P < 0.05). CONCLUSIONS: This study reveals the role of H2S in enhancing glucose uptake and utilization in chicks. The results suggest that NaHS is involved in glucose uptake in skeletal muscle in a fiber type-dependent way. The AMPK/p38 pathway and protein S-sulfhydration promote glucose uptake in fast-twitch glycolytic muscle fibers, which provides a muscle fiber-specific potential therapeutic target to ameliorate glucose metabolism.

Chicken embryo thermal manipulation alleviates postnatal heat stress-induced jejunal inflammation by inhibiting Transient Receptor Potential V4.

Intestinal inflammation induced by heat stress is an important factor restricting the healthy growth of broilers. The aim of this study was to evaluate the effect of chicken embryo thermal manipulation (39.5 â„ƒ and 65 % RH for 3 h daily during 16-18 th embryonic age) on intestinal inflammation in broilers under postnatal heat stress and to investigate whether transient receptor potential V4 (TRPV4) plays a role in this process. Our results suggest that broilers with embryo thermal manipulation experience could delay the rising of rectal temperature during postnatal heat stress (P < 0.05), and had better production performance (P < 0.05), intestinal morphological parameters (P < 0.05) and higher expression of tight junction related genes (P < 0.05). The increased serum lipopolysaccharide (LPS) content, activation of nuclear factor-kappa B (NF-κB) signaling pathway and the increased expression of pro-inflammatory cytokines interleukin (IL)-1ß, IL-6 and tumor necrosis factor alpha (TNF-α) in jejunum during postnatal heat stress were alleviated by embryo thermal manipulation (P < 0.05). Postnatal heat stress induced an increase in mRNA and protein expression of TRPV4 in jejunum (P < 0.05), but had no effect on broilers which experienced embryo thermal manipulation (P > 0.05). Inhibition of TRPV4 reduced LPS-induced Ca2+ influx and restrained the activation of NF-κB signaling pathway and the expression of downstream pro-inflammatory cytokines (P < 0.05). The expression of DNA methyltransferase (DNMT) in the jejunum of broilers exposed to postnatal heat stress was increased by embryo thermal manipulation (P < 0.05). The DNA methylation level of TRPV4 promoter region was detected, and the results showed that embryo thermal manipulation increased the DNA methylation level of TRPV4 promoter region (P < 0.05). In conclusion, Chicken embryo thermal manipulation can alleviate jejunal inflammation in broilers under postnatal heat stress. This may be due to the decreased circulating LPS or the increased DNA methylation level in the promoter region of TRPV4, which inhibits TRPV4 expression, thereby reducing Ca2+ influx, and finally alleviating inflammation by affecting NF-κB signaling pathway. The work is an attempt to understand the mechanism involved in alleviation of adverse effects of heat stress during postnatal life through prenatal thermal manipulation and to reveal the important role of epigenetics.

The regulating pathway of creatine on muscular protein metabolism depends on the energy state.

Creatine (Cr) is beneficial for increasing muscle mass and preventing muscle atrophy via involving in energy metabolism through the Cr and phosphocreatine (PCr) system. This study aimed to evaluate the supplemental effect of Cr on protein metabolism under normal and starvation conditions. The primary myoblasts were obtained from the breast muscle of chicks. The mammalian target of rapamycin (mTOR)/P70S6 kinase (P70S6K), ubiquitin-proteasome (UP) pathways, and mitochondrial function of myotubes were evaluated at normal or starvation state and with or without glucose supplementation. Under normal condition, Cr supplementation enhanced protein synthesis rate as well as upregulated the total and phosphorylated P70S6K expressions. Cr had little influence on protein catabolism and mitochondrial function. In a starvation state, however, Cr alleviated myotube atrophy and enhanced protein accretion by inhibiting Atrogin1 and myostatin (MSTN) expression. Furthermore, Cr treatment upregulated the transcriptional coactivators peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) expression and decreased reactive oxygen species (ROS) accumulation under starvation condition. In the presence of glucose, however, the favorable effect of Cr on protein content and myotube diameter did not occur under starvation condition. The present result indicates that at a normal state, Cr stimulated protein synthesis via the mTOR/P70S6K pathway. In a starvation state, Cr mainly takes a favorable effect on protein accumulation via suppression of the UP pathway and mediated mitochondrial function mainly by serving as an energy supplier. The result highlights the potential clinical application for the modulation of muscle mass under different nutritional conditions.

Bi-allelic SPATA22 variants cause premature ovarian insufficiency and nonobstructive azoospermia due to meiotic arrest.

The genetic causes of idiopathic premature ovarian insufficiency (POI) and nonobstructive azoospermia (NOA) remain unclear. We performed whole-exome sequencing (WES) in members of a consanguineous family with two POI and two NOA patients to screen for potential pathogenic variants for familial POI and NOA. And a homozygous variant in SPATA22 (c.400C>T:p.R134X) was identified. Histological analysis and spermatocyte spreading assay demonstrated that the spermatogenesis was arrested at a zygotene-like stage in the proband with NOA. The candidate gene was further screened in the in-house WES database of idiopathic POI-affected patients. One additional compound heterozygous variant in SPATA22 (c.900+1G>A and c.31C>T:p.R11X) was found in one patient with sporadic POI and validated by minigene assay. Thus, this is the first report identifying SPATA22 as the causative gene for human POI. Combined with the observations in the familial patient with NOA, our findings highlighted the essential role of meiotic HR genes in gametogenesis and gonadal function maintenance.

Dietary L-citrulline influences body temperature and inflammatory responses during nitric oxide synthase inhibition and endotoxin challenge in chickens.

Recent studies have revealed the role of L-citrulline (L-CIT) in thermoregulation, but very little is known about the mechanisms involved. In this study, nitric oxide synthase inhibition and endotoxin-induced fever were used to investigate the effects of L-CIT on body temperature and inflammatory responses. In experiment 1, NW-nitro-L-arginine methyl ester (L-NAME, 150 mg/kg BW), was i. p. injected into chicks fed with basal (CON) or L-CIT diets for 14 days. In experiment 2, Lipopolysaccharide (LPS, 2 mg/kg BW) was i. p. injected following 21d feeding with CON or L-CIT diets. In experiment 3, chickens were injected with either L-NAME, LPS, or L-NAME + LPS following 26 days feeding with CON or L-CIT diets. The rectal (RT), ear (ET), and core body temperature (CBT) of chickens were examined. Results showed that L-NAME effectively decreased the RT, ET, CBT, and plasma NO concentration. In contrast, LPS increased NO levels and initiated hyperthermia by increasing RT, ET, CBT, and PGE2 levels. L-CIT diet reduced the mean CBT in experiment 1 and diminished the NO level, PGE2 level, and mean RT in experiment 3. Co-administration of L-CIT + LPS upregulated IL-6 expression, whereas, LPS injection alone induced IL-10, IL-1ß, and TLR4 gene expressions. Therefore, this study reveals that L-CIT-induced hypothermia was related to NO inhibition and a decrease in PGE2 concentration. Conversely, LPS induced hyperthermia was associated with an increase in both NO and PGE2 concentrations.

Preliminary experience with 3D digital image microscope system on the treatment of varicocele.

The three-dimension digital image microscope system (3D-DIM) with a better ergonomic design and equipment characteristics can contribute to the achievement of good results during microsurgery. In this study, the safety and efficiency of 3D-DIM assisted varicocelectomy was evaluated. From July 2019 to November 2019, fifteen cases with varicocele (20 sides of varicocele in total) were included, seven cases underwent 3D-DIM-assisted modified microsurgical subinguinal varicocelectomy, and eight cases underwent modified microsurgical subinguinal varicocelectomy under standard operating microscope (SOM). The mean operative time of 3D-DIM group (67 ± 12.3 min) was a little longer than that of SOM group (55 ± 12.9 min) (p < 0.05). There was no significant difference between the two groups in the number of internal spermatic arteries, internal spermatic vein, lymphatics, gubernacular vein, external spermatic vein and post-operation complications. The 3D-DIM showed a significant difference in image definition for nurse (p < 0.01) and in doctor-nurse cooperation (p < 0.05) over SOM. The 3D-DIM with better ergonomic design and image definition can be applied to perform microsurgical subinguinal varicocelectomy, and could improve the surgeon's fatigue and doctor-nurse cooperation. We believe that the 3D-DIM would be widely used in the field of male infertility microsurgery in the near future.

Effects of dietary L-citrulline supplementation on nitric oxide synthesis, immune responses and mitochondrial energetics of broilers during heat stress.

L-Citrulline is a non-protein amino acid that promotes arginine recycling and muscle protein synthesis. This study investigated whether L-citrulline can exert these functions in heat-stressed chickens. Arbor acre broilers were fed either basal diets (Control) or basal diets supplemented with 1% L-citrulline (L-Cit). At 28 d old, broilers were subjected to two environmental temperatures, 35 °C for 8 h/d (HS) or 24 °C for 24 h/d (TNZ) for 2 weeks. The experiment was designed as a 2 by 2 factorial arrangement. Results showed that HS increased the core body temperature (CBT) and rectal temperature of broilers compared to the TNZ condition. The CBT and mean CBT decreased by ∼0.5oC in TNZ + L-Cit broilers compared to the TNZ + Control group (P < 0.05). L-Cit supplementation at TNZ significantly (P < 0.05) lowered the plasma malondialdehyde content but this was increased during HS. Exposure to HS significantly (P < 0.05) elevated plasma malondialdehyde compared to TNZ condition. Plasma immunoglobulin A, G, and M were increased (P < 0.05) by TNZ + L-Cit compared to the TNZ + Control group. Plasma nitric oxide and endothelial nitric oxide synthase (NOS) activity was increased during HS, whereas the total and inducible NOS enzymes were decreased. In addition, L-Cit supplementation increased both the inducible and endothelial NOS isoforms (P < 0.05). The mRNA expression of peroxisome proliferator-activated receptor γ coactivator 1-α, avian uncoupling protein, cytochrome C oxidase subunit 3, and ATP synthase F1 subunit beta (ATP5ß) in the breast muscle were downregulated (P < 0.05) during HS. However, L-Cit supplementation upregulated the mitochondrial transcription factor A and during HS, L-Cit increased ATP5ß expression similar to TNZ housed broilers. Therefore, this study demonstrates that dietary L-Cit can lower the body temperature, decrease lipid peroxidation and promote the immune status of broilers under thermoneutrality. Also, L-Cit would act to promote muscle ATP generation during heat stress in broilers.

Vasal vessel-sparing microsurgical single-armed vasoepididymostomy to epididymal obstructive azoospermia: A retrospective control study.

This study aimed to evaluate the efficacy and safety of vasal vessel-sparing modified single-armed 2-suture longitudinal intussusception vasoepididymostomy (SA-LIVE) to epididymal obstructive azoospermia patients. Forty consecutive epididymal obstructive azoospermia cases, who underwent microsurgical vasoepididymostomy in Shanghai General Hospital from January 2019 to October 2019, were included in this study. Twenty cases underwent SA-LIVE (group A), and 20 cases underwent vasal vessel-sparing SA-LIVE (group B). Until March 2021, the mean follow-up period was 16.9 ± 4.1 (12-23) months. The overall patency rate was 82.5%, and 80% and 85% for group A and group B respectively. The mean time to achieve patency was 4.11 ± 2.74 months. The overall natural pregnancy rate was 51.5%(17/33) at the mean follow-up of 16.9 months. The natural pregnancy rate was 50.0% for group A and 52.9% for group B (p > .05). At the time of 6 months post-operation, the patency rate was 70% for group A and 80% for group B (p = .465); the natural pregnancy rate was 0% for group A and 31.3% for group B (p = .022). Vasal vessel-sparing SA-LIVE is safe and effective to achieve favourable patency and pregnancy rates. Preserving vasal vessel would improve natural pregnancy rate at a very early stage.

Dusk feeding in laying hens is shifted by light program via involvement of clock genes.

Dusk feeding is practised probably to satisfy the energy requirement during night. However, little is known on the changes with clock gene expressions during this feeding behaviour. In our present study, the linkage of clock gene expressions and feeding behaviour in dusk feeding was investigated in laying hens under two lighting programs: the conventional lighting program (Control) with a light period from 05:00 AM to 21:00 PM and a dark period from 21:00 PM to 05:00 AM; or the shifted lighting program group (SLP) with a light period from 02:00 AM to 18:00 PM and a dark period from 18:00 PM to 02:00 AM. The gene expression-related appetite and circadian rhythm were investigated in hypothalamus and proventriculus at 1, 3 and 5 h before scotophase. The results demonstrated that dusk feeding was synchronously shifted with altered lighting program, dusk feeding was observed from 5 h before scotophase in both groups. The expressions of anorexigenic gene proopiomelanocortin (POMC) and cocaine- and amphetamine-regulated transcript (CART) were downregulated (p < 0.05) during dusk feeding and changed in pace with lighting program. The expressions of clock gene period 2 (Per2) and cryptochrome 1 (Cry1) in hypothalamus were downregulated (p < 0.05) during dusk feeding and shifted by lighting program. In the proventriculus, ghrelin expression was decreased (p < 0.05) during dusk feeding by lighting program. In conclusion, the expressions of clock genes Per2 and Cry1 are linked with the downregulated expressions of anorexigenic genes, POMC and CART, and in turn the augmented feed intake at dusk.

Ghrelin serves as a signal of energy utilization and is involved in maintaining energy homeostasis in broilers.

Ghrelin, one of the most important appetite regulating peptides, is involved in the regulation of energy homeostasis. The anorexia effect of ghrelin in chickens is contrary to that of ghrelin in mammals. In the present study, the effects of feeding status and dietary energy level on plasma total ghrelin levels and expression were studied in broilers. The gene expression of ghrelin and its receptor GHS-R1a were measured in the hypothalamus, proventriculus, duodenum, liver, and abdominal fat pad. The results showed that ghrelin mRNA and GHS-R1a mRNA are moderately expressed in liver and abdominal fat. Ghrelin secretion was increased by fasting and refeeding. The gene expression of ghrelin and GHS-R1a in the hypothalamus, proventriculus, liver, and abdominal fat pad were changed by feeding status and dietary energy level. The results suggest that ghrelin is a signal of energy utilization in chickens. The abundant expression of ghrelin and GHS-R1a in liver and abdominal fat pad may be associated with energy balance.

Dexamethasone and insulin stimulate ghrelin secretion of broilers in a different way.

Ghrelin is one of the most important appetite regulating peptides, involved in the regulation of energy homeostasis. The role of ghrelin on the appetite and fat metabolism in chickens is different from that of ghrelin in mammals. Glucocorticoids and insulin are important hormones and work differently in energy regulation of body. In this study, the effects of dexamethasone (DEX, 2.0 mg/kg BW), subcutaneous insulin injection (40 µg/kg BW), and glucose load on ghrelin secretion and expression were determined in broilers. DEX treatment increased circulating ghrelin concentration in broiler fed with either a low-energy diet (11.05 MJ/kg of metabolizable energy) or a high-energy diet (14.44 MJ/kg of metabolizable energy). The expression levels of ghrelin were increased while both ghrelin and its receptor GHS-R1a expression levels were stimulated by DEX. A single subcutaneous insulin injection (40 µg/kg BW) or oral glucose infusion (2 g/kg BW) rise circulating ghrelin level. Ghrelin expression in the proventriculus was increased by insulin treatment but unchanged by glucose load. DEX had no detectable influence on ghrelin and GHS-R1a expression in the hypohtalamus, whereas insulin suppressed their expression. In conclusion, both insulin and glucocorticoid stimulate ghrelin secretion in chickens, in contrast to mammals. Glucocorticoids evoke peripheral ghrelin/GHS-R1a system while insulin increases peripheral ghrelin expression and suppress the activation of central ghrelin/GHS-R1a system. The result suggests that ghrelin involved in the modulating network of energy homeostasis in concert with glucocorticoids and insulin.

Effect of energy level of pullet diet and age on laying performance and expression of hypothalamus-pituitary-gonadal related genes in laying hens.

Dietary energy density influences feed intake (FI) and development of layer-type pullets. A total of 384 nine-wk-old Hy-Line Brown pullets were randomly assigned to one of 3 dietary treatments: fed a diet with 2,600, 2,750, and 2,900 Kcal metabolizable energy/kg (ME/kg) from 10 to 21 wk of age. The results showed that the 2,900 and 2,600 ME groups had lower feed and ME intake (P < 0.01) from 10 to 21 wk of age. The 2,600 ME pullets had heavier body weight (BW) and longer shank length (P < 0.05) at 21 wk of age than the 2,750 ME group. The eggshell percentage was increased by the 2,600 and 2,900 kcal/kg treatments (P = 0.002). Serum concentration of 17-ß-estradiol (E2), follicle-stimulating hormone (FSH), and luteinizing hormone (LH) decreased at 70 wk of age (P < 0.05). Pullet diet and its interaction with age had a significant influence (P < 0.001) on the expression of gonadotropin-releasing hormone 1 (GnRH-1) and gonadotropin-inhibitory hormone (GnIH) in the hypothalamus and of gonadotropin releasing hormone 1 receptor (GnRH-1R) and gonadotropin-inhibitory hormone receptor (GnIHR) in the pituitary. In the hypothalamus, GnRH-1 expression increased from 9 to 40 wk of age and then decreased; however, GnIH expression was highest at 70 wk of age. Follicle-stimulating hormone receptor (FSHR) expression increased (P < 0.001) at wk 40 and decreased at wk 70 compared to wk 21 at various follicular stages. In conclusion, the energy level of pullet diet had no unfavorable influence on feed intake, laying rate, egg mass, and FCR, whereas change egg weight and mortality during the laying period from 21 to 70 wk of age. during the laying period. These results suggest that pullet dietary energy can activate the expression of genes related to reproduction in the hypothalamus, whereas it plays a minor role in the regulation of genes in the pituitary and ovary. Age-induced gene expression in the hypothalamus-pituitary-gonadal (HPG) axis is associated with laying performance in hens.

Allicin Promotes Glucose Uptake by Activating AMPK through CSE/H2S-Induced S-Sulfhydration in a Muscle-Fiber Dependent Way in Broiler Chickens.

SCOPE: Allicin, a product of enzymatic reaction when garlic is injured, plays an important role in maintaining glucose homeostasis in mammals. However, the effect of allicin on glucose homeostasis in the state of insulin resistance remains to be elucidated. This study investigates the effect of allicin on glucose metabolism using different muscle fibers in a chicken model. METHODS AND RESULTS: Day-old male Arbor Acres broilers are randomly divided into three groups and fed a basal diet supplemented with 0, 150, or 300 mg kg-1 allicin for 42 days. Results show that allicin improves the zootechnical performance of broilers at the finishing stage. The glucose loading test (2 g kg-1 body mass) indicates the regulatory role of allicin on glucose homeostasis. In vitro results demonstrate allicin increases glutathione (GSH) level and the expression of cystathionine γ lyase (CSE), leading to endogenous hydrogen sulfide (H2S) production in M. pectoralis major (PM) muscle-derived myotubes. Allicin stimulates adenosine monophosphate-activated protein kinase (AMPK) S-sulfhydration and AMPK phosphorylation to promote glucose uptake, which is suppressed in the presence of d,l-propargylglycine (PAG, a CSE inhibitor). CONCLUSION: This study demonstrates that allicin induces AMPK S-sulfhydration and AMPK phosphorylation to promote glucose uptake via the CSE/H2S system in a muscle fiber-dependent manner.

Prevotella and succinate treatments altered gut microbiota, increased laying performance, and suppressed hepatic lipid accumulation in laying hens.

BACKGROUND: This work aimed to investigate the potential benefits of administering Prevotella and its primary metabolite succinate on performance, hepatic lipid accumulation and gut microbiota in laying hens. RESULTS: One hundred and fifty 58-week-old Hyline Brown laying hens, with laying rate below 80% and plasma triglyceride (TG) exceeding 5 mmol/L, were used in this study. The hens were randomly allocated into 5 groups and subjected to one of the following treatments: fed with a basal diet (negative control, NC), oral gavage of 3 mL/hen saline every other day (positive control, PC), gavage of 3 mL/hen Prevotella melaninogenica (107 CFU/mL, PM) or 3 mL/hen Prevotella copri (107 CFU/mL, P. copri) every other day, and basal diet supplemented with 0.25% sodium succinate (Succinate). The results showed that PM and P. copri treatments significantly improved laying rate compared to the PC (P < 0.05). The amount of lipid droplet was notably decreased by PM, P. copri, and Succinate treatments at week 4 and decreased by P. copri at week 8 (P < 0.05). Correspondingly, the plasma TG level in Succinate group was lower than that of PC (P < 0.05). Hepatic TG content, however, was not significantly influenced at week 4 and 8 (P > 0.05). PM treatment increased (P < 0.05) the mRNA levels of genes PGC-1ß and APB-5B at week 4, and ACC and CPT-1 at week 8. The results indicated enhanced antioxidant activities at week 8, as evidenced by reduced hepatic malondialdehyde (MDA) level and improved antioxidant enzymes activities in PM and Succinate groups (P < 0.05). Supplementing with Prevotella or succinate can alter the cecal microbiota. Specifically, the abundance of Prevotella in the Succinate group was significantly higher than that in the other 4 groups at the family and genus levels (P < 0.05). CONCLUSIONS: Oral intake of Prevotella and dietary supplementation of succinate can ameliorate lipid metabolism of laying hens. The beneficial effect of Prevotella is consistent across different species. The finding highlights that succinate, the primary metabolite of Prevotella, represents a more feasible feed additive for alleviating fatty liver in laying hens.

Research Note: Effect of fasting time on the fasting heat production, blood metabolites, and body components of layer-type pullets.

Fasting heat production (FHP) is used to assess the maintenance net energy requirement of animals. Herein, the FHP of layer-type pullets was estimated. In trial 1, 16 40-day-old Jingfen layer-type pullets were divided into 4 groups of 4 chickens and placed in 4 respiratory chambers. Pullets had free access to feed and water. After 4-d acclimatization, feed was withdrawn, and chickens were measured for FHP for 3 consecutive days. In trial 2, twenty-four 40-day-old pullets were placed in 4 respiratory calorimetry chambers, with 6 pullets per chamber. After 4-d acclimatization, one chamber was randomly selected and all pullets in the chamber was sampled at 5, 25, 50, or 65 h after feed withdrawal. The result showed that FHP declined with fasting time and reached the lowest level between 48 and 72 h. Respiratory quotient was decreased (P < 0.05) between 24 and 48 h compared with that in the first 24 h after fasting. The FHP in the light period showed a significant to decline with fasting time (P < 0.01), whereas the FHP in the dark period was decreased (P < 0.01) 24 h after fasting. Body weight, thigh mass, and abdominal fat decreased (P < 0.05) at 25 h after fasting. Serum glucose were increased (P < 0.01) and while triglycerides were significantly decreased (P < 0.01) at 50 h compared with that at 5 and 25 h time point. The result suggests that the adequate measuring period for FHP for layer-type pullets is from 24 to 48 h after fasting. The FHP of 7-wk-old layer-type pullets was 562.20 kJ/kg of BW0.75/d under a 10-h light and 14-h dark lighting regime.

Daily feeding frequency affects feed intake and body weight management of growing layers.

The objective of this study was to investigate the effect of feeding behavior on feed intake and body weight in growing layers and the underlying mechanisms, thereby providing a scientific foundation for optimal feeding practices in growing layers' management. A total of 144 Hy-line brown growing layers of 10 wk old and similar body weight, were divided into 3 treatment groups with different feeding frequency and equal cumulative daily feeding amount: the once-a-day feeding group (F1) was fed at 9:00 am every day, with feeding amount of 150 g/layer; the twice-a-day feeding group (F2) were fed at 9:00 am and 13:00 pm every day, with each feeding amount of 75 g/layer; the 4 times-a-day feeding group (F4) were fed at 9:00 am, 11:00 am, 13:00 pm, and 15:00 pm every day, with each feeding amount of 37.5 g/layer. Pre-experiment lasted for 1 wk and formal experiment lasted for 8 wk. The results indicated that the daily feed intake and body weight were decreased (P < 0.05) while feed conversion ratio was not affected (P > 0.05) as daily feeding times increased. The glandular stomach proportion was significantly increased in twice-a-day feeding group, while liver proportion and ileum length were significantly increased in 4 times-feeding group (P < 0.05). Additionally, 4 times-feeding daily resulted in a significant elevation of blood glucose levels, which may have suppressed feed intake (P < 0.05). In 4 times-feeding group, the plasma triglyceride levels increased as feeding times, accompanied by a notable up-regulation in the mRNA level of appetite-suppressing gene, hypothalamic pro-opiomelanocortin (POMC) and glandular stomach ghrelin. This modulation effectively suppressed the subsequent feed intake and body weight. Therefore, 4 times feeding daily is recommended in growing layers' management, because it reduced the feed cost without affecting the feed conversion efficiency.