Showing papers by "Lei Sun published in 2019"

Mitochondrial functionality modifies human sperm acrosin activity, acrosome reaction capability and chromatin integrity

Abstract: Study question In addition to sperm motility, which major biological characteristics of sperm fertility potential are associated with mitochondrial functionality? Summary answer Sperm fertilization capacities, including acrosin activity, acrosome reaction (AR) capability and chromatin integrity, are related to the mitochondria functionality as evaluated by the mitochondrial membrane potential (MMP). What is known already Correlative studies suggest a potential role of sperm MMP in predicting sperm fertilization ability and ensuring sperm motility. However, researches characterizing other determinants of sperm fertility potential according to MMP are lacking. Study design, size, duration The sperm MMP was examined in 627 young college students in the Male Reproductive Health in Chongqing College Students (MARHCS) cohort study in 2014. Among these participants, acrosin activity and chromatin integrity were measured in 378 and 604 subjects, respectively. These two determinants of sperm fertility potential were first compared among high-, moderate- and low-MMP groups in the college population. The effects of MMP collapse caused by carbonyl cyanide 3-chlorophenylhydrazone (CCCP) on acrosin activity, AR, DNA fragmentation, reactive oxygen species (ROS) production, and ATP content in human spermatozoa were evaluated in vitro. Participants/materials, setting, methods The sperm MMP was evaluated by using JC-1 staining, acrosin activity was measured using a N-α-benzoyl-dl-arginine-para-nitroanilide HCl (BAPNA) substrate method, the integrity of chromatin represented by DNA fragmentation index (DFI) was measured by sperm chromatin structure assay (SCSA), AR was evaluated with chlortetracycline staining, and intracellular ROS production was evaluated with dihydroethidium. ATP concentration was determined with luciferase. Measurements were performed by spectrophotometry or flow cytometry. Main results and the role of chance Nonparametric analysis revealed significantly higher acrosin activity and a lower DFI in subjects with moderate or high MMP compared to those with low MMP. After adjustment for potential confounders, increases of 7.9 and 44.4% in sperm acrosin activity and deceases of 12.0 and 25.2% in the sperm DFI were found in the moderate- and high-MMP groups, respectively. The MMP dissipation induced by CCCP caused significant declines in acrosin activity and AR capacity and increased DFI in human spermatozoa. Moreover, sperm MMP dissipation induced ROS overproduction and decreased ATP content. Limitations, reasons for caution We cannot exclude a contribution of leukocytes to ROS production and no size gating was used to exclude these cells from the FACS measurements. No simultaneous live-dead staining was done and a contribution of dead sperm to the MMP and acrosome assays cannot be excluded. Wider implications of the findings Mitochondrial functionality might be necessary to maintain sperm acrosin activity, AR and chromatin integrity. Tests of mitochondrial functionality should be developed and used independently of or in addition to conventional semen parameters in infertility diagnosis or risk-assessment processes. Study funding/competing interest(s) This study was supported by the Key Program of the National Natural Science Foundation of China (No. 81630087) and the National Natural Science Foundation of China (No. 81703254). None of the authors have any competing interests to declare.

Mechanisms of Stress-Induced Spermatogenesis Impairment in Male Rats Following Unpredictable Chronic Mild Stress (uCMS)

Abstract: The negative association between psychological stress and male fertility has been known for many years. This study was aimed at (i) identifying spermatogenesis impairment induced by psychological stress in rats and (ii) exploring the role of glucocorticoid receptor (GR) signaling in these adverse effects (if they exist). Male Sprague Dawley rats were exposed to a six-week period of unpredictable chronic mild stress (uCMS) along with cotreatment of GR antagonist RU486 (1 mg/kg/day). Testicular damage was assessed by testicular pathological evaluation, epididymal sperm concentration, serum testosterone levels, testicular apoptotic cell measurements, and cell cycle progression analyses. Rats in the uCMS group had decreased levels of serum testosterone and decreased epididymal sperm concentration. The uCMS-treated rats also had decreased numbers of spermatids and increased levels of apoptotic seminiferous tubules; additionally, cell cycle progression of spermatogonia was arrested at the G0/G1 phase. Furthermore, uCMS exposure caused an increase in serum corticosterone level and activated GR signaling in the testes including upregulated GR expression. RU486 treatment suppressed GR signaling and alleviated the damaging effects of stress, resulting in an increased epididymal sperm concentration. Overall, this work demonstrated for the first time that the activation of GR signaling mediates stress-induced spermatogenesis impairment and that this outcome is related to cell apoptosis and cell cycle arrest in germ cells.

Kisspeptin Protein in Seminal Plasma Is Positively Associated with Semen Quality: Results from the MARHCS Study in Chongqing, China.

Abstract: Objectives. To study the associations between kisspeptin levels in seminal plasma and blood plasma and semen quality. Materials and Methods. We conducted a male reproductive health survey in June 2014. A total of 666 volunteers were recruited from colleges in Chongqing, China. All volunteers completed a questionnaire including information on domestic characteristics and some potential confounders. We tested the kisspeptin levels in both blood and seminal plasma. Total seminal kisspeptin was calculated as the concentration of kisspeptin in seminal plasma multiplied by semen volume. Semen samples were tested according to the 2010 World Health Organization’s (WHO) guidelines. Spearman correlation and multivariate linear regression were used to explore the association between kisspeptin concentrations in seminal plasma and blood plasma and semen quality. Potential confounders that were adjusted for included age, abstinence time, body mass index (BMI), grade, and smoking. Results. The median of kisspeptin levels in seminal plasma was 60,000 times higher than kisspeptin in blood plasma (28.0 × 106 pg/ml versus 448.9 pg/ml). Each interquartile range (IQR) of kisspeptin in seminal plasma was associated with a 4.6% (95% confidence interval [CI]: 1.6%–7.6%) increase in sperm concentration. Each IQR of total kisspeptin was associated with a 7.7% (95% CI: 4.4%–11.0%) increase in total sperm number and a 7.8% (95% CI: 4.0%–11.7%) increase in total motile sperm count. Kisspeptin levels were further classified into quartiles and Q1 was set as the reference level. Subjects in the high total kisspeptin group had 57.5% (95% CI: 33.2%–86.2%) higher total sperm number than the reference group. Conclusion. The positive association between kisspeptin levels in seminal plasma and semen quality supported an important role for the KISS1/GPR54 system in male reproductive health. Kisspeptin may be a potential marker of male reproductive health and an alternative strategy for treating infertility.

Genome Sequencing Illustrates the Genetic Basis of the Pharmacological Properties of Gloeostereum incarnatum.

Abstract: Gloeostereum incarnatum is a precious edible mushroom that is widely grown in Asia and known for its useful medicinal properties. Here, we present a high-quality genome of G. incarnatum using the single-molecule real-time (SMRT) sequencing platform. The G. incarnatum genome, which is the first complete genome to be sequenced in the family Cyphellaceae, was 38.67 Mbp, with an N50 of 3.5 Mbp, encoding 15,251 proteins. Based on our phylogenetic analysis, the Cyphellaceae diverged ~174 million years ago. Several genes and gene clusters associated with lignocellulose degradation, secondary metabolites, and polysaccharide biosynthesis were identified in G. incarnatum, and compared with other medicinal mushrooms. In particular, we identified two terpenoid-associated gene clusters, each containing a gene encoding a sesterterpenoid synthase adjacent to a gene encoding a cytochrome P450 enzyme. These clusters might participate in the biosynthesis of incarnal, a known bioactive sesterterpenoid produced by G. incarnatum. Through a transcriptomic analysis comparing the G. incarnatum mycelium and fruiting body, we also demonstrated that the genes associated with terpenoid biosynthesis were generally upregulated in the mycelium, while those associated with polysaccharide biosynthesis were generally upregulated in the fruiting body. This study provides insights into the genetic basis of the medicinal properties of G. incarnatum, laying a framework for future characterization of bioactive proteins and pharmaceutical uses of this fungus.

Pleurotus eryngii Genomes Reveal Evolution and Adaptation to the Gobi Desert Environment.

Abstract: Pleurotus eryngii (King Oyster) is one of the most highly prized edible mushrooms. Among the diverse varieties within P. eryngii, P. eryngii var. eryngii is the commonest one, with a worldwide distribution, while P. eryngii var. ferulae is only distributed in Europe and China, and is especially adapted to the Gobi Desert in Xinjiang Autonomous Region of China. However, little is known about the genome-wide pattern of evolution and adaptation to the divergent environments of P. eryngii. Here, we present the high-quality genome sequences of P. eryngii var. eryngii strain PEE81 originating from Europe and P. eryngii var. ferulae strain PEF12 originating from the Gobi Desert of China. The assembled genome sizes of PEE81 and PEF12 were 53.6 and 48.0 Mbp, respectively, which are larger than other reported genomes in the genus Pleurotus. We propose that the selective amplification of long terminal repeat (LTR) retrotransposons increases the genome size of the genus Pleurotus, and may play a key role in driving their rapid species diversification. Molecular clock analyses of five Pleurotus species, namely PEE81, PEF12, P. tuoliensis, P. ostreatus and P. cf. floridanus suggest that the divergence estimates of the genus Pleurotus over time scales ranged from ∼4 to ∼38 million years ago (Mya), and PEE81 and PEF12 diverged at ∼13 Mya. The whole genome resequencing of 33 geographically diverse strains of P. eryngii var. eryngii and var. ferulae was then performed and the genome variation among and within these two populations were investigated. Comparative analyses of these two populations detected several candidate genes related to stress responses and DNA repair that are putatively involved in adaptation to the Gobi Desert environment. These findings offer insights into genome evolution of the genus Pleurotus and provide valuable genomic resources for King Oyster mushroom breeding.

Genomic Analyses Reveal Evidence of Independent Evolution, Demographic History, and Extreme Environment Adaptation of Tibetan Plateau Agaricus bisporus

Abstract: Agaricus bisporus distributed in the Tibetan Plateau of China has high-stress resistance that is valuable for breeding improvements. However, its evolutionary history, specialization, and adaptation to the extreme Tibetan Plateau environment are largely unknown. Here, we performed de novo genome sequencing of a representative Tibetan Plateau wild strain ABM and comparative genomic analysis with the reported European strain H97 and H39. The assembled ABM genome was 30.4 Mb in size, and comprised 8,562 protein-coding genes. The ABM genome shared highly conserved syntenic blocks and a few inversions with H97 and H39. The phylogenetic tree constructed by 1,276 single-copy orthologous genes in nine fungal species showed that the Tibetan Plateau and European A. bisporus diverged ∼5.5 million years ago. Population genomic analysis using genome resequencing of 29 strains revealed that the Tibetan Plateau population underwent significant differentiation from the European and American populations and evolved independently, and the global climate changes critically shaped the demographic history of the Tibetan Plateau population. Moreover, we identified key genes that are related to the cell wall and membrane system, and the development and defense systems regulated A. bisporus adapting to the harsh Tibetan Plateau environment. These findings highlight the value of genomic data in assessing the evolution and adaptation of mushrooms and will enhance future genetic improvements of A. bisporus.

Genome Analysis of Hypomyces perniciosus, the Causal Agent of Wet Bubble Disease of Button Mushroom (Agaricus bisporus).

Abstract: The mycoparasitic fungus Hypomyces perniciosus causes wet bubble disease of mushrooms, particularly Agaricus bisporus. The genome of a highly virulent strain of H. perniciosus HP10 was sequenced and compared to three other fungi from the order Hypocreales that cause disease on A. bisporus. H. perniciosus genome is ~44 Mb, encodes 10,077 genes and enriched with transposable elements up to 25.3%. Phylogenetic analysis revealed that H. perniciosus is closely related to Cladobotryum protrusum and diverged from their common ancestor ~156.7 million years ago. H. perniciosus has few secreted proteins compared to C. protrusum and Trichoderma virens, but significantly expanded protein families of transporters, protein kinases, CAZymes (GH 18), peptidases, cytochrome P450, and SMs that are essential for mycoparasitism and adaptation to harsh environments. This study provides insights into H. perniciosus evolution and pathogenesis and will contribute to the development of effective disease management strategies to control wet bubble disease.

Genome Sequencing of Cladobotryum protrusum Provides Insights into the Evolution and Pathogenic Mechanisms of the Cobweb Disease Pathogen on Cultivated Mushroom.

Abstract: Cladobotryum protrusum is one of the mycoparasites that cause cobweb disease on cultivated edible mushrooms. However, the molecular mechanisms of evolution and pathogenesis of C. protrusum on mushrooms are largely unknown. Here, we report a high-quality genome sequence of C. protrusum using the single-molecule, real-time sequencing platform of PacBio and perform a comparative analysis with closely related fungi in the family Hypocreaceae. The C. protrusum genome, the first complete genome to be sequenced in the genus Cladobotryum, is 39.09 Mb long, with an N50 of 4.97 Mb, encoding 11,003 proteins. The phylogenomic analysis confirmed its inclusion in Hypocreaceae, with its evolutionary divergence time estimated to be ~170.1 million years ago. The genome encodes a large and diverse set of genes involved in secreted peptidases, carbohydrate-active enzymes, cytochrome P450 enzymes, pathogen–host interactions, mycotoxins, and pigments. Moreover, C. protrusum harbors arrays of genes with the potential to produce bioactive secondary metabolites and stress response-related proteins that are significant for adaptation to hostile environments. Knowledge of the genome will foster a better understanding of the biology of C. protrusum and mycoparasitism in general, as well as help with the development of effective disease control strategies to minimize economic losses from cobweb disease in cultivated edible mushrooms.

Genomic Analyses Provide Insights Into the Evolutionary History and Genetic Diversity of Auricularia Species.

Abstract: Species in the genus Auricularia play important roles for people’s food and nutrition especially Auricularia cornea and A. heimuer. To understand their evolutionary history, genome structure, and population-level genetic variation, we performed a high-quality genome sequencing of Auricularia cornea and the corresponding comparative genomic analysis. The genome size of A. cornea was similar to Auricularia subglabra, but 1.5 times larger than that of A. heimuer. Several factors were responsible for genome size variation including gene numbers, repetitive elements, and gene lengths. Phylogenomic analysis revealed that the estimated divergence time between A. heimuer and other Auricularia is ∼79.1 million years ago (Mya), while the divergence between A. cornea and A. subglabra occurred in ∼54.8 Mya. Population genomic analysis also provided insight into the demographic history of A. cornea and A. heimuer, indicating that their populations fluctuated over time with global climate change during Marine Isotope Stage 5-2. Moreover, despite the highly similar external morphologies of A. cornea and A. heimuer, their genomic properties were remarkably different. The A. cornea genome only shared 14% homologous syntenic blocks with A. heimuer and possessed more genes encoding carbohydrate-active enzymes and secondary metabolite biosynthesis proteins. The cross-taxa transferability rates of simple sequence repeat (SSR) and insertion or deletion (InDel) markers within the genus Auricularia were also lower than that previously observed for species within the same genus. Taken together, these results indicate a high level of genetic differentiation between these two Auricularia species. Consequently, our study provides new insights into the genomic evolution and genetic differentiation of Auricularia species that will facilitate future genetic breeding.

A Comparison of the Physical, Chemical, and Structural Properties of Wild and Commercial Strains of Button Mushroom, Agaricus bisporus (Agaricomycetes).

Abstract: Agaricus bisporus is well known for its nutraceutical properties. To meet consumer market demand, there is an urgent need for new strains with disease resistance and a diverse nutrient profile for commercial cultivation. Wild germplasm resources provide a good source for the breeding of new strains for this purpose. In this study, we evaluated the physical, chemical, and structural properties of wild domesticated (CCMJ1351) and major commercially cultivated strains (CCMJ1013, CCMJ1028, and CCMJ1343) of A. bisporus from China. The results showed significant differences among the strains for all parameters measured. In terms of morphological characteristics, CCMJ1351 possessed the highest stipe thickness, fruiting body individual weight, cohesiveness, and springiness; CCMJ1013 demonstrated maximum pileus diameter and thickness; CCMJ1028 exhibited the highest textural hardness and color characteristics; and strain CCMJ1343 had the highest yield. CCMJ1351 ranked top among all the strains for proximate composition, rheological profile, and structural and mechanical properties, containing 21.93% crude protein and the highest dry matter, crude fat, and fiber contents. However, the bioactive chemical constituents present in the four strains were very similar, especially β-(1→3)-glucan, according to Fourier transform infrared spectroscopy analysis, while some minimal peaks varied among the different strains. Therefore, in combination with previously identified high disease-resistance traits, the wild domesticated strain CCMJ1351 constitutes a good candidate for further exploitation in breeding programs and is suitable for fresh consumption as well as incorporation into various food products.

Rat noise and carbon monoxide integrated combination exposure contamination experiment device

Abstract: The invention provides a rat noise and carbon monoxide integrated combination exposure contamination experiment device to overcome the shortcoming that an existing device cannot meet simultaneous exposure contamination of carbon monoxide and noise. A gas blending chamber and a noise generation chamber provided with a Bluetooth speaker are assembled and then embedded into a combination exposure cabin, a rat is placed into a mouth and nose contamination cup through a door, the carbon monoxide is led into the gas blending chamber through a gas inlet, and the noise enters the combination exposure cabin through a noise transmission hole. As the mouth and nose contamination cup is communicated with the combination exposure cabin, the rat in the mouth and nose contamination cup can be simultaneously subjected to combination exposure to the carbon monoxide and the noise. The device can meet the experimental requirement of simultaneous exposure to the carbon monoxide and the noise. In addition, the mouth and nose contamination cup can limit activities of the rat, the true situation of activity limitation of cabin operators can be simulated, and the distance between the ear or noise of the rat and a noise source can be basically the same as that between the ear or noise of the rat and a gas pollution source, and scientificity of experimental results is improved.