Histone H3K79 methylation by DOT1L promotes Aurora B localization at centromeres in mitosis.

Centromere localization of the chromosome passenger complex (CPC) is paramount for achieving accurate sister chromosome segregation in mitosis. Although it has been widely recognized that the recruitment of CPC is directly regulated by two histone codes, phosphorylation of histone H3 at threonine 3 (H3T3ph) and phosphorylation of histone H2A at threonine 120 (H2AT120ph), the regulation of CPC localization by other histone codes remains elusive. We show that dysfunction of disruptor of telomeric silencing 1 like (DOT1L) leads to mislocation of the CPC in prometaphase, caused by disturbing the level of H3T3ph and its reader Survivin. This cascade is initiated by over-dephosphorylation of H3T3ph mediated by the phosphatase RepoMan-PP1, whose scaffold RepoMan translocalizes to chromosomes, while the level of H3K79me2/3 is diminished. Together, our findings uncover a biological function of DOT1L and H3K79 methylation in mitosis and give insight into how genomic stability is coordinated by different histone codes.

Heavy metal ions exchange driven protein phosphorylation cascade functions in genomic instability in spermatocytes and male infertility.

DNA double-strand breaks (DSBs) are functionally linked to genomic instability in spermatocytes and to male infertility. The heavy metal cadmium (Cd) is known to induce DNA damage in spermatocytes by unknown mechanisms. Here, we showed that Cd ions impaired the canonical non-homologous end-joining (NHEJ) repair pathway, but not the homologous recombination (HR) repair pathway, through stimulation of Ser2056 and Thr2609 phosphorylation of DNA-PKcs at DSB sites. Hyper-phosphorylation of DNA-PKcs led to its premature dissociation from DNA ends and the Ku complex, preventing recruitment of processing enzymes and further ligation of DNA ends. Specifically, this cascade was initiated by the loss of PP5 phosphatase activity, which results from the dissociation of PP5 from its activating ions (Mn), that is antagonized by Cd ions through a competitive mechanism. In accordance, in a mouse model Cd-induced genomic instability and consequential male reproductive dysfunction were effectively reversed by a high dosage of Mn ions. Together, our findings corroborate a protein phosphorylation-mediated genomic instability pathway in spermatocytes that is triggered by exchange of heavy metal ions.

SNP rs12794714 of CYP2R1 is associated with serum vitamin D levels and recurrent spontaneous abortion (RSA): a case-control study.

PURPOSE: Vitamin D (VD) deficiency seems to be associated with the risk of recurrent spontaneous abortion (RSA). Vitamin D receptor (VDR) and cytochrome P450 family 2 subfamily R member 1 (CYP2R1) are two genes which are vital for VD metabolism and actions. However, whether single-nucleotide polymorphisms (SNPs) in these genes are correlated with the risk of RSA are poorly understood. Therefore, we aimed to characterize the relationships among VDR SNPs, CYP2R1 SNPs and RSA. METHODS: This case-control study enrolled 75 RSA patients and 83 controls. Serum VD and some cytokines were detected with LC-MS/MS and flow cytometry, respectively. Genotyping for three SNPs of CYP2R1 (rs10741657, rs10766197 and rs12794714) and five SNPs of VDR (rs7975232, rs1544410, rs2189480, rs2228570 and rs2239179) was done with polymerase chain reaction (PCR) and high-throughput sequencing. All the data were analyzed with appropriate methods and in different models. RESULTS: The results revealed a significant correlation between the AG genotype of CYP2R1 rs12794714 and VD levels (OR 0.686; 95% CI 0.49-0.96; p = 0.028). Besides, the AG and GG genotypes of CYP2R1 rs12794714 were markedly related to the risk of RSA (OR 52.394, 59.497; 95% CI 2.683-1023.265, 3.110-1138.367; p = 0.009, 0.007, respectively). CONCLUSION: Our results indicate that CYP2R1 rs12794714 might be a risk factor for RSA. Hence, early screening of pregnant women for CYP2R1 rs12794714 is necessary to warrant proactive counseling and treatment against RSA.

Dual apodization with cross-correlation combined with robust Capon beamformer applied to ultrasound passive cavitation mapping.

PURPOSE: Passive acoustic mapping (PAM) has received increasing attention in recent years and has an extremely widespread application prospect in real-time monitoring of ultrasound treatment. When using a diagnostic ultrasound transducer, such as a linear-array transducer, the initially used time exposure acoustics (TEA) algorithm will produce high-level artifacts. To address this problem, we recently proposed an enhanced algorithm for linear-array PAM by introducing dual apodization with the cross-correlation (DAX) method into TEA. But due to that the delay and sum beamformer used to create RX1 and RX2 is non-adaptive, the remaining X-type artifacts cannot be completely suppressed, yielding unsatisfactory image quality. This study aims to propose an improved version by combining DAX and robust Capon beamformer (DAX-RCB). METHODS: Different from the delay and sum beamformer in the DAX-TEA algorithm, in the proposed version, the two sets of channel signals from a pair of complementary receive apodizations are beamformed by the RCB method, which may make passive cavitation images much less sensitive to X-type artifacts. The performance of the DAX-RCB algorithm is validated by simulations and in vitro experiments and compared with the initially used TEA algorithm and the previous DAX-TEA and RCB algorithms. Four indexes, including passive energy beam (PEB) size, image signal-to-background ratio (ISBR), energy estimation ratio (EER), and computing time, are used to evaluate the algorithm performance. RESULTS: Consider an example of the 8-8 alternating pattern (a pair of complementary apodizations are obtained by extracting eight elements every eight elements), the experimental results show that the A-6dB area (2D PEB size) of the proposed DAX-RCB is significantly reduced by 11.0 and 6.8 mm2 when compared with TEA and DAX-TEA and is not significantly reduced when compared with RCB, the ISBR is significantly improved by 19.6, 10.8, and 5.6 dB compared with TEA, DAX-TEA, and RCB, and the EER of DAX-RCB is over 90%. The simulation tests indicate that the DAX-RCB algorithm is also applicable to the image enhancement in the double-source scenario and the high-level noise scenario but at a risk of low energy estimation. The improvement of algorithm performance is accompanied by an increase in the computing time. The proposed DAX-RCB consumes 113.3%, 29.5%, and 17.8% more time than TEA, DAX-TEA, and RCB. CONCLUSIONS: The proposed DAX-RCB can be considered as an effective reconstruction algorithm for passive cavitation mapping and provide an appropriate monitoring means for ultrasound therapy, especially for cavitation-mediated applications.

LSD1 negatively regulates autophagy in myoblast cells by driving PTEN degradation.

Lysine-specific demethylase 1 (LSD1) is a well characterized transcriptional regulator functioning on the chromatin to remove mono- and di-methyl groups from lysine 4 or lysine 9 of histone 3 (H3K4 or H3K9). LSD1 also has non-transcriptional activities via targeting non-histone substrates that participate in diverse biological processes. In this report, we determined that LSD1 negatively regulates autophagy in skeletal muscle cells by promoting PTEN degradation in a transcription-independent mechanism. In C2C12 cells, LSD1 inhibition or depletion significantly induced the initiation of autophagy; and autophagy resulted from LSD1 inhibition is associated with AKT/mTORC1 inactivation. Notably, the proteins of PTEN, a prominent repressive AKT modulator, are stabilized by LSD1 inhibition despite a decrease of its mRNA levels. Further data demonstrated that LSD1 interacts with PTEN protein and enhances its ubiquitination and degradation. Together, our findings identify a novel biological function of LSD1 in autophagy, mediated by regulating the stability of PTEN and the activity of AKT/mTORC1.

The effects of long-term exposure to low doses of cadmium on the health of the next generation of mice.

Cadmium (Cd) is an important toxic chemical due to its increasing levels in the environment and bioaccumulation in humans and animals. The present study was performed to evaluate the effects of long-term exposure to 1, 10, or 100 µg/L Cd in drinking water on the development, reproduction and neurotoxicity of offspring when administered to mice from parental puberty to postnatal 10 weeks in offspring. The development parameters measured in offspring included physical development, reflex ontogeny, body weight and body size. The reproductive indices measured consisted of anogenital distances (AGDs), estrous cycle, sperm quality, specific gene expression in Leydig or Sertoli cells, seminiferous epithelium cycle, sex hormone levels, histological morphology and apoptosis in testis or ovary, and the levels of oxidative stress. The determination of neurotoxicity included learning and memory ability, anxiety, and related serum indicators. In addition, blood lipid level, liver and kidney function were also determined by serum biochemical assays. The results showed that exposure to Cd in the present model had no adverse effects on development, but had some reproductive toxicity and neurotoxicity, including alteration of spermatogenic epithelial staging in testis and inducing anxiety in offspring. Furthermore, the levels of total protein, globulins, total bile acid and direct bilirubin were also significantly altered, especially in female offspring. The present study suggested that long-term exposure to low doses of Cd had adverse effects on the health of the next generation, and some harmful effects showed gender differences in offspring. The present study demonstrated that attention should be paid to Cd pollution in the environment, especially before pregnancy.

Delay multiply and sum beamforming method applied to enhance linear-array passive acoustic mapping of ultrasound cavitation.

PURPOSE: Passive acoustic mapping (PAM) has been proposed as a means of monitoring ultrasound therapy, particularly nonthermal cavitation-mediated applications. In PAM, the most common beamforming algorithm is a delay, sum, and integrate (DSAI) approach. However, using DSAI leads to low-quality images for the case where a narrow-aperture receiving array such as a standard B-mode linear array is used. This study aims to propose an enhanced linear-array PAM algorithm based on delay, multiply, sum, and integrate (DMSAI). METHODS: In the proposed algorithm, before summation, the delayed signals are combinatorially coupled and multiplied, which means that the beamformed output of the proposed algorithm is the spatial coherence of received acoustic emissions. We tested the performance of the proposed DMSAI using both simulated and experimental data and compared it with DSAI. The reconstructed cavitation images were evaluated quantitatively by using source location errors between the two algorithms, full width at half maximum (FWHM), size of point spread function (A50 area), signal-to-noise ratio (SNR), and computational time. RESULTS: The results of simulations and experiments for single cavitation source show that, by introducing DMSAI, the FWHM and the A50 area are reduced and the SNR is improved compared with those obtained by DSAI. The simulation results for two symmetric or nonsymmetric cavitation sources and multiple cavitation sources show that DMSAI can significantly reduce the A50 area and improve the SNR, therefore improving the detectability of multiple cavitation sources. CONCLUSIONS: The results indicate that the proposed DMSAI algorithm outperforms the conventionally used DSAI algorithm. This work may have the potential of providing an appropriate method for ultrasound therapy monitoring.

Passive cavitation mapping using dual apodization with cross-correlation in ultrasound therapy monitoring.

Recently, passive acoustic mapping (PAM) has been successfully applied for dynamic monitoring of ultrasound therapy by beamforming acoustic emissions of cavitation activity during ultrasound exposure. The most widely used PAM algorithm in the literature is time exposure acoustics (TEA), which is a standard delay, sum, and integrate algorithm. However, it results in large point spread function (PSF) and serious imaging artifacts for the case where a narrow-aperture receiving array such as a standard B-mode linear array is used, therefore degrading the quality of cavitation image. To address these challenges, in this paper, we proposed a novel PAM algorithm namely dual apodization with cross-correlation (DAX)-based TEA, in which DAX was originally used as a reconstruction algorithm in medical ultrasound imaging. In the proposed algorithm, two sets of signals were beamformed by two receive apodization functions with alternating elements enabled, and the cross-correlation coefficient of the two signals served as a weighting factor that would be multiplied to the sum of the two signals. The performance of the proposed algorithm was tested on simulated channel data obtained using a multi-bubble model, and experiments were also performed in an in vitro vessel phantom with flowing microbubbles as cavitation nuclei. The reconstructed cavitation images were evaluated quantitatively using established quality metrics including full width at half maximum (FWHM), A-6dB area, and signal-to-noise ratio (SNR). The results suggested that the proposed algorithm significantly outperformed the conventionally used TEA algorithm. This work may have the potential of providing a useful tool for highly accurate localization of cavitation activity during ultrasound therapy.

The Protective Effect of Autophagy on DNA Damage in Mouse Spermatocyte-Derived Cells Exposed to 1800 MHz Radiofrequency Electromagnetic Fields.

BACKGROUND/AIMS: The effects of exposure to radiofrequency electromagnetic fields (RF-EMFs) on the male reproductive system have raised public concern and studies have shown that exposure to RF-EMFs can induce DNA damage and autophagy. However, there are no related reports on the role of autophagy in DNA damage in spermatocytes, especially after exposure to RF-EMFs. The aim of the present study was to determine the mechanism and role of autophagy induced by RF-EMFs in spermatozoa cells. METHODS: Mouse spermatocyte-derived cells (GC-2) were exposed to RF-EMFs 4 W/kg for 24 h. The level of reactive oxygen species (ROS) was determined by ROS assay kit. Comet assay was utilized to detect DNA damage. Autophagy was detected by three indicators: LC3II/LC3I, autophagic vacuoles, and GFP-LC3 dots, which were measured by western blot, transmission electron microscopy, and transfection with GFP-LC3, respectively. The expression of the molecular signaling pathway AMP-activated protein kinase (AMPK)/mTOR was determined by western blot. RESULTS: The results showed that RF-EMFs induced autophagy and DNA damage in GC-2 cells via ROS generation, and the autophagy signaling pathway AMPK/mTOR was activated by ROS generation. Furthermore, following inhibition of autophagy by knockdown of AMPKα, increased DNA damage was observed in GC-2 cells following RF-EMFs exposure, and overexpression of AMPKα promoted autophagy and attenuated DNA damage. CONCLUSIONS: These findings demonstrated that the autophagy which was induced by RF-EMFs via the AMPK/mTOR signaling pathway could prevent DNA damage in spermatozoa cells.

ATM signals to AMPK to promote autophagy and positively regulate DNA damage in response to cadmium-induced ROS in mouse spermatocytes.

Cadmium (Cd) is a toxic heavy metal and harmful to human health due to its ability to accumulate in organs. Previous studies have shown that Cd can induce DNA damage and autophagy. Autophagy can stabilize genetic material and DNA integrity. The aim of the present study was to determine the exact mechanism and role of autophagy induced by Cd in spermatozoa cells. Mouse spermatocyte-derived cells (GC-2) were treated with 20 µM Cd chloride for 24 h. The level of reactive oxygen species (ROS), DNA damage, autophagy and the expression of the molecular signaling pathway ATM/AMP-activated protein kinase (AMPK)/mTOR were determined. The results showed that Cd induced autophagy and DNA damage in GC-2 cells via ROS generation, and the autophagy signal pathway AMPK/mTOR was activated by ATM which is a DNA damage sensor. Melatonin, a well-known antioxidant, ameliorated DNA damage, and inhibited autophagy via the AMPK/mTOR signal pathway. Furthermore, after inhibition of autophagy by knockdown of AMPKα, increased DNA damage by Cd treatment was observed in GC-2 cells. These findings demonstrated the protective role of autophagy in DNA damage and suggested that the mechanism of autophagy induced by Cd was through the ATM/AMPK/mTOR signal pathway in spermatozoa cells.

A Preliminary Study about the Potential Effects of Heavy Metals on the Human Male Reproductive Parameters in HIV-Infected Population in China.

Due to the inconsistent effects of human immunodeficiency virus (HIV) on the human male reproduction in previous studies and the impacts of environmental exposures, such as heavy metals, on male reproduction receiving little attention in HIV-infected population, the aim of present study was to investigate whether heavy metals have potential effects on reproductive parameters in HIV-infected men. The current study assessed the associations between semen quality or serum hormone and concentration of the three heavy metal toxicants (lead (Pb), cadmium (Cd), and zinc (Zn)) in seminal, urine, and serum, and 50 HIV-infected men were recruited in the present study. Concentrations of Pb, Cd, and Zn were measured in three fluids by graphite furnace atomic absorption spectrophotometer. Semen analyses were performed according to World Health Organization criteria. Serum samples were analyzed for follicle-stimulating hormone, luteinizing hormone, and testosterone. HIV RNA viral load was determined by HIV virus loads kit. Spearman's rank correlations were used for correlation analyses. The results showed that the concentrations of Pb, Cd, and Zn were significantly correlated with semen quality and serum hormone. HIV-1 virus loads were significantly associated with increased seminal Pb. However, HIV-1 virus loads were not statistically associated with semen quality and serum hormone. Our findings suggested that environmental heavy metals had potential effects on reproductive parameters in HIV-infected men in China.

Occupational exposure to 50 Hz magnetic fields does not alter responses of inflammatory genes and activation of splenic lymphocytes in mice.

OBJECTIVES: The objective of the present study was to observe the effects of 50 Hz magnetic fields (MFs) on the immune function of splenic lymphocytes in mice. MATERIAL AND METHODS: Twenty male Kunming mice (6 weeks old), weighing 18- 25 g, were randomly divided into sham exposure (N = 10) and 500 µT MFs (N = 10) groups. The mice in the MFs group were exposed to 500 µT MFs for 8 h daily (5 days/week) for up to 60 days. In vitro study was carried out to examine the effects of 50 Hz MFs on the expression of inflammatory factor genes and a cluster of differentiation 69 (CD69) in mouse prime splenic lymphocytes activated by para-Methoxyamphetamine (PMA) and ionomycin. In the in vitro experiments, lymphocytes were isolated from the spleen of 10 healthy Kunming mice, the cells were cultured in the Roswell Park Memorial Institute 1640 medium (RPMI-1640) and exposed to 0 µT, 250 µT, 500 µT, or 1 mT MFs in an incubator under 5% carbon dioxide (CO2) at 37°C for 6 h. The levels of interleukin-2 (IL-2), IL-4, interferon-gamma (IFN-γ), GATA binding protein 3 (GATA-3) and T cell-specific T-box transcription factor (T-bet) were assessed by the real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR), respectively. The expression of CD69 was checked using the flow cytometry. RESULTS: Under our experimental conditions, body weight of the mice exposed to occupational, extremely low frequency- electromagnetic fields (ELF-EMFs) significantly decreased on day 20 and day 30. There were no significant changes observed in vivo in spleen weight, splenic coefficient, splenic histology profile and cytokine production in spleen tissues. Our in vitro experiments showed that 50 Hz MFs had no effect on the expression of these genes and CD69 to primary splenic cells. CONCLUSIONS: In conclusion, under the applied experimental conditions, occupational exposure to 50 Hz magnetic field did not alter responses of inflammatory genes and activation of splenic lymphocytes in mice, except for body weight.

The Protective Effects of Melatonin Against Oxidative Stress and Inflammation Induced by Acute Cadmium Exposure in Mice Testis.

Cadmium (Cd) is widely used in daily life and was recently recognized as a possible source of human toxicity due to its ability to accumulate in organs. Previous studies have shown that Cd exposure may cause testicular toxicity through oxidative stress and an inflammatory effect. Melatonin has been demonstrated to be an effective anti-oxidant and has an anti-inflammatory effect. The aim of the present study was to investigate the toxicological effects of Cd on reproduction in male mice and the potential protective action of melatonin against these adverse effects. Adult male mice were injected intraperitoneally with Cd at a dose of 2 mg/kg body weight per day for seven consecutive days with or without melatonin pretreatment. Sex organ weight, sperm parameters including sperm quality, apoptosis, acrosome integrity, mitochondrial membrane potential, testicular morphology, serum sex hormone, inflammatory status, and oxidative stress were evaluated. The results showed that significant adverse effects were observed in the male reproductive system after Cd exposure, including alterations in sperm parameters, increased DNA damage, and sex hormone disturbance. Acute Cd exposure also significantly increased malondialdehyde (MDA) contents, decreased glutathione (GSH) and superoxide dismutase (SOD) activities, and upregulated levels of the pro-inflammatory cytokines, tumor necrosis factor-alpha (TNF-α), and interleukin-1beta (IL-1ß), in the testis. In contrast, melatonin pretreatment significantly alleviated these toxic effects, and its mechanism may involve inhibiting MDA level, restoring GSH and SOD activities, and reducing the upregulation of TNF-α and IL-1ß. Our data suggest that oxidative stress and inflammation are involved in Cd-induced toxicity in the male reproductive system and that co-administration of melatonin exerts a protective effect against Cd-induced male reproductive toxicity.

Effects of low-dose cadmium exposure during gestation and lactation on development and reproduction in rats.

Cadmium (Cd) is an important toxic chemical due to its increasing levels in the environment and its resulting accumulation in humans and animals. The present study was performed to evaluate the long-term effects of low doses of Cd administered in offspring by oral route to rats during pregnancy and lactation. There were no adverse effects on the physical and sexual development in the pups, except to delay the development of offspring. The relative weights of livers and kidneys in the adult female offspring were significantly decreased after exposure to 10 ppm Cd. These results indicated that there were adverse effects on growth and development from exposure to 5 or 10 ppm Cd in utero and during lactation. The results also showed differential gender sensitivity effects on the organ weights.

Melatonin Improves mitochondrial function by promoting MT1/SIRT1/PGC-1 alpha-dependent mitochondrial biogenesis in cadmium-induced hepatotoxicity in vitro.

Melatonin is an indolamine synthesized in the pineal gland that has a wide range of physiological functions, and it has been under clinical investigation for expanded applications. Increasing evidence demonstrates that melatonin can ameliorate cadmium-induced hepatotoxicity. However, the potentially protective effects of melatonin against cadmium-induced hepatotoxicity and the underlying mechanisms of this protection remain unclear. This study investigates the protective effects of melatonin pretreatment on cadmium-induced hepatotoxicity and elucidates the potential mechanism of melatonin-mediated protection. We exposed HepG2 cells to different concentrations of cadmium chloride (2.5, 5, and 10 µM) for 12 h. We found that Cd stimulated cytotoxicity, disrupted the mitochondrial membrane potential, increased reactive oxygen species production, and decreased mitochondrial mass and mitochondrial DNA content. Consistent with this finding, Cd exposure was associated with decreased Sirtuin 1 (SIRT1) protein expression and activity, thus promoted acetylation of PGC-1 alpha, a key enzyme involved in mitochondrial biogenesis and function, although Cd did not disrupt the interaction between SIRT1 and PGC-1 alpha. However, all cadmium-induced mitochondrial oxidative injuries were efficiently attenuated by melatonin pretreatment. Moreover, Sirtinol and SIRT1 siRNA each blocked the melatonin-mediated elevation in mitochondrial function by inhibiting SIRT1/ PGC-1 alpha signaling. Luzindole, a melatonin receptor antagonist, was found to partially block the ability of melatonin to promote SIRT1/ PGC-1 alpha signaling. In summary, our results indicate that SIRT1 plays an essential role in the ability of moderate melatonin to stimulate PGC-1 alpha and improve mitochondrial biogenesis and function at least partially through melatonin receptors in cadmium-induced hepatotoxicity.

Bisphenol A exposure at an environmentally relevant dose induces meiotic abnormalities in adult male rats.

Whether environmental exposure to bisphenol A (BPA) may induce reproductive disorders is still controversial but certain studies have reported that BPA may cause meiotic abnormalities in C. elegans and female mice. However, little is known about the effect of BPA on meiosis in adult males. To determine whether BPA exposure at an environmentally relevant dose could induce meiotic abnormalities in adult male rats, we exposed 9-week-old male Wistar rats to BPA by gavage at 20 µg/kg body weight (bw)/day for 60 consecutive days. We found that BPA significantly increased the proportion of stage VII seminiferous epithelium and decreased the proportion of stage VIII. Consequently, spermiation was inhibited and spermatogenesis was disrupted. Further investigation revealed that BPA exposure delayed meiosis initiation in the early meiotic stage and induced the accumulation of chromosomal abnormalities and meiotic DNA double-strand breaks (DSBs) in the late meiotic stage. The latter event subsequently activated the phosphatidylinositol 3-kinase-related protein kinase (ATM). Our results suggest that long-term exposure to BPA may lead to continuous meiotic abnormalities and ultimately put mammalian reproductive health at risk.

Analysis of genetic diversity and phylogenetic relationship of red deer subspecies in XinJiang, China.

Polymorphisms for seven microsatellite loci in three red deer subspecies (9 populations) found in XinJiang were detected by polymerase chain reaction (PCR), 12% nondenaturation polyacrylamide gel electrophoresis and the Sanguinetti silver staining method. Numbers of alleles, average effective numbers of alleles (E) and the average rate of homozygosity, allelic frequencies of seven microsatellite loci, polymorphism information content (PIC), mean heterozygosity (H) and genetic distances among the populations were calculated for each population. Dendrograms were constructed based on genetic distances by the neighbor-joining method (NJ), utilizing molecular evolutionary genetics analysis software PHYLIP (3.6). The phylogenetic tree was constructed based on allelic frequencies using maximum likelihood (ML); the bootstrap value was estimated by bootstrap test in the tree. Lastly, phylogenesis was analyzed. The results showed that four of the seven microsatellite loci were highly polymorphic, but BMS2508 and Celjp0023 showed no polymorphism and BM5004 was a neutral polymorphism. It is our conclusion that the four microsatellite loci are effective DNA markers for the analysis of genetic diversity and phylogenetic relationships among the three red deer subspecies. The mean PIC, H and E-values across the microsatellite loci were 0.5393, 0.5736 and 2.64, which showed that these microsatellite loci are effective DNA markers for the genetic analysis of red deer. C.e. songaricus populations from Regiment 104, 151 and Hami are clustered together. C.e. yarkandensis populations from Regiment 35, Xaya and Alaer are clustered together. These two clusters also cluster together. Lastly, C.e. sibiricus populations from Burqin, Regiment 188 and the first two clusters were clustered together. The phylogenetic relationship among different red deer populations is consistent with the known origin, history of breeding and geographic distributions of populations.

Sex control by Zfy siRNA in the mouse.

The objective of this work was to detect the influence of Y sperm forming of Mus musculus by silencing the Zfy gene during spermatogenesis. The recombination expression vectors pSilencer5.1/Zfy215 and pSilencer5.1/Zfy2102 were constructed. 64 male KunMing Mus were divided into four groups randomly and averagely. The two recombination expression vectors were injected into two groups, respectively, through testis. The other two groups were injected with the same volume of physiological saline and empty vector pSilencer5.1-H1 Retro, respectively. They were injected every ten days for a total of four injections. Seventeen days after the fourth injection, 8 male Mus of each group mated with 8 female Mus. The testis tissue of the other 8 male Mus of each group was collected, and the expression level of Zfy mRNA was determined by fluorescence quantitation real time PCR (qRT-PCR). The result showed that the expression of Zfy mRNA decreased significantly after injection of pSilencer5.1/Zfy2102 (P < 0.01), and that 72.3% of the offspring were female, a number significantly higher than in the control group (P < 0.01). In the pSilencer5.1/Zfy215 group, the expression of Zfy mRNA was significantly lower than in the control group (P < 0.05), but the female rate of offspring was not. It was concluded that the Zfy gene could play a role in the process of Y sperm formation.

Antibody and cytokine responses to hydatid in experimentally infected Kazakh sheep with hydatidosis resistance haplotype.

Different MHC haplotype of Kazakh sheep has different resistance and susceptibility of hydatidosis. Notably, the MvaIbc-SacIIab-Hin1Iab haplotype of MHC-DRB1 exon two was associated with resistance hydatidosis. In order to analyze the antibody and cytokine responses to hydatidosis in Kazakh sheep with hydatidosis resistance haplotype, eight Kazakh sheep with the haplotype of MvaIbc-SacIIab-Hin1Iab were chosen as the test group, and other eight, which were not associated with hydatidosis resistance or susceptibility, were taken as control. After experimentally infected with hydatid orally, the blood was collected on 0, 7, 14, 30, 45, 60, 75, 90, 105, and 120 days. Serum and mRNA level of the cytokines IL-2, IFN-γ, TNF-α, IL-4, and IL-10 were evaluated by ELISA and fluorescence quantitative real-time polymerase chain reaction, respectively. The total white blood cells and leukomonocytes were determined by automation cytoanalyze. The level of IgE, IgG, and IgM were evaluated by ELISA. The results showed that the total white blood cells and leukomonocytes in test group were significantly higher than in control on 7, 45, 90, and 105 days post-infection (p.i.). The serum level of IL-2 in test group was significantly higher than in control on 45 days p.i., while the difference of IL-2 mRNA expression between test and control group was not significant. The serum level of TNF-α in test group was significantly higher than in control at 90 and 105 days p.i., and the TNF-α mRNA in test group was also significantly higher than in control on 90 days p.i. The level of IgE, IgG, and IgM in test group was higher than in control, but none was significant. The results suggested that the test group, which was predominant of Th1, could induce the protective immunity, while the control, which was predominant of Th2, could induce the susceptibility to infection of hydatidosis.