Moniezia benedeni drives the SNAP-25 expression of the enteric nerves in sheep's small intestine.

BACKGROUND: The neuroimmune network plays a crucial role in regulating mucosal immune homeostasis within the digestive tract. Synaptosome-associated protein 25 (SNAP-25) is a presynaptic membrane-binding protein that activates ILC2s, initiating the host's anti-parasitic immune response. METHODS: To investigate the effect of Moniezia benedeni (M. benedeni) infection on the distribution of SNAP-25 in the sheep's small intestine, the recombinant plasmid pET-28a-SNAP-25 was constructed and expressed in BL21, yielding the recombinant protein. Then, the rabbit anti-sheep SNAP-25 polyclonal antibody was prepared and immunofluorescence staining was performed with it. The expression levels of SNAP-25 in the intestines of normal and M. benedeni-infected sheep were detected by ELISA. RESULTS: The results showed that the SNAP-25 recombinant protein was 29.3 KDa, the titer of the prepared immune serum reached 1:128,000. It was demonstrated that the rabbit anti-sheep SNAP-25 polyclonal antibody could bind to the natural protein of sheep SNAP-25 specifically. The expression levels of SNAP-25 in the sheep's small intestine revealed its primary presence in the muscular layer and lamina propria, particularly around nerve fibers surrounding the intestinal glands. Average expression levels in the duodenum, jejunum, and ileum were 130.32 pg/mg, 185.71 pg/mg, and 172.68 pg/mg, respectively. Under conditions of M. benedeni infection, the spatial distribution of SNAP-25-expressing nerve fibers remained consistent, but its expression level in each intestine segment was increased significantly (P < 0.05), up to 262.02 pg/mg, 276.84 pg/mg, and 326.65 pg/mg in the duodenum, jejunum, and ileum, and it was increased by 101.06%, 49.07%, and 89.16% respectively. CONCLUSIONS: These findings suggest that M. benedeni could induce the SNAP-25 expression levels in sheep's intestinal nerves significantly. The results lay a foundation for further exploration of the molecular mechanism by which the gastrointestinal nerve-mucosal immune network perceives parasites in sheep.

Moniezia benedeni infection enhances neuromedin U (NMU) expression in sheep (Ovis aries) small intestine.

BACKGROUND: Neuromedin U (NMU) plays an important role in activating the group 2 innate lymphoid cells (ILC2s) and initiating the host's anti-parasitic immune responses. It is aimed to explore the distribution characteristics of NMU in the sheep small intestine and the influence of Moniezia benedeni infection on them. In the present study, the pET-28a-NMU recombinant plasmids were constructed, and Escherichia coli. BL21 (DE3) were induced to express the recombinant protein. And then, the rabbit anti-sheep NMU polyclonal antibody was prepared and immunofluorescence staining was performed with it. The expression levels of NMU in the intestine of normal and Moniezia benedeni-infected sheep were detected by ELISA. RESULTS: The results showed that the molecular weight of the obtained NMU recombinant protein was consistent with the expected molecular (13 kDa) and it was expressed in the form of inclusion body. The titer and specificity of obtained rabbit anti-sheep NMU polyclonal antibody were good. The results of immunofluorescence analysis showed that the nerve fibers which specifically expressed NMU mainly extended from the ganglion in the submucosal to lamina propria (LP) in the sheep small intestine, and the expression level was relatively high; especially on the nerve fibers of LP around the intestinal glands. The expression levels were gradually increased from the duodenum to the ileum, and the levels in the jejunum and ileum were significantly higher than that in the duodenum (P < 0.05). In addition, scattered NMU positive cells were distributed in the epithelium of the jejunal crypts. Moniezia benedeni infection increased the expression of NMU in each intestinal segment, especially in the jejunum and ileum there were significant increase (P < 0.05). CONCLUSIONS: It was suggested that Moniezia benedeni infection could be detected by the high expression of NMU in sheep enteric nervous, and which laid the foundation for further studies on whether NMU exerts anti-parasitic immunity by activating ILC2s. In addition, NMU was expressed in some intestinal gland epitheliums, which also provided a basis for studying its roles in regulation of the immune homeostasis. The present study laid the foundation for further revealing the molecular mechanism of sheep's neural-immune interaction network perceiving the colacobiosis of parasites.

Profiling the DNA methylation patterns of imprinted genes in abnormal semen samples by next-generation bisulfite sequencing.

PURPOSE: Changes in DNA methylation modifications have been associated with male infertility. With the development of assisted reproductive technologies (ARTs), abnormal DNA methylation in sperm, especially in imprinted genes, may impact the health of offspring and requires an in-depth study. METHODS: In this study, we collected abnormal human semen samples, including asthenospermic, oligospermic, oligoasthenospermic and deformed sperm, and investigated the methylation of imprinted genes by reduced representation bisulfite sequencing (RRBS) and bisulfite amplicon sequencing on the Illumina platform. RESULTS: The differentially methylated regions (DMRs) of imprinted genes, including H19, GNAS, MEG8 and SNRPN, were different in the abnormal semen groups. MEG8 DMR methylation in the asthenospermic group was significantly increased. Furthermore, higher methylation levels of MEG8, GNAS and SNRPN DMR in the oligospermic and oligoasthenospermic groups and a decrease in the H19 DMR methylation level in the oligospermic group were observed. However, the methylation levels of these regions varied greatly among the different semen samples and among individual sperm within the same semen sample. The SNP rs2525883 genotype in the H19 DMR affected DNA methylation. Moreover, DNA methylation levels differed in the abnormal semen groups in the non-imprinted genomic regions, including repetitive sequence DNA transposons and long/short interspersed nuclear elements (LINEs and SINEs). CONCLUSION: Our study established that imprinted gene DMRs, such as H19, GNAS, SNRPN and MEG8, were differentially methylated in the abnormal semen groups with obvious inter- and intra-sample heterogeneities. These results suggest that special attention needs to be paid to possible epigenetic risks during reproduction.

Comparative transcriptomics and histopathological analysis of crucian carp infection by atypical Aeromonas salmonicida.

Aeromonas salmonicida is a ubiquitous fish pathogen known to cause furunculosis. With the emergence of new subtypes and the expansion of the host range, it has threatened the health of a variety of marine and freshwater fish, particularly the non-salmonids, manifesting differently from the classical furunculosis. Although there have been reports of infection by atypical strains on the crucian carp, the pathogenesis and tissue pathology remain unclear. In this study, transcriptomics and histopathology were used to analyze the immune response and lesions of crucian carp infected with A. salmonicida. Comparative analysis showed 6579 differentially expressed genes (DEGs) (3428 down-regulated and 3151 up-regulated) were identified on day 5 post-infection (5 dpi). Further annotation and analysis revealed that the DEGs were enriched in enzyme regulator activity, response to oxidative stress, iron ion homeostasis and other functions, and mitogen-activated protein kinase (MAPK), nuclear factor-κB (NF-κB), toll-like receptor (TLR), and nucleotide-binding oligomerization domain (NOD)-like receptor (NLR) etc., and immune-related signaling pathways. Meanwhile, the four C-type lysozyme genes found in all DEGs were significantly up-regulated after infection. In addition, there was severe bleeding on the body of the infected fish. Also, the intestine, liver, spleen, and kidney showed varying degrees of inflammatory damage, especially the goblet cell hyperplasia of intestinal mucosa epithelium and degeneration and necrosis of renal tubular epithelium cells. Additionally, with the increase in pathogen concentration, the cumulative mortality increased, the severity of lesions in the hindgut and head-kidney tissues increased. The relative expression levels of four immune-related genes (TNF-α, IL-1ß, IL-11, C-lysozyme) were also significantly upregulated, compared with the control (P < 0.05). In conclusion, this study provides a scientific basis for further study on the immune response, pathological diagnosis, and prevention of crucian carp infection caused by atypical A. salmonicida.

Heterologous overexpression of Vigna radiata epoxide hydrolase in Escherichia coli and its catalytic performance in enantioconvergent hydrolysis of p-nitrostyrene oxide into (R)-p-nitrophenyl glycol.

Two native epoxide hydrolases (EHs) were previously discovered from mung bean powder (Vigna radiata), both of which can catalyze the enantioconvergent hydrolysis of p-nitrostyrene oxide (pNSO). In this study, the encoding gene of VrEH1 was successfully cloned from the cDNA of V. radiata by RT-PCR and rapid amplification of cDNA ends (RACE) technologies. High homologies were found to two putative EHs originated from Glycine max (80%) and Medicago truncatula (79%). The vreh1 gene constructed in pET28a(+) vector was then heterologously overexpressed in Escherichia coli BL21(DE3), and the encoded protein was purified to homogeneity by nickel affinity chromatography. It was shown that VrEH1 has an optimum activity at 45 °C and is very thermostable with an inactivation energy of 468 kJ mol(-1). The enzyme has no apparent requirement of metal ions for activity, and its activity was strongly inhibited by 1 mM of Ni(2+), Cu(2+), Fe(2+), or Co(2+). By adding 0.1% Triton X-100, the enzyme activity could be significantly increased up to 340%. VrEH1 shows an unusual ability of enantioconvergent catalysis for the hydrolysis of racemic pNSO, affording (R)-p-nitrophenyl glycol (pNPG). It displays opposite regioselectivity toward (S)-pNSO (83% to Cα) in contrast to (R)-pNSO (87% to Cß). The K M and k cat of VrEH1 were determined to be 1.4 mM and 0.42 s(-1) for (R)-pNSO and 5.5 mM and 6.2 s(-1) for (S)-pNSO. This thermostable recombinant VrEH1 with enantioconvergency is considered to be a promising biocatalyst for the highly productive preparation of enantiopure vicinal diols and also a good model for understanding the mechanism of EH stereoselectivity.

Moniezia benedeni drives CD3+ T cells residence in the sheep intestinal mucosal effector sites.

Introduction: T cells are the core of the cellular immunity and play a key role in the regulation of intestinal immune homeostasis. In order to explore the impact Moniezia benedeni (M. benedeni) infection on distributions of CD3+ T cells in the small intestine of the sheep. Methods: In this study, sheep pET-28a-CD3 recombinant plasmid were constructed and expressed in BL21 receptor cells, then the rabbit anti-sheep CD3 polyclonal antibody was prepared through recombinant protein inducing. The M. benedeni-infected sheep (infection group, n = 6) and healthy sheep (control group, n = 6) were selected, and the distributions of CD3+ T cells in intestinal laminae propria (LP) and mucous epitheliums were observed and analyzed systematically. Results: The results showed that the rabbit anti-sheep CD3 polyclonal antibody had good potency and specificity. In the effector area of small intestine, a large number of CD3+ T cells were mainly diffusely distributed in the intestinal LP as well as in the mucous epitheliums, and the densities of intestinal LP from duodenum to jejunum to ileum were 6.01 cells/104 µm2, 7.01 cells/104 µm2 and 6.43 cells/104 µm2, respectively. Their distribution densities in mucous epitheliums were 6.71 cells/104 µm2, 7.93 cells/104 µm2 and 7.21 cells/104 µm2, respectively; in the infected group, the distributions of CD3+ T cells were similar to that of the control group, and the densities in each intestinal segment were all significantly increased (p < 0.05), meanwhile, the total densities of CD3+ T cells in duodenum, jejunum and ileum were increased by 33.43%, 14.50%, and 34.19%. In LP and mucous epitheliums, it was increased by 33.57% and 27.92% in duodenum; by 25.82% and 7.07% in jejunum, and by 27.07% and 19.23% in ileum, respectively. Discussion: It was suggested that M. benedeni infection did not change the spatial distributions of CD3+ T cells in the small intestine of sheep, but significantly increased their densities, which lays a foundation for further research on the regulatory mechanism of sheep intestinal mucosal immune system against M. benedeni infection.

Epigenetic editing of BRCA1 promoter increases cisplatin and olaparib sensitivity of ovarian cancer cells.

Drug resistance is the primary contributor to the high mortality rate of ovarian cancer (OC). The loss of BRCA1/2 function is linked to drug sensitivity in OC cells. The aim of this study is to enhance the drug sensitivity of OC cells by inducing BRCA1 dysfunction through promoter epigenetic editing. Epigenetic regulatory regions within the BRCA1 promoter, affecting gene expression, were initially discerned through analysis of clinical samples. Subsequently, we designed and rigorously validated epigenetic editing tools. Ultimately, we evaluated the cisplatin and olaparib sensitivity of the OC cells after editing. The BRCA1 promoter contains two CpG-rich regions, with methylation of the region covering the transcription start site (TSS) strongly correlating with transcription and influencing OC development, prognosis, and homologous recombination (HR) defects. Targeting this region in OC cells using our designed epigenetic editing tools led to substantial and persistent DNA methylation changes, accompanied by significant reductions in H3K27ac histone modifications. This resulted in a notable suppression of BRCA1 expression and a decrease in HR repair capacity. Consequently, edited OC cells exhibited heightened sensitivity to cisplatin and olaparib, leading to increased apoptosis rates. Epigenetic inactivation of the BRCA1 promoter can enhance cisplatin and olaparib sensitivity of OC cells through a reduction in HR repair capacity, indicating the potential utility of epigenetic editing technology in sensitization therapy for OC.

Prenatal exposure to per- and polyfluoroalkyl substances and DNA methylation in the placenta: A prospective cohort study.

Epidemiological studies regarding the relationship between per- and polyfluoroalkyl substances (PFAS) and DNA methylation were limited. We investigated the associations of maternal PFAS concentrations with placental DNA methylation and examined the mediating role of methylation changes between PFAS and infant development. We measured the concentrations of 11 PFAS in maternal plasma during early pregnancy and infant development at six months of age. We analyzed genome-wide DNA methylation in 16 placental samples using reduced representation bisulfite sequencing. Additionally, we measured DNA methylation levels using bisulfite amplicon sequencing in 345 mother-infant pairs for five candidate genes, including carbohydrate sulfotransferase 7 (CHST7), fibroblast growth factor 13 (FGF13), insulin receptor substrate 4 (IRS4), paired like homeobox 2Ap (PHOX2A), and plexin domain containing 1 (PLXDC1). We found that placental DNA methylation profiles related to PFOA mainly enriched in angiogenesis and neuronal signaling pathways. PFOA was associated with hypomethylation of IRS4 and PLXDC1, and PFNA was associated with PLXDC1 hypomethylation. There were positive associations of CHST7 methylation with PFTrDA and IRS4 methylation with PFDoA and PFTrDA. PLXDC1 hypomethylation mediated the association between PFOA and suspected developmental delay in infants. Future studies with larger sample sizes are warranted to confirm these findings.

Novel histone post-translational modifications in Alzheimer's disease: current advances and implications.

Alzheimer's disease (AD) has a complex pathogenesis, and multiple studies have indicated that histone post-translational modifications, especially acetylation, play a significant role in it. With the development of mass spectrometry and proteomics, an increasing number of novel HPTMs, including lactoylation, crotonylation, ß-hydroxybutyrylation, 2-hydroxyisobutyrylation, succinylation, and malonylation, have been identified. These novel HPTMs closely link substance metabolism to gene regulation, and an increasing number of relevant studies on the relationship between novel HPTMs and AD have become available. This review summarizes the current advances and implications of novel HPTMs in AD, providing insight into the deeper pathogenesis of AD and the development of novel drugs.

Moniezia benedeni infection increases IgE+ cells in sheep (Ovis aries) small intestine.

The concentration of immunoglobulin (Ig) E is the lowest among serum Igs, but it can induces type I hypersensitivity and plays an important role in anti-parasitic infection. The present study aimed to explore the residence characteristics of IgE+ cells in the sheep small intestine and the impact of Moniezia benedeni infection on them. The recombinant plasmids pET-28a-IgE were constructed and induced and expressed in Escherichia coli. BL21 (DE3). The rabbit anti-sheep IgE polyclonal antibody was prepared using the obtained recombinant protein as antigen. Finally, the levels of IgE+ cells in the small intestine of healthy (Control group) and naturally M. benedeni-infected (Infected group) sheep were detected analyzed. The results showed that the rabbit anti-sheep IgE polyclonal antibody with good immunogenicity (titer = 1: 128000) could specifically bind to the heavy chain of natural sheep IgE. In the Control group, the IgE+ cells were mainly distributed in lamina propria of the small intestine, and the densities were significantly decreased from duodenum to ileum (P<0.05), with respective values of (4.28 cells / 104 µm2, 1.80 cells / 104 µm2, and 1.44 cells / 104 µm2 in duodenum, jejunum, and ileum. In the Infected group, IgE+ cells density were 6.26 cells / 104 µm2, 3.01 cells / 104 µm2, and 2.09 cells / 104 µm2 in duodenum, jejunum and ileum respectively, which were significantly higher in all segments compared to the Control group (P<0.05), increasing by 46.26%, 67.22% and 45.14%, respectively. In addition, compared with the Control group, the IgE protein levels were significantly increased in all intestinal segments of the Infected group (P<0.01), however, there was no significant differences among the different intestinal segments within the same group (P>0.05). The results demonstrated that M. benedeni infection could significantly increase the content of IgE and the distribution density of its secreting cells in sheep small intestine. The intestinal mucosal immune system of sheep presented obvious specificity against M. benedeni infection. This lays a good foundation for further exploring molecular mechanisms of the intestinal mucosal immune system monitoring and responding to M. benedeni infection.

The Degradation of Intramuscular Connective Tissue In Vitro with Purified Cathepsin L from Bovine Pancreas.

To investigate the possible degradation of the intramuscular connective tissue (IMCT) with cathepsin L, isolated IMCTs were incubated with purified cathepsin L in vitro. Here, we prepared purified cathepsin L from bovine pancreas by using DEAE Sephacel, Sephacryl S-100 HR, SP Sepharose FF, and con A-Sepharose affinity chromatography in sequence. An SDS-PAGE analysis of CNBr-digested peptides showed that the degradation of collagen in IMCT could take place on terminal non-helical peptides rather than the triple helix region. Decorin (DCN) was clearly degraded at a pH of 5.0. The TP and TO of intramuscular connective tissue decreased to 41.41 °C and 43.79 °C, respectively. In the cathepsin L treatment of pH 5.0, the decreases in the TP and TO of IMCT were more sensitive than they were at pH 5.5~6.5.

Spirituality Concern Toward Patients in Hospice Care during COVID-19 Pandemic in China.

Tension, anxiety, or fear spread all over the world during COVID-19 pandemic. How health professionals provide terminally ill patients with humanistic care to deal with fear is worth consideration. A more abundant spirituality concern in the body-mind-spirit care mode on patients in hospice care is appied in our practice.

Moniezia benedeni Infection Restrain IgA+, IgG+, and IgM+ Cells Residence in Sheep (Ovis aries) Small Intestine.

Secreted immunoglobulin A (SIgA), IgG, and IgM play a crucial role in forming the intestinal mucosal immune barrier, and parasites could disturb the host's immune response by releasing various immunomodulatory molecules. Moniezia benedeni is an important pathogen parasitizing in the sheep small intestine. It is aimed to explore the residence characteristics of IgA+, IgG+, and IgM+ cells in the sheep small intestine, and the influence of Moniezia benedeni infection on them. Control group (n = 6) and infected group (n = 6) were selected, respectively, and the three subtype cells residing in the small intestine were systematically observed and analyzed. The results showed that in the Control group, the three types of positive cells were all distributed diffusely, and the total densities in jejunum, duodenum and ileum was gradually declined in turn. Notably, the change trend of IgA+ and IgG+ cells densities were both congruent with the total densities, and the differences among them were significant, respectively (P < 0.05); the IgM+ cells density was the highest in duodenum, followed by jejunum and ileum, there was no significant difference between duodenum and jejunum (P > 0.05), but both significantly higher than in ileum (P < 0.05). In the Infected group, their total densities in duodenum, jejunum and ileum were gradually declined in turn. Notably, the IgA+ and IgM+ cells densities change trend was the same as the total densities, and the differences among them were significant, respectively (P < 0.05). The IgG+ cells density in duodenum was the highest, followed by ileum and jejunum and there was significantly difference among them (P < 0.05). The comparison results between Control and Infected groups showed that from the duodenum, jejunum to ileum, IgA+, IgG+, and IgM+ cells were all reduced significantly, respectively. The results suggest that the three types of positive cells were resided heterogeneously in the small intestinal mucosa, that is, significant region-specificity; Moniezia benedeni infection could not change their diffuse distribution characteristics, but strikingly, reduce their resident densities, and the forming mucosal immune barrier were significantly inhibited. It provided powerful evidence for studying on the molecular mechanism of Moniezia benedeni evasion from immune surveillance by strongly inhibiting the host's mucosal immune barrier.

Expression characteristics of polymeric immunoglobulin receptor in Bactrian camel (Camelus bactrianus) lungs.

Polymeric immunoglobulin receptor (pIgR), the transmembrane transporter of polymeric immunoglobulin A and M, has multiple immune functions. To explore the characteristics of pIgR expression in Bactrian camel lungs, twelve healthy adult (2-7 years old) Bactrian camels were systematically studied. The results showed that pIgR was mainly expressed in the cytoplasm and membrane of ciliated cells, as well as in the cytoplasm and membrane of basal cells, serous cells of bronchial glands, club cells and alveolar type 2 cells in Bactrian camel lungs. Specially, as the bronchial branches extended, the pIgR expression level in ciliated cells significantly declined (p<0.05), and the corresponding bronchial luminal areas obviously decreased (p<0.05). However, pIgR was not expressed in goblet cells, endocrine cells, alveolar type 1 cells and mucous cells of bronchial glands. The results demonstrated that ciliated cells continuously distributed throughout the whole bronchial tree mucosa were the major expression sites of pIgR, and pIgR was also expressed in basal cells, serous cells of bronchial glands, club cells and alveolar type 2 cells, which would facilitate secretory immunoglobulin A (SIgA) transmembrane transport by pIgR and form an intact protective barrier. Moreover, the pIgR expression level in ciliated cells was positively correlated with the bronchial luminal areas; but negatively correlated with the cleanliness of airflow through the bronchial cross-sections, showing that the pIgR expression level in the bronchial epithelium was inhomogeneous. Our study provided a foundation for further exploring the regulatory functions of immunoglobulins (i.e., SIgA) after transport across the membrane by pIgR in Bactrian camel lungs.

Hysterectomy by Transvaginal Natural Orifice Transluminal Endoscopic Surgery versus Transumbilical Laparoscopic Single-Site Surgery: A Single-Center Experience from East China.

Objective: To compare hysterectomy by transvaginal natural orifice transluminal endoscopic surgery (VNOTES) versus transumbilical laparoscopic single-site surgery (LESS) as a minimal invasive technique. Materials and Method. The women undergoing hysterectomy for benign diseases by VNOTES and LESS from January 2020 to June 2021 in a tertiary hospital in Shanghai were retrospectively analyzed. Results: 361 women were included in our study, with 228 in the VNOTES groups, 129 in the LESS groups, and 4 conversions from VNOTES to LESS technique. The length of a VNOTES hysterectomy was shorter than that of LESS (80.76 min versus 112.09 min; MD -31.34 min; 95% CI -40.24 to -22.43 min; P < 0.001). VNOTES hysterectomy has a quicker gas passage by the anus (18.80 versus 36.49 hours, MD -17.68 hours, 95% CI -20.23 to -15.14 hours, P < 0.001) and associated with a shorter length of hospital stay (2.31 versus 3.77 days, MD -1.46 days, 95% CI -1.75 to -1.17 days, P < 0.001), while with no increase in blood loss during the operation (median 50 versus 50 ml, P = 0.25). Besides, the VAS pain score in the 24th hour after the operation was lower (median 0 versus 0.5, P < 0.001) in the VNOTES group. Four unique phases of the learning curve were identified using cumulative analysis: the mean operation time of phase I was 82.81 ± 31.45 min (the initial learning curve of 43 cases), phase II was 72.48 ± 23.66 min (the acquisition of command of 91 cases), phase III was 103.77 ± 45.69 min (the further learning of 26 cases), and phase IV was 73.18 ± 26.89 min (postlearning in 68 cases). Conclusions: VNOTES is noninferior to LESS as a new minimal invasive procedure for hysterectomy, which also allows patients a faster recovery from surgery and to suffer less pain, and its efficiency and feasibility in large uterine need further exploring.

Age-related changes in the anatomical characteristics of Peyer's patches in small intestine of Bactrian camels (Camelus bactrianus).

The distribution, size, and appearance of Peyer's patches vary according to species. In order to determine the anatomical characteristics of Peyer's patches in small intestine of Bactrian camel, and age-related changes in the number of Peyer's patches, 40 Bactrian camels of the following four age groups were studied: young (0.5-2 years), pubertal (3-5 years), middle-aged (6-16 years), and old (17-20 years). The exact number of Peyer's patches was recorded, and the appearance of Peyer's patches was described in detail. The results indicated that Peyer's patches of Bactrian camels not only have a particular anatomical location and distinct appearance but also change with age. They were distributed in the whole small intestine and there were four distinct types of Peyer's patches: nodular, faviform, cup-shaped, and cystic form Peyer's patches. However, the nodular and cystic form Peyer's patches are specific to Bactrian camel, which have not been found in other animals including Dromedary camel. In addition, the distribution density of Peyer's patches in ileum was the maximum, then was jejunum and duodenum. Further statistical analysis showed that the number of Peyer's patches was altered with age. The number peaked in 5-year-old camels and declined subsequently with age. However, there was little change in the size of Peyer's patches in different age groups; no age-related macroscopic variations in the shape or size of the Peyer's patches were found. Results obtained from this study provide the basic information to further study on the gastrointestinal mucosal immunity of Bactrian camel.

Bacterial community analysis on the different mucosal immune inductive sites of gastrointestinal tract in Bactrian camels.

The microbial communities colonize the mucosal immune inductive sites could be captured by hosts, which could initiate the mucosal immune responses. The aggregated lymphoid nodule area (ALNA) and the ileal Payer's patches (PPs) in Bactrian camels are both the mucosal immune inductive sites of the gastrointestinal tract. Here, the bacteria community associated with the ALNA and ileal PPs were analyzed using of 16S rDNA-Illumina Miseq sequencing. The mutual dominant bacterial phyla at the two sites were the Bacteroidetes, Firmicutes, Verrucomicrobia and Proteobacteria, and the mutual dominant genus in both sits was Prevotella. The abundances of the Fibrobacter, Campylobacter and RFP12 were all higher in ALNA than in ileal PPs. While, the abundances of the 5-7N15, Clostridium, and Escherichia were all higher in ileal PPs than in ALNA. The results suggested that the host's intestinal microenvironment is selective for the symbiotic bacteria colonizing the corresponding sites, on the contrary, the symbiotic bacteria could impact on the physiological functions of this local site. In ALNA and ileal PPs of Bactrian camel, the bacteria which colonized different immune inductive sites have the potential to stimulate different immune responses, which is the result of the mutual selection and adaptation between microbial communities and their host.

siRNA-based breast cancer therapy by suppressing 17ß-hydroxysteroid dehydrogenase type 1 in an optimized xenograft cell and molecular biology model in vivo.

PURPOSE: Hormone-dependent breast cancer is the most common form of breast cancer, and inhibiting 17ß-HSD1 can play an attractive role in decreasing estrogen and cancer cell proliferation. However, the majority of existing inhibitors have been developed from estrogens and inevitably possess residual estrogenicity. siRNA knockdown provides a highly specific way to block a targeted enzyme, being especially useful to avoid estrogenicity. Application of 17ß-HSD1-siRNA in vivo is limited by the establishment of an animal model, as well as the potential nuclease activity in vivo. We tried to reveal the in vivo potential of 17ß-HSD1-siRNA-based breast cancer therapy. MATERIALS AND METHODS: To establish a competent animal model, daily subcutaneous injection of an estrone micellar aqueous solution was adopted to provide the substrate for estradiol biosynthesis. The effects of three different doses of estrone (0.1, 0.5, and 2.5 µg/kg/day) on tumor growth in T47D-17ß-HSD1-inoculated group were investigated and compared with the animals inoculated with wild type T47D cells. To solve in vivo delivery problem of siRNA, "17ß-HSD1-siRNA/LPD", a PEGylated and modified liposome-polycation-DNA nanoparticle containing 17ß-HSD1-siRNA was prepared by the thin film hydration method and postinsertion technology. Finally, "17ß-HSD1-siRNA/LPD" was tested in the optimized model. Tumor growth and 17ß-HSD1 expression were assessed. RESULTS: Comparison with the untreated group revealed significant suppression of tumor growth in "17ß-HSD1-siRNA/LPD"-treated group when HSD17B1 gene expression was knocked down. CONCLUSION: These findings showed promising in vivo assessments of 17ß-HSD1-siRNA candidates. This is the first report of an in vivo application of siRNA for steroid-converting enzymes in a nude mouse model.

The distributive and structural characteristics of bronchus-associated lymphoid tissue (BALT) in Bactrian camels (Camelus bactrianus).

BACKGROUND: Bronchus-associated lymphoid tissue (BALT), distributed in the bronchial mucosa, plays a critical role in maintaining the mucosal immune homeostasis of the lower respiratory tract. The bronchial tree is a functional structure for gas exchange with the outside environment and maintains basic lung morphology. METHODS: To explore the structural and distributive characteristics of BALT in Bactrian camels, twelve healthy adult Bactrian camels were divided into two groups (six in each group). The lungs, bronchial tree and BALT were observed and analysed systematically through anatomical and histological methods. RESULTS: The results showed that Bactrian camel lungs were constituted by the left cranial lobe, left caudal lobe, right cranial lobe, right caudal lobe and accessory lobe, but lacked the middle lobe. The cranial lobe was narrow and small, the caudal lobe was extremely developed (almost four times the cranial lobe in size), and the accessory lobe was smaller than the cranial lobe; the bronchial tree, an unequal dichotomy with a tracheobronchial branch, was composed of dorsal, ventral, lateral and medial bronchiole systems. Isolated lymphoid follicles (the chief type) and aggregates of lymphoid follicles revealed two types of BALT, and germinal centres, follicle-associated epithelium and high endothelial venules could be observed in some well-developed BALT. Additionally, BALT was scattered along the bronchial tree in the entire lung, and the density increased from the trachea to the lower graded branches (densest in the bronchioles) and then decreased, with the occasional location around respiratory bronchioles or among the pulmonary mesenchyme. In the conducting portion, BALT was primarily located in the mucosa lamina propria but was also found in the submucosa, under the muscular layer, and around the submucosal glands and cartilage. CONCLUSION: The results demonstrated that the lung morphology of Bactrian camels was similar to that of horses, but the bronchial branches were more closely related to those of ruminants. These characteristics were in accordance with the morphological and structural variation regularity of lungs with species evolution. BALT was mainly scattered in the conducting portion, and bronchioles, as the final "checkpoint" in the surveillance, capture and recognition of antigens before pulmonary exchange, were the pivotal locational position of BALT. However, BALT at different depths of the bronchial wall of the conducting portion might be at different developmental stages. Our study provided evidence for further insight into the mucosal immunomodulatory mechanism of BALT in the respiratory system of Bactrian camels.

Hypermethylation of the PTTG1IP promoter leads to low expression in early-stage non-small cell lung cancer.

Despite the clinical requirement for early diagnosis, the early events in lung cancer and their mechanisms are not fully understood. Pituitary tumor transforming gene 1 binding factor (PTTG1IP) is a tumor-associated gene; however, to the best of our knowledge, its association with lung cancer has not been reported. The present study analyzed PTTG1IP expression in early-stage non-small cell lung cancer (NSCLC) samples and investigated its epigenetic regulatory mechanisms. The results revealed that the mRNA level of PTTG1IP in NSCLC tissues was significantly downregulated by 43% compared with that in adjacent tissues. In addition, overexpression of this gene significantly inhibited cell proliferation. According to data from The Cancer Genome Atlas, a significant negative correlation was identified between the PTTG1IP gene methylation level and expression level in lung adenocarcinoma and lung squamous cell carcinoma cases. Reduced representation bisulfite sequencing (RRBS) analysis of six paired early-stage NSCLC tissue samples indicated that the CpG island shore of the PTTG1IP promoter is hypermethylated in lung cancer tissues, which was further validated in 12 paired early-stage NSCLC samples via bisulfite amplicon sequencing. Following treatment with 5-aza-2'-deoxycytidine to reduce DNA methylation in the promoter region, the PTTG1IP mRNA level increased, indicating that the PTTG1IP promoter DNA methylation level negatively regulates PTTG1IP transcription. In conclusion, in early-stage NSCLC, the PTTG1IP gene is regulated by DNA methylation in its promoter region, which may participate in the development and progression of lung cancer.