Diltiazem HCl suppresses porcine reproductive and respiratory syndrome virus infection in susceptible cells and in swine.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a pathogen for swine, resulting in substantial economic losses to the swine industry. However, there has been little success in developing effective vaccines or drugs for PRRSV control. In the present study, we discovered that Diltiazem HCl, an inhibitor of L-type Ca2+ channel, effectively suppresses PRRSV replication in MARC-145, PK-15CD163 and PAM cells in dose-dependent manner. Furthermore, it demonstrates a broad-spectrum activity against both PRRSV-1 and PRRSV-2 strains. Additionally, we explored the underlying mechanisms and found that Diltiazem HCl -induced inhibition of PRRSV associated with regulation of calcium ion homeostasis in susceptible cells. Moreover, we evaluated the antiviral effects of Diltiazem HCl in PRRSV-challenged piglets, assessing rectal temperature, viremia, and gross and microscopic lung lesions. Our results indicate that Diltiazem HCl treatment alleviates PRRSV-induced rectal temperature spikes, pulmonary pathological changes, and serum viral load. In conclusion, our data suggest that Diltiazem HCl could serve as a novel therapeutic drug against PRRSV infection.

Effects of Concentrate Feeding Sequence on Growth Performance, Nutrient Digestibility, VFA Production, and Fecal Microbiota of Weaned Donkeys.

In this study, effects on the growth performance, nutrient digestibility, volatile fatty acids (VFA) production, and fecal microbiota of weaned donkeys were observed using different concentrate feeding sequences. Fifteen healthy 6-month-old weaned male donkeys with a body weight of 117.13 ± 10.60 kg were randomly divided into three treatment groups, including group C1 (roughage-then-concentrate), group C2 (concentrate-then-roughage), and group C3 (total mixed ration, TMR). The experiment lasted 35 d. We measured nutrient digestion by the acid-insoluble ash method and analyzed the fecal microbiota of the weaned donkeys by high-throughput sequencing of 16s rRNA genes in the V3-V4 region. The results show that group C3 obtained the best growth performance, and the digestibility of crude protein (CP) and crude extract (EE) was significantly higher than that of group C1 (p < 0.05). Acetic acid, isobutyric acid, valeric acid, isovaleric acid, and caproic acid were notably different among all groups (p < 0.05). In addition, we observed that Firmicutes and Bacteroidetes were dominant in the fecal microbes of each group, and Firmicutes was significantly higher in group C3 (p < 0.05). At the genus level, the different genera were Treponema, Rikenellaceae-RC9-gut-group, Unidentified-F082, and Bacteroidales-RF16-group (p < 0.05). The prediction of fecal microbiota function by PICRUSt indicated that different feeding sequences had minimal impact on the function of the fecal microbiota, particularly on the high-abundance pathway. In summary, the concentrate feeding sequence changed the composition of the fecal microbe of weaned donkeys.

Integrated analysis of transcriptome and proteome for exploring mechanism of promoting proliferation of equine satellite cells associated with leucine.

The proliferation and differentiation of skeletal muscle satellite cells (SCs) are necessary for the development of mature skeletal muscle. Leucine (Leu) is both an essential amino acid (EAA) and a branched-chain amino acid (BCAA), which has attracted worldwide attention due to its ability to repair and become new fibers. We separated the equine SCs into the control group (CON) and the Leu-supplemented group (LEU), which the cells were cultured in Leu-deprived and Leu-supplemented media respectively. We combined the transcriptome (RNA-Seq) and quantitative proteome (TMT) profiling analyses on proliferation of equine SCs associated with Leu. 1839 up-regulated and 631 down-regulated genes made up the 2470 differentially expressed genes (DEGs), and the 253 differentially abundant proteins (DEPs) included 118 up-regulated and 135 down-regulated proteins. 110 overlapping genes were verified based on the mRNA and protein translation relationship. Moreover, by comparing overlapped pathways through enrichment analysis, we found 13 genes not only appeared among 110 key DEGs/DEPs but also enriched in the KEGG overlapping signaling pathway, including CCL26, STAT2, PCK2, ASNS, GPT2, SHMT2, PHGDH, PGAM2, PSAT1, FTL, HMOX1, STEAP1 and STEAP2. To our knowledge, this is the first report in the world to systematically show how Leu regulated the growth of equine SCs. Leu deficiency inhibits the proliferation of equine SCs and development of fresh muscle fibers was proved in this paper. The main genes in charge of the Leu-induced proliferation of horse SCs have been found. These genes will make it easier to understand the mechanism at work and offer new information for enhancing the performance of sport horses and alleviating the equine muscle damage during exercise in the future.

Donkey Oil-Based Ketogenic Diet Prevents Tumor Progression by Regulating Intratumor Inflammation, Metastasis and Angiogenesis in CT26 Tumor-Bearing Mice.

Colon cancer is one of the typical malignant tumors, and its prevalence has increased yearly. The ketogenic diet (KD) is a low-carbohydrate and high-fat dietary regimen that inhibits tumor growth. Donkey oil (DO) is a product with a high nutrient content and a high bioavailability of unsaturated fatty acids. Current research investigated the impact of the DO-based KD (DOKD) on CT26 colon cancer in vivo. Our findings revealed that DOKD administration significantly lowered CT26+ tumor cell growth in mice, and the blood ß-hydroxybutyrate levels in the DOKD group was significantly higher than those in the natural diet group. Western blot results showed that DOKD significantly down-regulated Src, hypoxia inducible factor-1α (HIF-1α), extracellular signal-related kinases 1 and 2 (Erk1/2), snail, neural cadherin (N-cadherin), vimentin, matrix metallopeptidase 9 (MMP9), signal transducer and activator of transcription 3 (STAT3), and vascular endothelial growth factor A (VEGFA), and it significantly up-regulated the expressions of Sirt3, S100a9, interleukin (IL)-17, nuclear factor-kappaB (NF-κB) p65, Toll-like receptor 4 (TLR4), MyD88, and tumor necrosis factor-α. Meanwhile, in vitro validation results showed that LW6 (a HIF-1α inhibitor) significantly down-regulated the expressions of HIF-1α, N-cadherin, vimentin, MMP9, and VEGFA, which supported those of the in vivo findings. Furthermore, we found that DOKD inhibited CT26+ tumor cell growth by regulating inflammation, metastasis, and angiogenesis by activating the IL-17/TLR4/NF-κB p65 pathway and inhibiting the activation of the Src/HIF-1α/Erk1/2/Snail/N-cadherin/Vimentin/MMP9 and Erk1/2/HIF-1α/STAT3/VEGFA pathways. Our findings suggest that DOKD may suppress colon cancer progression and help prevent colon cancer cachexia.

Characteristics and epidemiological investigation of equid herpesvirus 8 in donkeys in Shandong, China.

Equid herpesvirus 8 (EHV-8), also known as asinine herpesvirus type 3 (AHV-3), can cause severe respiratory disease, abortion in mares, and neurological disorders. There is limited information on the prevalence of EHV-8 in donkeys in China. In this study, we investigated EHV-8 infection in donkeys using PCR, resulting in the identification of a field strain, termed EHV-8 SD2020113, which was isolated using RK-13 cells and characterized by high-throughput sequencing and transmission electron microscopy. Our data indicated that 38.7% (457/1180) of donkeys showed the presence of EHV-8 in blood samples. Analysis of the ORF70 gene showed the highest similarity (99.8-99.9% identity) to EHV-8 IR/2015/40 (MF431614.1) and SDLC66 (MW816102), and, in phylogenetic analysis, it clustered with EHV-8 SDLC66 from China. The findings of this study indicate that EHV-8 is likely to represent a threat to the donkey industry, and breeders and veterinarians who care for donkey farms should be aware of this.

Effect of Sperm Cryopreservation in Farm Animals Using Nanotechnology.

Sperm cryopreservation is one of the sublime biotechnologies for assisted reproduction. In recent decades, there has been an increasing trend in the use of preserved semen. Post-thaw semen quality and values vary among animals of the same species. Similarly, there are species-specific variations in sperm morphology, i.e., sperm head, kinetic properties, plasma membrane integrity, and freezability. Similarly, the viability of sperm varies in the female reproductive tract, i.e., from a few hours (in cattle) to several days (in chicken). Various steps of sperm cryopreservation, i.e., male health examination, semen collection, dilution, semen centrifugation, pre- and post-thaw semen quality evaluation, lack standardized methodology, that result in differences in opinions. Assisted reproductive technologies (ART), including sperm preservation, are not applied to the same extent in commercial poultry species as in mammalian species for management and economic reasons. Sperm preservation requires a reduction in physiological metabolism by extending the viable duration of the gametes. Physiologically and morphologically, spermatozoa are unique in structure and function to deliver paternal DNA and activate oocytes after fertilization. Variations in semen and sperm composition account for better handling of semen, which can aid in improved fertility. This review aims to provide an update on sperm cryopreservation in farm animals.

MRKNs: Gene, Functions, and Role in Disease and Infection.

The makorin RING finger protein (MKRN) gene family encodes proteins (makorins) with a characteristic array of zinc-finger motifs present in a wide array from invertebrates to vertebrates. MKRNs (MKRN1, MKRN2, MKRN3, MKRN4) as RING finger E3 ligases that mediate substrate degradation are related with conserved RING finger domains that control multiple cellular components via the ubiquitin-proteasome system (UPS), including p53, p21, FADD, PTEN, p65, Nptx1, GLK, and some viral or bacterial proteins. MKRNs also served as diverse roles in disease, like MKRN1 in transcription regulation, metabolic disorders, and tumors; MKRN2 in testis physiology, neurogenesis, apoptosis, and mutation of MKRN2 regulation signals transduction, inflammatory responses, melanoma, and neuroblastoma; MKRN3 in central precocious puberty (CPP) therapy; and MKRN4 firstly reported as a novel E3 ligase instead of a pseudogene to contribute to systemic lupus erythematosus (SLE). Here, we systematically review advances in the gene's expression, function, and role of MKRNs orthologs in disease and pathogens infection. Further, MKRNs can be considered targets for the host's innate intracellular antiviral defenses and disease therapy.

The Effect of Inhibin Immunization in Seminiferous Epithelium of Yangzhou Goose Ganders: A Histological Study.

The current study investigated the effect of inhibin immunization on germ cell numbers (spermatogonia, spermatocytes, round, and elongated spermatids), seminiferous tubules (ST) diameter, Johnsen's score, epithelial height (µm), luminal tubular diameter (µm), and number of ST per field (ST/field) of Yangzhou goose ganders. Histological evaluation showed apoptosis and regression of testes after inhibin (INH) immunization, with a concomitantly marked reduction in the round and elongated spermatids in the experiment (INH) group compared to the control group. The diameter of seminiferous tubules (ST) and epithelial height (EH) were positively correlated at 181, 200, and 227 days of age. In comparison, luminal tubular diameter (LD) was negatively correlated on day 227 to ST diameter and epithelial height. On day 227, many seminiferous tubules per field (ST/field) were negatively correlated to ST diameter, EH, and LD. INH immunization elevated ST diameter, EH, and LD, while Johnsen's score and number of ST/field had reciprocal expression. In conclusion, the concomitant effect of INH immunization and seasonality in breeding regressed germ cells and damaged spermatogenesis in seminiferous epithelium Yangzhou ganders.

Improving Gander Reproductive Efficacy in the Context of Globally Sustainable Goose Production.

The goose is a popular poultry species, and in the past two decades the goose industry has become highly profitable across the globe. Ganders low reproductive performance remains a barrier to achieving high fertility and hatchability in subsequent flocks. To address the global demand for cheaper animal protein, various methodologies for improving avian (re)production should be explored. A large amount of literature is available on reproduction traits and techniques for commercial chicken breeder flocks, while research on improved reproduction in ganders has been carried out to a lesser extent. The present review aims to provide a comprehensive literature overview focusing on recent advancements/techniques used in improving gander reproductive efficacy in the context of ensuring a globally sustainable goose industry.

Spermiogenesis, Stages of Seminiferous Epithelium and Variations in Seminiferous Tubules during Active States of Spermatogenesis in Yangzhou Goose Ganders.

The past three decades revolutionized the goose industry in the world. China holds the world's largest goose breeds stock by 95% of the global total. To optimize the goose industry and cope with ever increasing poultry meat and egg demands, there is a dire need to focus on reproduction, as most geese breeds exhibit poor reproductive performance. The present study was conducted with the aim to add a contribution in the goose industry and research by the histological visualizing step wise development of germ cells during spermatogenesis by microscopy and a histological technique. Yangzhou goose is a synthetic breed developed by using local goose germplasm resources of China. It is popular in the Chinese goose industry due to high productivity and adaptability. This research evaluated the steps of spermiogenesis and stages along with morphological changes in the seminiferous epithelium in Yangzhou goose ganders. For the assessment of various stages of the seminiferous epithelium cycle, testis sections were embedded in molten paraffin wax. The initial steps of spermiogenesis were depicted by changes in acrosomic granules, whereas further stages were identified by nuclear morphological changes. Ten steps of spermiogenesis and nine stages of seminiferous epithelium were identified. Four types of spermatogonia Ad, Ap1, Ap2 and B were recognized. The results depicted a clear variation in the diameter of seminiferous tubules (ST), epithelium height (EH), luminal tubular diameter (LD), number of seminiferous tubules per field and the Johnsen score. Microscopy indicated that the stages of seminiferous epithelium were similar to other birds and mammals and the ST diameter, EH, LD and Johnsen score are positively correlated while the number of seminiferous tubules per field is negatively correlated with the ST diameter, EH, LD and Johnsen score. Fertility in Yangzhou ganders can further be improved by visualizing the histological development of germs cells in testis tissues during spermatogenesis after onset of breeding season and maturity. Our results suggest that Yangzhou ganders reach complete sexual maturity at 227 days of age.

The role of active immunization against inhibin α-subunit on testicular development, testosterone concentration and relevant genes expressions in testis, hypothalamus and pituitary glands in Yangzhou goose ganders.

The present study was designed to investigate the potential role of immunization against inhibin on testicular development, plasma testosterone concentration and expression of relevant genes in hypothalamus, pituitary, Leydig and Sertoli cells in Yangzhou ganders. For this purpose, Yangzhou ganders, n = 30 were divided into groups A and B. Group B ganders were actively immunized against inhibin α-subunit, while group A ganders were immunized with bovine serum albumin (BSA), which served as control. Immunization against inhibin elevated testes weights. In addition, immunization against inhibin elevated GnRH, StAR, CYP11A1 and AMH mRNA transcription expressions as depicted by qRT-PCR. Furthermore, hypothalamic GnRH-I mRNA expressions were up regulated, while GnIH mRNA transcription expression showed reciprocal expression on day 227. LH-ß mRNA transcription expression remained unaffected. In conclusion, our findings suggest that active immunization against inhibin affect spermatogenesis and testicular development through regulations of hypothalamic, pituitary and testicular genes expressions.

Time course effect of lipopolysaccharide on Toll-like receptors expression and steroidogenesis in the Chinese goose ovary.

The ovary of Chinese goose is easily infected by microorganisms because of the mating behaviour in water, which causes decreased laying performance. This study investigated the time course effect of lipopolysaccharide (LPS) on the steroidogenesis and mRNA expression of Toll-like receptors (TLRs), a class of key pattern recognition receptor, in the breeding goose ovary. The laying geese were treated intravenously with LPS for 0, 6, 12, 24 and 36 h, and all birds were slaughtered approximately 8 h after oviposition. The expression levels of TLRs in the white and yellowish follicles, and granulosa and theca layers of hierarchical follicles were examined by real-time PCR. All 10 members of avian TLR family were differentially expressed among the different follicular tissues. Moreover, at 24 and 36 h after LPS treatment, the hierarchical follicle morphological structure was altered, but the expression levels of TLRs were still higher than the control. Furthermore, during LPS treatment period, the expression pattern of TLRs 2A and 4 genes was similar to that of TLR15 in the white follicles, TLRs 1B, 5 and 15 in the yellowish follicles, TLRs 7 and 15 in the granulosa layer, and TLRs 1A, 2B, 3, 7 and 15 in the theca layer, which had a negative correlation with the kinetics of plasma P4 and E2 concentrations. In conclusion, the mechanism by which pathogen infection inhibited goose follicular growth and further decreased egg production may involve a gradually enhanced inflammatory response and reduced endocrine function. This may be due to stimulated TLRs in the ovary.

Methods of synthesis of hydrogels A review.

Hydrogels are being investigated recently for the bioactive molecules (in particular pharmaceutical proteins) controlled release, such as matrices, and for the living cells encapsulation. Biodegradable nature of hydrogels has created much interest for drug delivery systems. The original three-dimensional structure disintegrates into nontoxic substances to ascertain an excellent biocompatibility of the gel. Chemical cross-linking is the highly resourceful method for the formation of hydrogels having an excellent mechanical strength. Cross-linkers used in hydrogel preparation should be extracted from the hydrogels before use due to their reported toxicity. Physically cross-linked methods for preparation of hydrogel are the alternate solution of cross-linker toxicity.

Effect of ethylene glycol dimethacrylate on swelling and on metformin hydrochloride release behavior of chemically crosslinked pH-sensitive acrylic acid-polyvinyl alcohol hydrogel.

BACKGROUND: The present work objective was to prepare and to observe the effect of ethylene glycol dimethacrylate on swelling and on drug release behavior of pH-sensitive acrylic acid-polyvinyl alcohol hydrogel. METHODS: In the present work, pH sensitive acrylic acid-polyvinyl alcohol hydrogels have been prepared by free radical polymerization technique in the presence of benzoyl peroxide as an initiator. Different crosslinker contents were used to observe its effect on swelling and on drug release. Dynamic and equilibrium swelling studies of prepared hydrogels were investigated in USP phosphate buffer solutions of pH 1.2, 5.5, 6.5 and 7.5 with constant ionic strengths. Hydrogels were evaluated for polymer volume fraction, solvent interaction parameter, molecular weight between crosslinks, number of links per polymer chain, diffusion coefficient, sol-gel fraction and porosity. To demonstrate the release pattern of the drug, zero-order, first-order, higuchi and korsmeyer-peppas models were applied. Quality and consistency of hydrogels was examined by FTIR and surface morphology of hydrogels was examined by SEM. RESULTS: Decrease in swelling and in drug release was seen by increasing content of ethylene glycol dimethacrylate. A remarkable high swelling was observed at high pH indicating the potential of this hydrogel for delivery of drugs to intestine. By increasing the concentration of ethylene glycol dimethacrylate, porosity decreased. Order of release was observed first order in all cases and the mechanism was non-fickian diffusion. FTIR confirmed the formation of network. SEM results showed the incorporation of drug. CONCLUSION: The prepared hydrogels can be suitably used for targeted drug delivery to the intestine.