Hydroxytyrosol mitigates Mycoplasma gallisepticum-induced pulmonary injury through downregulation of the NF-κB/NLRP3/IL-1ß signaling pathway in chicken.

In this study, the anti-inflammatory and antiapoptotic effects of hydroxytyrosol (HT) in Mycoplasma gallisepticum (MG)-infected chicken were investigated, and the underlying molecular mechanisms were explored. The results revealed severe ultrastructural pathological changes after MG infection in the lung tissue of chicken, including inflammatory cell infiltration, thickening of the lung chamber wall, visible cell swelling, mitochondrial cristae rupture, and ribosome shedding. MG possibly activated the nuclear factor κB (NF-κB)/nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3)/interleukin (IL)-1ß signaling pathway in the lung. However, HT treatment significantly ameliorated MG-induced pathological damage of the lung. HT reduced the magnitude of pulmonary injury after MG infection by reducing apoptosis and releasing the proinflammatory factors. Compared with the MG-infected group, the HT-treated group exhibited significant inhibition of the expression of NF-κB/NLRP3/IL-1ß signaling-pathway-related genes; for example, the expressions of NF-κB, NLRP3, caspase-1, IL-1ß, IL-2, IL-6, IL-18, and TNF-α significantly decreased (P < 0.01 or <0.05). In conclusion, HT effectively inhibited MG-induced inflammatory response and apoptosis and protected the lung by blocking the activation of NF-κB/NLRP3/IL-1ß signaling pathway and reducing the damage caused by MG infection in chicken. This study revealed that HT may be a suitable and effective anti-inflammatory drug against MG infection in chicken.

Kisspeptin-10 in cryodiluent improves the post-thaw quality of Nili-Ravi buffalo (Bubalus bubalis) bull spermatozoa.

Effects of kisspeptin-10 as antioxidant in cryodiluent were evaluated on post-thaw quality of buffalo spermatozoa. Qualified semen samples from five bulls were pooled, divided into five aliquots and extended in Tris-citric acid cryodiluent containing differential doses of kisspeptin-10 (5, 10, 15, and 20 µmol L-1 and negative control. Extended sperm suspension was cooled to 4°C, packaged in 0.54 ml straws and cryopreserved. At post-thawing, catalase (unit mg-1 ), peroxidase (unit mg-1 ) and reduced glutathione (µmol L-1 ) levels were highest (p < 0.05) with 20 µmol L-1 of kisspeptin-10 as compared to negative control. Moreover, lipid peroxidation (nmol L-1  min-1  mg protein-1 ) level was lowest (p < 0.05) with 20 µmol L-1 of kisspeptin-10. Sperm progressive motility (%), rapid velocity (%) and kinematics were higher (p < 0.05) with 15 and 20 µmol L-1 of kisspeptin-10 as compared to negative control. Supra-vital plasma membrane integrity (%), viable sperm with intact acrosome (%) and DNA integrity (%) were improved (p < 0.05) with all doses of kisspeptin-10 as compared to negative control. It was concluded that the addition of 15 and 20 µmol L-1 kisspeptin-10 in cryodiluent ameliorated the overall frozen-thawed quality parameters of Nili-Ravi buffalo spermatozoa.

Validation of double freezing protocol for Beetal buck (Capra hircus) spermatozoa.

The study aimed to validate the double versus single freezing protocol for Beetal buck (Capra hircus) spermatozoa in tris-citric acid (TCA) based extender both in terms of quality and fertilization potential. Computer-assisted sperm motion and kinematic (CASA) variables, i.e. total (%), and progressive motilities (TM and PM, %) and rapid velocity (RV, %), average path (VAP, µm/s), straight line (VSL, µm/s) and curved line velocities (VCL, µm/s), straightness, (VSL/VAP, %) and linearity, (VSL/VCL, %) as well as supra-vital plasma membrane integrity (SV-PMI, %), mitochondrial membrane potential (MMP, %), viable/intact acrosome (V-IACR, %) and DNA integrity (DNA-I, %) had significantly greater values (p < .05) during single freeze-thawing as compared with the double freeze-thawing at 0, 30, 90, 150 and 210 days, respectively. All CASA and other assays alone did not show significant differences (p > .05) between both freeze-thaw cycles at all treatment durations, respectively. No statistical significance (p > .05) was observed for the in vivo fertility between single (n = 84/141 = 59.72%) and double freeze-thawing (n = 72/136 = 52.9%) cycles, respectively. In conclusion, sperm motion, kinematics, plasma membrane, acrosome, mitochondria and DNA integrities and in vivo fertility are acceptable after the double freezing protocol despite being lower than after one freeze cycle in Beetal buck.

Ginsenoside Rb1 prevents deoxynivalenol-induced immune injury via alleviating oxidative stress and apoptosis in mice.

Deoxynivalenol (DON) is considered to be a grave threat to humans and animals. Ginsenoside Rb1 (Rb1) has been reported for its antioxidant potential and medicinal properties. However, the shielding effects of Rb1 and the precise molecular mechanisms against DON-induced immunotoxicity in mice have not been reported yet. In the present research, 4-weeks old healthy C57BL/6 mice were randomly assigned into four experimental groups (n = 12), viz., CON, DON 3 mg/kg BW, Rb1 50 mg/kg BW and DON 3 mg/kg + Rb1 50 mg/kg BW (DON + Rb1). Feed intake and body weight gain were monitored during the entire experiment (15 d). Our results demonstrated that Rb1 markedly increased the ADG (30%) and ADFI (25.10%) of mice compared with DON group. Furthermore, Rb1 alleviated the DON-induced immune injury by relieving the splenic histopathological alteration, enhancing the T-lymphocytes subsets (CD4+, CD8+), the levels of cytokines (IL-2, IL-6, IFN-γ, and TNF-α), as well as production of immunoglobulins (IgA, IgM, and IgG). Moreover, Rb1 ameliorated DON-inflicted oxidative stress by reducing the ROS, MDA and H2O2 contents and boosting the antioxidant defense system (T-AOC, T-SOD, CAT, and GSH-Px). Additionally, Rb1 significantly reversed the DON-induced excessive splenic apoptosis via modulating the mitochondria-mediated apoptosis pathway in mice, depicting the decreased percentage of splenocyte apoptotic cells by 26.65%, down-regulated the mRNA abundance of Bax, caspase-3, caspase-9, and protein expression of Bax, cleaved caspase-3, and Cyt-c. Simultaneously, Rb1 markedly rescued both Bcl-2 mRNA and protein expression levels. Taken together, Rb1 mitigates DON-induced immune injury by suppressing the oxidative damage and regulating the mitochondria-mediated apoptosis pathway in mice. Conclusively, our current research provides an insight into the preventive mechanism of Rb1 against DON-induced immune injury in mice and thus, presents a scientific baseline for the therapeutic application of Rb1.

A systematic review and meta-analysis of spermatozoa cryopreservation, in vitro and in vivo fertility practices in water buffalo.

We explored different aspects of buffalo spermatozoa during cryopreservation. The meta-data comprised of 285 studies, published from January 2008 to March 2020. A free web tool CADIMA as well as PRISMA 2009 Flow Diagram were used for carrying out this study. The inter-reviewer agreement among studies allocated was satisfactory for criteria A (selection bias), B (performance bias), C (detection bias) and D (attrition bias), respectively. India led the percent (%) research ladder with 34.4, followed by Pakistan (29.5), Egypt (12.3), Iran (7.7), Italy (5.6), Indonesia (3.2), China (2.1), Brazil (1.4), Thailand (1.1), Philippines and Bulgaria (0.7 each), Saudi Arabia, Turkey, Vietnam, and USA (0.4 each). Among four categories of studies, Group-1 evaluated only supplements/additives/media in the freezing semen extender (n = 191/285; 67.02%); Group-2 conducted in vivo fertilization (n = 62/285; 21.75%) and Group-3 conducted in vitro fertilization/ cleavage rate/penetration rate/ blastocyst yields (n = 28/285; 9.82%) with their specific cryodiluents/media, respectively. Group-4 conducted different experimental supplements/additives/media and carried out both in vitro and in vivo fertilization simultaneously (n = 4/285; 1.40%). Conventional spermatozoa cryopreservation was reported by 51.9% studies followed by programmable fast freezing by 20.7% studies. A few leading extender types included BioXcell (3.9%); Soyamilk-skim (3.5%); and Andromed (2.1%). The study also describes French straws for semen filling, cooling temperatures, extension time, equilibration time, cryopreservation stages, thawing temperatures, seasons, thawing time, and stains used during semen evaluation assays. The study concludes that the research on spermatozoa cryopreservation of buffalo is largely conducted at quality level and a need of applying these findings for evaluation of fertility potential (in vivo and in vitro) is indispensable for effective genetic improvement.

The protective effect of walnut oil on lipopolysaccharide-induced acute intestinal injury in mice.

Walnut oil (WO) is widely used in traditional medicine, and it has become a dietary supplement in many countries. We isolated walnut oil from Juglans sigillata and evaluated its protective effects on acute intestinal injury, and Toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling pathway in lipopolysaccharide (LPS)-induced mice was studied. The results showed that the LPS + WO group significantly decreased serum tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IL-1ß levels and increased the jejunum superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels compared with the LPS group. Walnut oil ameliorated the pathological morphology of the LPS-induced acute jejunum injury and decreased jejunum cells apoptosis rate and TLR4/NF-κB protein expression. Furthermore, the expression of the TLR4/NF-κB pathway key gene mRNA significantly reduced after treatment with walnut oil. This study concludes that walnut oil can exert the protective effect on LPS-induced acute intestinal injury in mice by inhibiting the TLR4/NF-κB signaling pathway.

Effect of walnut (Juglans sigillata) oil on intestinal antioxidant, anti-inflammatory, immunity, and gut microbiota modulation in mice.

The study investigated the anti-oxidant, anti-inflammatory, immunity, and gut microbiota modulation in mice (n = 60; 15 mice/group) after intragastric administration of walnut oil (WO; three groups (low (LD), medium (MD), and high doses (HD): 2.5, 5, and 10 ml/kg, respectively) and normal control (NC, saline). WO significantly increased the median villous height/crypt depth (VH/CD) ratio, the activities of superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) in intestinal mucosa. WO exerted the anti-inflammatory effects by decreasing the expression of tumor necrosis factor-α (TNF-α) in the duodenal mucosa. All groups shared 157 operational taxonomic units (OTUs; 97% similarity) representing nine phyla. The relative abundance in gut microbiota shifted from more pathogenic bacteria-Helicobacter (NC: 22% versus MD: 3%) toward probiotic-Lactobacillus (NC: 19% versus MD: 40%). The immune organ index (spleen) and contents of secretory immunoglobulin A (S-IgA) were increased from small intestine. In conclusion, WO decreased the oxidative stress, inflammation, and improved the immunity and beneficial gut microbiota in the mice. PRACTICAL APPLICATIONS: Walnut oil (WO) is widely used in traditional medicine around the world and is prescribed as beneficial food oil in agro-industry. However, the intestinal benefits of WO have not been explored extensively, and even its therapeutic mechanism still remains unknown in modern medicine. In this study, WO from Juglans sigillata was investigated for its preventive and protective effects on the intestinal mucosa in mice including anti-oxidant, anti-inflammatory, immunity, and gut microbiota modulation. WO decreased the oxidative stress, inflammation, and improved immunity and beneficial gut microbiota in the mice. WO has shown strong probiotic effect on the gut, and thus, can be considered as a potential candidate in food. The study outcome would enhance utilization of WO for the prevention of gastrointestinal diseases (e.g., Helicobacter, etc.) both in animals and human (inflammatory bowel diseases, IBD) and the formulation of functional foods.

Comparison of Growth Characteristics and Genomics of Two Canine Distemper Virus Strains Isolated From Minks in China.

Canine distemper (CD), caused by the CDV variant strain with HI542N/Y549H, has become an epidemic in fur-bearing animals in China since 2012. To well understand the genomic and replicated characteristics of the CDV variants, we determined the viral growth kinetics and completed the genome sequences of two CDV strains, namely SDZC(17)M2 and LNDL(17)M4, isolated from CDV-infected minks from Shandong and Liaoning province in China, in 2017. SDZC(17)M2 showed higher viral titers and extensive syncytia in BHK-minkSLAM (BMS) cells than LNDL(17)M4. Although both two strains belong to the Asia-1 genotype and clustered an independent clade in the phylogenetic tree, SDZC(17)M2, harboring I542N/Y549H substitutions in the H protein, shared high identity (99.3-99.6% nt) with the other variant strains, whereas LNDL(17)M4, with the only Y549H substitution, shared a lower identity (97.7%-97.9% nt) with the other variant strains. Furthermore, a novel R223K substitution was identified in the conserved cleavage site (RRQRR → RRQKR) of the F protein in the SDZC(17)M2 strain. However, it which did not significantly affect the cell to cell fusion activity when combined with the CDV H/minkSLAM in BHK-21 cells. The key variations in the genome contributed to the virulence and the evolutionary trend need to be determined in the future.

Identification of proteins that mediate the role of androgens in antler regeneration using label free proteomics in sika deer (Cervus nippon).

Deer antlers offer a unique model to study organ regeneration in mammals. Antler regeneration relies on the pedicle periosteum (PP) cells and is triggered by a decrease in circulating testosterone (T). The molecular mechanism for antler regeneration is however, unclear. Label-free liquid chromatography-mass spectrometry (LC-MS/MS) was used to identify differentially-expressed proteins (DEPs) in the regeneration-potentiated PP (under low T environment) over the non-regeneration-potentiated PP (under high T environment). Out of total 273 DEPs, 189 were significantly up-regulated and 84 were down-regulated from these comparisons: after castration vs before castration, natural T vs before castration, and exogenous T vs before castration. We focused on the analysis only of those DEPs that were present in fully permissive environment to antler regeneration (low T). Nine transduction pathways were identified through the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, including the estrogen signaling pathway. A total of 639 gene ontology terms were found to be significantly enriched in regeneration-potentiated PP (low T) from the DEPs. Reliability of the label free LC-MS/MS was determined by qRT-PCR to estimate the expression level of selected genes. The results suggest that up-regulated heat shock proteins (HSP90AB1, HSP90B1), peptidyl-prolyl cis-trans isomerase 4 (FKBP4), mitogen-activated protein kinase 3 (MAPK3) and calreticulin (CALR) and down-regulated SHC-transforming protein 1 (SHC1), heat shock protein family A member 1A (HSPA1A) and proto-oncogene tyrosine-protein kinase (SRC) may be associated directly or indirectly with antler regeneration. Further studies are required to investigate the roles of these proteins in regeneration using appropriate in vivo models.

Effect of kisspeptin-10, LH and hCG on serum testosterone concentrations in stallions, donkeys and mules.

This study was conducted to determine the response of serum testosterone (T) in male equines (stallions, donkeys and mules) after administering intravenous doses of kisspeptin-10 (KP-10), human chorionic gonadotropin (hCG) and luteinizing hormone (LH) and saline as a control. The animals were divided into four groups of three each: Group I, 3 ml of 0.95% saline; Group II, 50 µg KP-10; Group III, 2500 IU hCG and group IV, 400 µg LH. The administration of KP-10 and hCG to stallions resulted in a significant increase in serum T concentration at 240 min; whereas it was significantly higher at 30, 60, 120, and 240 min with LH treatment as compared to pre-dose concentrations. Both KP-10 and hCG significantly elevated the T concentrations in donkeys at 120 and 240 min, respectively; whereas it was significantly higher at 60, 120, and 240 min with LH treatment as compared to pre-dose concentration. Both KP-10 and LH elevated T in donkeys at 240 min as compared to the control and hCG concentrations. After 120 and 240 min, T concentrations in mules were higher (p < 0.05) with administration of KP-10, hCG and LH as compared to the control. In conclusion, the administration of KP-10, hCG and LH elevate the serum T concentration in normal male equines. It is suggested that KP-10 may be useful in situations where an increase in T is desired. Further work is required to determine the effect of KP-10 on T in male equids with reproductive abnormalities before it can be used in clinical situations.

Cryoprotection synergism between glycerol and dimethyl sulfoxide improves the mitochondrial transmembrane potential, plasmalemma, acrosomal and DNA integrities, and in vivo fertility of water buffalo (Bubalus bubalis) spermatozoa.

The objective of the study was to devise a cryoprotection synergism between glycerol and dimethyl sulfoxide (DMSO) for water buffalo spermatozoa. Additionally, the effect of best evolved concentrations of glycerol and DMSO in extender was assessed on in vivo fertility of buffalo spermatozoa. Ejaculates (n = 30) were equally distributed into five aliquots; first aliquot was diluted at 37 °C in extender having 7 % glycerol (control); second aliquot was diluted at 37 °C as well as at 4 °C in extender having 3.5 % DMSO (Group 1); third aliquot was diluted at 37 °C in extender having 3.5 % glycerol and then at 4 °C in extender having 3.5 % DMSO (Group 2); fourth aliquot was diluted at 37 °C in extender having 3.5 % DMSO and then at 4 °C in extender having 3.5 % glycerol (Group 3); fifth aliquot was diluted in extenders having 1.75 % glycerol and 1.75 % DMSO at 37 as well as at 4 °C (Group 4). At post thawing, sperm progressive motility (%), rapid velocity (%), average path velocity (µm/s), curved line velocity (µm/s), in vitro longevity (%), structural and functional integrity of plasmalemma (%), mitochondrial transmembrane potential (%) and viable sperm with intact acrosome (%) were higher (P < 0.05) in Group 4 compared to other treatment groups and control. Regarding sperm DNA integrity (%); it was higher (P < 0.05) in Group 4 compared to Group 1, 3 and control. The in vivo fertility (%) of buffalo spermatozoa was significantly higher with Group 4 compared to control (69.45 vs. 59.81). In conclusion, synergism exists between glycerol and DMSO (Group 4) in improving the quality and in vivo fertility of cryopreserved water buffalo spermatozoa.