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Heavy metals intoxication causes several health problems that necessitate finding new protective and therapeutic approaches. This study aimed to evaluate the impact of Musa sp. leaves extract (MLE) on hepato-renal toxicities induced by cadmium (Cd) in male mice. The phytochemical screening, metal chelating activity (MCA), and the median lethal dose (LD50) of MLE were determined. Fifty CD-1 male mice were used and intraperitoneally (i.p.) injected with MLE (1000 to 5000 mg/kg b.wt) for MLE LD50 determination. Another 50 mice were used for evaluating the effect of MLE on Cd toxicity. Blood samples were collected for hematological, liver, and kidney functions assessments. Liver tissue homogenates were used for determination of oxidant/antioxidant parameters. Liver and kidney tissues were harvested for histopathological and molecular investigations. MLE showed potent in vitro antioxidant activities. The MCA and LD50 of the MLE were 75 µg/mL and 3000 mg/kg b.wt, respectively. MLE showed beneficial therapeutic activity against hepato-renal toxicities in Cd-intoxicated mice, evidenced by improving the hematological, biochemical, histopathological, and molecular alterations.

Intestinal acute rejection (AR) lacks a reliable non-invasive biomarker and AR surveillance is conducted through frequent endoscopic biopsies. Although citrulline and calprotectin have been suggested as AR biomarkers, these have limited clinical value. Using a mouse model of intestinal transplantation (ITx), we performed a proteome-wide analysis and investigated rejection-related proteome changes that may eventually be used as biomarkers. ITx was performed in allogenic (Balb/C to C57Bl) and syngeneic (C57Bl) combinations. Graft samples were obtained three and six days after transplantation (n = 4/time point) and quantitative proteomic analysis with iTRAQ-labeling and mass spectrometry of whole tissue homogenates was performed. Histology showed moderate AR in all allografts post-transplantation at day six. Nine hundred and thirty-eight proteins with at least three unique peptides were identified in the intestinal grafts. Eighty-six proteins varying by >20% between time points and/or groups had an alteration pattern unique to the rejecting allografts: thirty-seven proteins and enzymes (including S100-A8 and IDO-1) were significantly upregulated whereas forty-nine (among other chromogranin, ornithine aminotransferase, and arginase) were downregulated. Numerous proteins showed altered expression during intestinal AR, several of which were previously identified to be involved in acute rejection, although our results also identified previously unreported proteome changes. The metabolites and downstream metabolic pathways of some of these proteins and enzymes may become potential biomarkers for intestinal AR.

Background This study determined whether the naturally attenuated, thermotolerant Newcastle disease vaccine virus I-2 could acquire virulence after five in vivo passages through SPF chickens. Methods Study design was to international requirements including European Pharmacopoeia, Ph. Eur., v9.0 04/2013:0450, 2013. I-2 Working Seed (WS) was compared with five-times-passaged I-2 WS (5XP WS) in intracerebral pathogenicity index (ICPI), Fo cleavage site sequencing and Safety tests. Results The first passage series used a 50% brain: 50% tracheal tissue challenge homogenate and was unsuccessful as I-2 was not detected after the fourth passage. A second passage series used 10% brain: 90% tracheal tissue homogenates. I-2 was isolated from tracheal tissue in each passage. However harvested titres were below the minimum challenge level (107 EID50) specified for the ICPI and Safety tests, possibly reflecting I-2’s inherently low pathogenicity (interestingly caecal tonsils yielded significant titres). Given this the WS and 5XP WS comparisons proceeded. ICPI values were 0.104 and 0.073 for the WS group and the 5XP WS group respectively confirming that I-2, whether passaged or not, expressed low pathogenicity. F0 amino-acid sequences for both WS and 5XP WS were identified as 112R-K-Q-G-R-↓-L-I-G119 and so compatible with those of avirulent ND viruses. In safety, no abnormal clinical signs were observed in both groups except for two chicks in the 5XP WS group, where one bird was withdrawn due to a vent prolapse, and another bird died with inconclusive necropsy results. Conclusions: These data, the issue of low passage titres with little or no virus isolation from brain tissues and the genomic copy approach suggest a need to amend Ph. Eur. v9.0 04/2013:0450, 2013 for naturally attenuated, low pathogenicity vaccine viruses such as I-2. From an international regulatory perspective, the study provides further definitive data demonstrating that Newcastle disease vaccine virus I-2 is safe for use.

The mechanisms underlying neutrophilic inflammation in chronic rhinosinusitis with nasal polyps (CRSwNP) remain poorly investigated. This study aimed to examine the factors that contribute to tissue neutrophilia in CRSwNP. The numbers of neutrophils and active caspase-3-positive apoptotic neutrophils in sinonasal tissues were assessed via immunofluorescence staining. The 95th percentile of tissue neutrophil numbers in control subjects was selected as a cut-off to define neutrophil-high (Neu-high) or neutrophil-low (Neu-low) nasal polyps (NPs). The levels of 34 inflammatory mediators in sinonasal tissues were analyzed using Bio-Plex assay. Purified human peripheral blood neutrophils were incubated with nasal tissue homogenates, and the apoptotic neutrophils were assessed via flow cytometry. The cut-off for Neu-high NPs was >10 myeloperoxidase positive cells/high-power field. Compared with Neu-low NPs, Neu-high NPs had higher tissue levels of IL-1β, IL-1Ra, IL-6, IL-8, G-CSF, MCP-1, and MIP-1α, but lower levels of IL-5, IL-13, IgE, and eosinophils. Principal component and multiple correspondence analyses revealed mixed type 1, type 2, and type 3 endotypes for Neu-low NPs, and predominant type 1 and type 3 endotypes for Neu-high NPs. Neu-high NPs had lower percentages of apoptotic neutrophils than Neu-low NPs. The numbers of neutrophils and the percentages of apoptotic neutrophils correlated with G-CSF and IL-6 levels in the NPs. Tissue homogenates from Neu-high NPs, but not those from Neu-low NPs, suppressed neutrophil apoptosis in vitro, which was reversed by anti-G-CSF treatment. Tissue neutrophil numbers were associated with difficult-to-treat disease in patients with CRSwNP after surgery. We propose that G-CSF promotes neutrophilic inflammation by inhibiting neutrophil apoptosis in CRSwNP.

Purpose: To determine the influence of fasudil on LPS-mediated acute kidney injury (AKI) in mice.Methods: Healthy C57 mice (n = 140) of largely similar weight were used in this study. They were assigned to a treatment group (n = 40), a model group (n = 50), and a blank control group (n = 50). Mice in treatment and model groups were injected with lipopolysaccharide (LPS). In the treatment group, each mouse was injected intravenously with fasudil daily before the establishment of the mouse model of AKI. All mice were sacrificed 6 h after establishing the AKI model. Portions of the kidney from mice were used for preparation of tissue homogenates, while the remaining portions were subjected to primary culture. Transformed C3H Mouse Kidney-1 (TCMK1) and mesangial cells from mouse glomeruli (SV40-MES-13) cells were used for assays of cell growth and apoptosis. Blood samples were alsocollected from the mice. Thereafter, the levels of blood urea nitrogen (BUN) and creatinine (Cr) in kidney homogenates of the three groups were determined. Moreover, levels of NLRP3, nuclear factor kappa-B (NF-κB), toll-like receptor 4 (TLR4), tumor necrosis factor-α (TNF-α), interleukin (IL)-6, and IL-1β in the homogenates and blood were assayed. Cell growth and apoptosis were also measured.Results: The treatment group and model group showed higher levels of BUN and Cr than the control group, with a higher level observed in model mice than in the treatment mice. There were significantly higher relative levels of NF-κB, NLRP3 and TLR4 in treatment and model groups than in controls, with a higher level observed in model mice than in treatment mice. There were significantly higher concentrations of inflammatory factors in treatment and model mice groups than in control mice, with higher levels observed in model mice than in treatment mice. The TCMK1 and SV40-MES-13 cells in the two groups showed slower cell growth and stronger apoptosis than those in control group (p < 0.05).Conclusion: Fasudil relieved LPS-mediated AKI in mice by suppressing TLR4/NF-κB signal pathway and lowering NLRP3. Thus, fasudil has potential as a new adjunctive agent for the treatment of AKI.

Seed transmission of endophytic microorganisms is a growing research area in plant biology and microbiology. We employed cultivation versus cultivation-independent approaches on excised embryos from watermelon seeds (6–12 months in storage) and on embryo-derived in vitro seedlings (EIVS) to assess the vertical transmission of endophytic bacteria. Surface-disinfected watermelon seeds bore abundant residual bacteria in the testa and perisperm tissues, predominantly Bacillus spp. propounding the essentiality of excluding all non-embryonic tissues for vertical transmission studies. Tissue homogenates from re-disinfected seed embryos displayed no cultivable bacteria during the 1-week monitoring. Bright-field live microscopy revealed abundant bacteria in tissue homogenates and in embryo sections as intracellular motile particles. Confocal imaging on embryo sections after SYTO-9 staining and eubacterial fluorescent in situ hybridization (FISH) endorsed enormous bacterial colonization. Quantitative Insights Into Microbial Ecology (QIIME)-based 16S rRNA V3–V4 taxonomic profiling excluding the preponderant chloroplast and mitochondrial sequences revealed a high bacterial diversity in watermelon seed embryos mainly Firmicutes barring spore formers followed by Proteobacteria, Bacteroidetes, and Actinobacteria, and other minor phyla. Embryo-base (comprising the radicle plus plumule parts) and embryo-cotyledon parts differed in bacterial profiles with the abundance of Firmicutes in the former and Proteobacteria dominance in the latter. EIVS displayed a higher bacterial diversity over seed embryos indicating the activation from the dormant stage of more organisms in seedlings or their better amenability to DNA techniques. It also indicated embryo-to-seedling bacterial transmission, varying taxonomic abundances for seed embryos and seedlings, and differing phylogenic profiles for root, hypocotyl, and cotyledon/shoot-tip tissues. Investigations on different watermelon cultivars confirmed the embryo transmission of diverse cultivation recalcitrant endophytic bacteria. Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes formed the core phyla across different cultivars with 80–90% similarity at genus to phylum levels. Conversely, freshly harvested seeds displayed a dominance of Proteobacteria. The findings revealed that dicot seeds such as in different watermelon cultivars come packaged with abundant and diverse vertical and seedling-transmissible cultivation recalcitrant endophytic bacteria with significant implications for plant biology.

Background This study determined whether the naturally attenuated, thermotolerant Newcastle disease vaccine virus I-2 could acquire virulence after five in vivo passages through SPF chickens. Methods Study design was to international requirements including European Pharmacopoeia, Ph. Eur., v9.0 04/2013:0450, 2013. I-2 Working Seed (WS) was compared with five-times-passaged I-2 WS (5XP WS) in intracerebral pathogenicity index (ICPI), Fo cleavage site sequencing and Safety tests. Results The first passage series used a 50% brain: 50% tracheal tissue challenge homogenate and was unsuccessful as I-2 was not detected after the fourth passage. A second passage series used 10% brain: 90% tracheal tissue homogenates. I-2 was isolated from tracheal tissue in each passage. However harvested titres were below the minimum challenge level (107 EID50) specified for the ICPI and Safety tests, possibly reflecting I-2’s inherently low pathogenicity (interestingly caecal tonsils yielded significant titres). Given this the WS and 5XP WS comparisons proceeded. ICPI values were 0.104 and 0.073 for the WS group and the 5XP WS group respectively confirming that I-2, whether passaged or not, expressed low pathogenicity. F0 amino-acid sequences for both WS and 5XP WS were identified as 112R-K-Q-G-R-↓-L-I-G119 and so compatible with those of avirulent ND viruses. In safety, no abnormal clinical signs were observed in both groups except for two chicks in the 5XP WS group, where one bird was withdrawn due to a vent prolapse, and another bird died with inconclusive necropsy results. Conclusions: These data, the issue of low passage titres with little or no virus isolation from brain tissues and the genomic copy approach suggest a need to amend Ph. Eur. v9.0 04/2013:0450, 2013 for naturally attenuated, low pathogenicity vaccine viruses such as I-2. From an international regulatory perspective, the study provides further definitive data demonstrating that Newcastle disease vaccine virus I-2 is safe for use.

Abstract Background: Betanodaviruses , members of the Nodaviridae family, are the causative agents of viral nervous necrosis (VNN) in fish, resulting in great economic losses worldwide. Methods: In this study, we isolated a virus strain named seahorse nervous necrosis virus (SHNNV) from cultured big-belly seahorses Hippocampus abdominalis in Xiamen city, Fujian Province, China. Cell isolation, PCR detection, phylogenetic analysis, qRT-PCR, fluorescence in situ hybridization (FISH) and histology were used for virus identification and analysis of virus histopathology. Furthermore, an artificial infection experiment was conducted for virulence testing. Results: Brain and eye tissue homogenates of diseased big-belly seahorses were inoculated onto a grouper spleen (GS) cell monolayer at 28°C. Tissue homogenates induced obvious cytopathic effects in GS cells. PCR and sequencing analyses revealed that the virus belonged to Betanodavirus and shared high sequence identity with red-spotted grouper nervous necrosis virus (RGNNV) isolates. qRT-PCR and fluorescence in situ hybridization revealed that SHNNV mainly attacked the brain and eye. Histopathological examination revealed that the virus led to cytoplasmic vacuolation in the brain and retinal tissues. Infection experiments confirmed that SHNNV was highly infectious, causing massive death in big-belly seahorses. Conclusion: A novel seahorse betanodavirus from the big-belly seahorse in China was discovered. This finding will contribute to the development of efficient strategies for disease management in aquaculture.

In order to determine the frequency of SARS-CoV-2 placental and fetal involvements, we analyzed placentas of 197 women positive for infection at delivery and fetal tissues in cases of pregnancy loss in women positive by SARS-CoV-2 PCR (N = 2) and COVID-19 serology (N = 4), using in situ hybridization (ISH), immunohistochemistry (IHC) and, in selected cases, RT-PCR of tissue homogenates. The virus was identified in situ, accompanied by intervillositis, in 2 of 197 placentas (1.02%). In three more cases, SARS-CoV-2 was detected by tissue PCR without in situ localization and placental inflammation. There were no maternal mortality or association of placental infection with the clinical severity of COVID-19. All tested neonates born to SARS-CoV-2-positive women (N = 172) were negative for the virus. There were three pregnancy losses among 197 infected women and in two cases available fetal tissues were negative for SARS-CoV-2. In one of four fetal autopsies performed in women with positive COVID-19 serology, the mother-to-child transmission (MTCT) could be inferred based on positive SARS-CoV-2 nucleocapsid IHC in fetal pulmonary endothelium. Placental involvement by SARS-CoV-2 is rare, but may be underestimated due to its transient nature. MTCT is even rarer, supporting the protective role of placenta in SARS-CoV-2 infection.

Amyloidosis is diagnosed by the histologic detection of amyloid deposits; however, this method has limitations such as a prolonged diagnosis time and the need for histologic proficiency. We aimed to develop a rapid and simple method for diagnosing amyloidosis by targeting amyloid-specific endogenous fluorescence, which has not been reported previously, to our knowledge. Fluorescence fingerprint analysis of amyloid extracts and tissue homogenates derived from amyloid A (AA) amyloidosis-affected cattle exhibited a specific intrinsic fluorescence pattern. Furthermore, principal component analysis using analytical data revealed that AA could be identified by peaks near λex 350 nm and λem 430 nm. Fluorescence spectrometry analysis using tissue homogenates, which does not require special histochemical staining, enables the rapid detection of bovine AA.