Development of a method for the isolation and culture of astrocytes from the canine cerebral cortex.

BACKGROUND: Astrocytes are considered key players in neuroimmunopathological processes, and they play a certain role in neuroinflammation. Rodent primary astrocyte cultures are commonly used in the study of human neuroinflammation. However, gene sequence homologies are closer between humans and dogs than between humans and rodents. NEW METHOD: We established protocols to isolate astrocytes from the canine forebrain. Cerebral hemispheres of 3-4-week-old dogs were used. The isolation procedure included the use of the Neural Tissue Dissociation Kit P, demyelination by the magnetic bead method, and separation and preparation by differential adhesion. RESULTS: We found a 96% astrocyte purification rate after isolation by differential adhesion. Purified canine astrocytes increased the secretion of interleukin-1ß, interleukin-6, and tumor necrosis factor-alpha, and increased the expression of glial fibrillary acidic protein after lipopolysaccharide stimulation. We sequenced the transcriptome of the purified canine astrocytes and analyzed the differentially expressed genes among the rodent, human, and canine astrocytes. Transcriptome profiling and gene ontology analysis of the genes co-expressed in humans and canines indicate that human and canine astrocytes may be different from their rodent counterparts in terms of mediated interactions with metals. COMPARED WITH THE EXISTING METHODS: The cells prepared by our method allow for the rapid separation of astrocytes with a relatively small resource scheme. The method also retains the cell phenotype and has an in vitro culture lifetime of approximately 2-3 months. CONCLUSION: We established a method for preparing canine astrocytes with high purity, which can be used to study the biological function of astrocytes in vitro.

Mechanism and therapeutic prospect of resveratrol combined with TRAIL in the treatment of renal cell carcinoma.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been reported to kill a wide variety of tumor cells with minimal effects on normal cell. However, renal cell carcinoma (RCC) cells 786-0 and OS-RC-2 were resistant to TRAIL. The present study examines the potential of combining polyphenolic compound resveratrol (RES) with TRAIL. We found that RES can sensitize RCC cells to TRAIL-induced death. Electron microscopy analyses showed that RES plus TRAIL can induce both autophagy and apoptosis in RCC cells. It was proved that the apoptosis is caspase-dependent and the activation of caspase-8, caspase-9, and caspase-3 was involved in this process. Besides, we also found that XIAP expression was significantly inhibited after RES plus TRAIL treatment in RCC cells. Furthermore, a fiber-modified replication-deficient adenovirus Ad5/35-TRAIL was generated to test the synergistic effect of RES and TRAIL in vivo. Our data demonstrated that RES plus Ad5/35-TRAIL significantly inhibited RCC xenograft growth in nude mice. These results suggest the possibility of a new combination therapeutic leading to the improvement of RCC treatment.

Induction and Suppression of Innate Antiviral Responses by Hepatitis A Virus.

Hepatitis A virus (HAV) belongs to the family Picornaviridae. It is the pathogen of acute viral hepatitis caused by fecal-oral transmission. RNA viruses are sensed by pathogen-associated pattern recognition receptors (PRRs) such as Toll-like receptor 3 (TLR3), retinoic acid-inducible gene I (RIG-I), and melanoma differentiation-associated gene 5 (MDA5). PRR activation leads to production of type 1 interferon (IFN-α/ß), serving as the first line of defense against viruses. However, HAV has developed various strategies to compromise the innate immune system and promote viral propagation within the host cells. The long coevolution of HAV in hosts has prompted the development of effective immune antagonism strategies that actively fight against host antiviral responses. Proteases encoded by HAV can cleave the mitochondrial antiviral signaling protein (MAVS, also known as IPS-1, VISA, or Cardif), TIR domain- containing adaptor inducing IFN-ß (TRIF, also known as TICAM-1) and nuclear factor-κB (NF-κB) essential modulator (NEMO), which are key adaptor proteins in RIG-I-like receptor (RLR), TLR3 and NF-κB signaling, respectively. In this mini-review, we summarize all the recent progress on the interaction between HAV and the host, especially focusing on how HAV abrogates the antiviral effects of the innate immune system.

In vitro and in vivo studies of the native isolates of nematophagous fungi from China against the larvae of trichostrongylides.

To screen potential nematophagous fungi candidates for the biological control of parasitic nematodes in livestock, in vitro and in vivo studies of the native isolates of nematophagous fungi against the larvae of trichostrongylides were conducted. The in vitro predatory activity of 16 native nematophagous fungal isolates on the larvae of trichostrongylides in sheep feces was assessed. In the ten isolates of Duddingtonia flagrans, the reduction percentage for the infective larvae (L3) of Trichostrongylus colubriformis ranged from 57.21 to 99.83%, and that of Haemonchus contortus ranged from 62.12 to 99.88%. The analysis of the same assay on five isolates of Arthrobotrys superba and one isolate of A. cookedickinson (Monacrosporium cystosporum) showed comparable results with those for D. flagrans. To determine the excretion time of fungal isolates in feces after oral administration, D. flagrans (SDH035) were studied in vivo in sheep and rabbits. Results showed that the tested fungal isolates existed in sheep feces from 12 to 72 h after fungal treatment, and the fungal excretion in rabbit feces occurred at 4 h, reached a peak at 10 h, and declined gradually 18 h after oral administration. All the native fungal isolates were assessed after passing through the gastrointestinal tract of sheep. Treatment with isolates of D. flagrans significantly reduced the number of developing larvae in the feces, and the efficacies ranged from 55.15 to 98.82%. One out of the five isolates of A. superba and A. cookedickinson (BS002) survived after passing through the gastrointestinal tract, and the L3 reduction rates were 83.79 and 81.33%, respectively. Results of the present study provide information about the in vitro predatory activity of nematophagous fungi from China on the L3 of trichostrongylides and their ability to pass through the gastrointestinal tract before administering them for biocontrol.