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The significance of the polypeptide N-acetyl-galactosaminyl transferase-3 (GalNAc-T3) and mucin 1 (MUC1) in solitary pulmonary adenocarcinoma (SPA) initially diagnosed as malignant solitary pulmonary nodule (≤ 3 cm), especially as a combined predictor of prognosis and recurrence, was explored in this study. A retrospective analysis of 83 patients with SPA (≤ 3 cm), which revealed postoperative pathological diagnosis was lung adenocarcinoma after complete resection. Immunohistochemical staining was used to detect the expression of GalNAc-T3 and MUC1 in primary tumor specimens. The relationship between expression and various clinicopathological factors was analyzed, as well as the effects of patients' overall survival (OS) and disease-free survival (DFS). In all patients, GalNAc-T3 was highly expressed in 53 (63.9%) cases; MUC1 was highly expressed in 31 (37.3%) cases. The GalNAc-T3 expression was correlated with differentiation, pathological risk group, N stage, and TNM stage. The group with high GalNAc-T3 expression and low MUC1 expression (GalNAc-T3 Hig /MUC1 Low ) is correlated to pathological differentiation and has a trend related to the TNM stage. The patients with better differentiation, lower pathological risk group, lower N stage, and GalNAc-T3 high expression had better overall survival, especially the GalNAc-T3 Hig /MUC1 Low group. Moreover, the moderate differentiation, N3 stage, and GalNAc-T3 Hig /MUC1 Low group were independent predictive factors for OS. Besides, patients with lower N stage, lower TNM stage, higher GalNAc-T3 expression got better disease-free survival (DFS), especially the GalNAc-T3 Hig /MUC1 Low group. The GalNAc-T3 Hig /MUC1 Low group was an independent predictive factor for DFS. In conclusion, GalNAc-T3 and MUC1 were combined predictors of prognosis and recurrence in SPA (≤ 3 cm).

Cervical cancer (CC) is a common gynecological malignancy, and improving cisplatin sensitivity has become a hot topic in CC chemotherapy research. Polyphyllin I (PPI), a potent bioactive compound found in Rhizoma Paridis, known for its anticancer properties, remains underexplored in CC resistance. In this study, we evaluated PPI's impact on cisplatin-resistant CC cells and elucidated its underlying mechanism. Our findings reveal that PPI enhances the sensitivity of cisplatin-resistant CC cells to the drug, promotes apoptosis, and inhibits cell migration. Mechanistically, PPI was found to regulate p53 expression and its target genes, and suppressing p53 expression reverses PPI's sensitizing effect in drug-resistant CC cells. In conclusion, PPI showed promise in sensitizing cisplatin-resistant human CC cells to cisplatin treatment, suggesting that it could serve as a potent adjunct therapy for cervical cancer, particularly for cases that have developed resistance to cisplatin, thereby providing a promising basis for further clinical investigation into PPI for enhancing the efficacy of existing chemotherapy regimens in resistant cervical cancer. [ABSTRACT FROM AUTHOR]

Objectives. To assess A-kinase anchor protein 95 (AKAP95), B-Raf, extracellular regulated protein kinases 1/2 (ERK1/2), and Elk-1 expression in colon cancer tissue, and characterize AKAP95 associations with B-Raf, ERK1/2, Elk-1, and colon cancer clinicopathological indices. Methods. The immunohistochemistry streptavidin-perosidase (SP) method was used to determine protein expression levels in 64 colon cancer and 32 para-carcinoma tissue specimens. Results. (1) Positive AKAP95 expression rates in colon cancer tissue were higher when compared with para-carcinoma tissue (92.19% vs. 59.38%, P < 0.05). Similar findings were determined for B-Raf (76.56% vs. 25%, P < 0.05), ERK1/2 (90.63% vs. 31.25%, P < 0.05), and Elk-1 levels (92.19% vs. 40.63%, P < 0.05). (2) No significant associations were identified between AKAP95, B-Raf, ERK1/2, and Elk-1 protein expression and degree of differentiation, histological type, and lymph node metastasis in colon cancer samples (P > 0.05); however, in The Cancer Genome Atlas and Gene Expression Omnibus datasets, AKAP95 was closely related to immune infiltration, and highly expressed AKAP95 was negatively associated with overall survival and relapse free survival rates in colon cancer patients. (3) Correlations were observed between AKAP95 and ERK1/2, AKAP95 and Elk-1, B-Raf and ERK1/2, B-Raf and Elk-1, and ERK1/2 and Elk-1 (all P < 0.05), but no correlation was observed between AKAP95 and B-Raf (P > 0.05). Conclusions. AKAP95 may affect immune infiltration levels in colon cancer by participating in ERK1/2–Elk-1 signal transduction. [ABSTRACT FROM AUTHOR]

Simple Summary: Andrias davidianus is one of the largest amphibian species in the world. To improve our understanding of Myxovirus resistance (Mx) genes in amphibians, particularly their function in virus infection, we cloned four full-length A. davidianus (adMx) cDNA sequences and characterized these genes by bioinformatics analysis and quantitative expression techniques. The four adMx genes ranged from 2008 to 2840 bp in length, and their conserved protein domains included the signature architectural feature of the dynamin superfamily. Deduced amino acid sequences exhibited relatively high sequence identity with Mx proteins from other vertebrates and phylogenetic analysis revealed close clustering with fish. The four adMx genes were broadly expressed in healthy A. davidianus, but differentially expressed in the spleen following Chinese giant salamander iridovirus (GSIV) infection. These findings imply that the adMx genes share major sequence and protein structures and similar functions with those of other species. Amphibians, including Andrias davidianus, are declining worldwide partly due to infectious diseases. The Myxovirus resistance (Mx) gene is a typical interferon (IFN)-stimulated gene (ISG) involved in the antiviral immunity. Therefore, knowledge regarding the antiviral immunity of A. davidianus can be used for improved reproduction in captivity and protection in the wild. In this study, we amplified and characterized four different A. davidianus Mx genes (adMx) and generated temporal mRNA expression profiles in healthy and Chinese giant salamander iridovirus (GSIV) infected A. davidianus by qualitative real-time PCR (qPCR). The four adMx genes ranged in length from 2008 to 2840 bp. The sequences revealed conserved protein domains including the dynamin superfamily signature motif and the tripartite guanosine-5-triphosphate (GTP)-binding motif. Gene and deduced amino acid sequence alignment revealed relatively high sequence identity with the Mx genes and proteins of other vertebrates. In phylogenetic analysis, the adMx genes clustered together, but also clustered closely with those of fish species. The four adMx genes were broadly expressed in healthy A. davidianus, but were differentially expressed in the spleen during the GSIV infection. Our results show that the adMx genes share major structural features with their homologs, suggesting similar functions to those in other species. [ABSTRACT FROM AUTHOR]

In immature lymphocytes, recombination activating genes 1 and 2 are necessary for antigen receptor V (D) J recombination, representing immature lymphocyte biomarkers. Herein, we cloned and sequenced rice-field eel rag1 and rag2 genes. Their expressions in the thymus, liver, and kidney were significant from 0 days post hatching (dph) to 45 dph, peaking at 45 dph in these three tissues. In situ hybridization detected high rag1 and rag2 expressions in the liver, kidney, and thymus of rice-field eel from 2 to 45 dph, suggesting that multiple tissues of rice-field eel contain lymphocyte lineage cells and undergo lymphopoiesis. Tissue morphology was used to observe lymphopoiesis development in these three tissues. The thymus primordium began to develop at 2 dph, while the kidney and liver have generated. Our findings verified that the thymus is the primary lymphopoietic tissue and suggested that, in rice-field eel, lymphocyte differentiation also occurs in the liver and kidney. [ABSTRACT FROM AUTHOR]

The cytoplasmic helicases retinoic acid‐inducible gene I (RIG‐I) and melanoma differentiation‐associated protein 5 (MDA5) are two pattern recognition receptor genes belonging to RIG‐I‐like receptor (RLR). They play a crucial role in host sensing of pathogen invasion. Herein, we identified and characterized complete cDNA of RIG‐I and MDA5 in Chinese giant salamander (adRIG‐I and adMDA5). To evaluate their expression levels, real‐time fluorescence quantitative RT‐PCR (qPCR) was performed between normal and Chinese giant salamander iridovirus (GSIV)‐infected tissues. The full‐length adMDA5 cDNA of 3,509 bp encoded a 968‐acid amino polypeptide, and the 3567‐bp adRIG‐I cDNA encoded a 951‐acid amino protein. The amino acid sequence analysis revealed adRIG‐I and adMDA5 kept the conserved domains of RLR, and the two genes both shared relatively high similarity homologs with other species. Two genes were expressed in most tissues of Andrias davidianus, with the highest expression in spleen. After GSIV infection, both adRIG‐I and adMDA5 transcripts were initially increased then decreased in kidney and spleen. These results suggest that adRIG‐I and adMDA5 can response to GSIV infection and may play a role in antiviral infections in DNA virus. [ABSTRACT FROM AUTHOR]

A novel GCRV strain isolated from healthy grass carp was named as grass carp reovirus - HH196 (GCRV-HH196), and its infection mechanism remains unclear. In this study, the grass carp ovary cell line (GCO cells) was used to investigate the cell death involved in GCRV-HH196 infection. The results showed that DNA damage, cells volume reduction and cytoplasm shrinkage happened during GCRV-HH196 infection. The mRNA expression levels of pro-apoptotic genes were up-regulated during infection. Two initiators of apoptosis, caspase 8 and caspase 9, and the executioner of apoptosis, caspase 3, were all significantly activated in GCRV-HH196-infected cells. Flow cytometry analysis showed that the number of apoptotic cells in infected cells was significantly higher than that in control cells as the infection progress. Meanwhile, autophagy was also involved in the regulation of GCRV - HH196 infection. We observed that LC3 puncta existed in cytoplasm in GCRV-HH196-infected cells. Furthermore, the protein level of LC3-Ⅱ and Beclin-1 increased, while that of p-Akt decreased in GCRV-HH196-infected cells. These results demonstrated that GCRV-HH196 may regulate apoptosis and autophagy for the virus proliferation and spread, which set a foundation for further research on the interaction between GCRV-HH196 and host. • Grass carp reovirus-HH196 (GCRV-HH196) infection leads to typical apoptosis GCO cells. • Caspases were activated during GCRV-HH196-induced apoptosis. • Pro-apoptotic genes levels up-regulated during GCRV-HH196-induced apoptosis. • LC3 protein shifted from diffuse distribution to punctate pattern in cytoplasmic upon infection. • The protein level of LC3-Ⅱ and Beclin-1 increased, while that of p-Akt decreased during infection. [ABSTRACT FROM AUTHOR]

Type I interferons, as a class of multipotent cytokines, play a key role in host antiviral immune responses. In this study, a type I IFN coding gene of gibel carp, Carassius auratus gibelio , Cag IFNa was cloned and sequenced. The full-length cDNA sequence of Cag IFNa consists of 724 nucleotides that encode a predicted protein of 183 amino acids. Cag IFNa has two highly conserved cysteine residues in the deduced protein, which is mostly conserved in the fish group I type I IFNs. Cag IFNa was identified as a member of the IFNa subgroup of group I type I IFNs by phylogenetic analysis. Cag IFNa transcripts were detected in all investigated tissues with higher levels in the liver, intestine, spleen and head kidney of gibel carp. Following injection with Cyprinid herpesvirus 2 (CyHV-2), Cag IFNa gene expression was significantly inhibited in the spleen but delayed and then increased in head kidneys. Similarly, while Cag IFNa expression was rapidly induced in gibel carp brain (GiCB) cells by poly I:C stimulation and its high induction level was delayed following CyHV-2 infection. Cag IFNa overexpression in GiCB cells drastically reduced virus CPE and titer. Furthermore, several genes associated with type I IFN signaling pathway including IRF3, IRF7, IRF9, STAT1, Mx1 and PKR were induced in GiCB cells overexpressing Cag IFNa upon CyHV-2 infection. These results show that Cag IFNa plays a role in antiviral immune system in gibel carp. • A type I IFN gene from gibel carp, Carassius auratus gibelio named Cag IFNa was cloned and characterized. • The expression of Cag IFNa was investigated with CyHV-2 infection or poy I:C stimulation. • The CyHV-2 propagation was remarkably inhibited in Cag IFNa over-expressed cells. • The genes associated with type I IFN system were significantly induced by Cag IFNa. [ABSTRACT FROM AUTHOR]

• Over-expression of AdBcl-w inhibited the apoptotic process during GSIV infection while RNAi of AdBcl-w promoted this process. • Over-expression of AdBcl-w can effectively reduce the replication of GSIV. • The interaction of AdBcl-w and AdBak reduced after GSIV infection. • Over-expression of AdBcl-w restrained the mRNA expression of molecules in p53 pathway during GSIV infection. B-cell lymphoma-2 (BCL-2) superfamily molecules play crucial roles in mitochondrial apoptosis induced by Chinese giant salamander iridovirus (GSIV). As an anti-apoptotic molecule in the BCL-2 family, the molecular mechanism of Bcl-w during GSIV infection remains unknown. In this study, we characterized for the first time an amphibian Bcl-w from Chinese giant salamander Andrias davidianus (AdBcl-w), and its function and regulatory mechanism during GSIV infection were investigated. AdBcl-w possesses the conserved structural features of Bcl-w and shares 35–54% sequence identities with other Bcl-w. mRNA expression of AdBcl-w was most abundant in liver and muscle. The AdBcl-w mRNA expression was regulated during GSIV infection. Western blotting assays revealed that the level of Bcl-w protein was downregulated markedly as the infection progresses. Confocal microscopy showed that overexpressed AdBcl-w was translocated to the mitochondria after infection with GSIV. Flow cytometry analysis demonstrated that compared with control, the apoptotic progress in cells transfected with AdBcl-w was reduced while that in cells transfected with AdBcl-w siRNA was enhanced. The number of virus major capsid protein gene copies was lower and protein synthesis was reduced in AdBcl-w overexpressing cells. In addition, AdBcl-w could bind directly to the pro-apoptotic molecule AdBak, while this interaction was weakened with GSIV infection. Moreover, p53 level was reduced and the mRNA expression levels of crucial regulatory molecules in the p53 pathway were regulated in AdBcl-w overexpressing cells during GSIV infection. These results suggested that AdBcl-w inhibit GSIV replication by regulating the virus induced mitochondrial apoptosis. [ABSTRACT FROM AUTHOR]

Rag1 and rag2 are two closely linked recombination activating genes required for V(D)J recombination of antigen receptors in immature lymphocytes, whose expression can serve as marker to identify the lymphopoietic tissues. To study the development of lymphopoietic tissues in Chinese giant salamander ( Andrias davidianus ), the Chinese giant salamander rag1 and rag2 coding sequences were cloned and determined. High transcript levels of rag1 and rag2 were co-detected in the thymus before 14 months of age, whereas levels were lower in spleen, liver and kidney at all stage of development. The spatial expression patterns of rag1 and rag2 were studied in combination with igY and tcrβ gene expression using in situ hybridization. Significant transcript signals for rag1 , rag2 , tcrβ and igY were detected not only in the thymus and spleen but also the liver and kidney of juvenile and adult Chinese giant salamanders, which suggests that cells of lymphocyte lineage are present in multiple tissues of the Chinese giant salamander. This implies that lymphopoiesis may take place in these tissues. The tissue morphology of thymus suggested that the branched thymic primordium developed into mature organ with the development of thymocyte from juvenile to adult. These results not only confirm that as expected the thymus and spleen are primordial lymphopoietic tissues but also suggest that the liver and kidney provide site of lymphocyte differentiation in Chinese giant salamander. [ABSTRACT FROM AUTHOR]

The Chinese giant salamander, Andrias davidianus , is the largest extant amphibian species in the world, which is of significance due to its specific position in the evolutionary history of vertebrates. Currently, limited information about the innate immune system of this animal is known. In this study, the toll-like receptor 7 (TLR7), designated CgsTLR7, was cloned from Chinese giant salamander, A. davidianus . The full-length cDNA of CgsTLR7 is 3747 bp, with an open reading frame of 3150 bp, encoding 1049 amino acids. The TLR family motifs, including the leucine-rich repeat (LRR) and Toll/interleukin (IL)-1 receptor (TIR) domain are conserved in CgsTLR7, which includes 19 LRRs and a TIR domain. The predicted amino acid sequence of CgsTLR7 has 71%, 65%, 63% and 55% identity with turtle, chicken, human and fugu TLR7 homologues, respectively. Phylogenetic analysis showed that CgsTLR7 is closest to that of frog TLR7 among the examined species. Quantitative real-time PCR analysis revealed broad expression of CgsTLR7 in tissues from apparently healthy Chinese giant salamanders with the highest expression in the liver and the lowest expression in the intestine. The mRNA expression was up-regulated and reached a peak level in the kidney, liver and spleen at 12 h, 24 h and 48 h after infecting the animals with the giant salamander iridovirus (GSIV), respectively. These results suggest that CgsTLR7 has a conserved gene structure and might play an important role in immune regulation against viral infections in the Chinese giant salamander. [ABSTRACT FROM AUTHOR]