Association between bladder cancer treatment and female sexual function.

INTRODUCTION: Bladder cancer ranks 17th in prevalence of cancer types among women, and the trend is rising. The increased risk of female sexual dysfunction (FSD) after radical cystectomy (RC) underscores the need for greater focus on preserving and mitigating FSD. OBJECTIVES: To place greater emphasis on the importance of female sexual function (FSF) in the treatment of bladder cancer and stimulate additional research to discover more effective solutions for enhancing the overall quality of life. METHODS: This review used a narrative approach. Previous reviews on FSF after RC have provided limited and 1-sided solutions due to the lack of research. What makes this review unique is its innovative approach: it includes all available measures curing FSD as well as comparative analyses based on experimental data, thus making the findings more comprehensive. A detailed perspective of treatments for female bladder cancer is provided, including nerve- and organ-sparing RC, robot-assisted RC, and radiotherapy. We also analyze the impact of treatments for female bladder cancer on postoperative FSD. Additionally, solutions for addressing or alleviating postoperative FSD are summarized, such as urinary diversion, vaginal reconstruction, and drug and nondrug treatment. RESULTS: Research has suggested that robot-assisted nerve- and organ-sparing RC is promising. Moreover, orthotopic neobladder among urinary diversions without a stoma helps to maintain a positive female body image. If part of the anterior vaginal wall must be removed during RC, vaginal reconstruction can restore the dimensions with synthetic grafts and biologic scaffolds. Additionally, postoperative measures, such as vaginal laser and hormone therapy, and use of vaginal dilators and lubricants have a significant role in reducing distress caused by FSD to provide maximum relief. CONCLUSIONS: To support FSF after RC, various interventions are needed, and urologists must focus on patient recovery while minimizing treatment impact on FSF as much as possible.

E2F3 induces DNA damage repair, stem-like properties and therapy resistance in breast cancer.

Therapy resistance is a major hurdle to the treatment of human malignant tumors. Both DNA damage repair and stem-like properties contribute to chemoresistance and radioresistance. E2F transcription factor 3 (E2F3) is overexpressed in breast cancer tissues, and promotes proliferation of breast cancer cells. Higher E2F3 level is associated with shorter survival of breast cancer patients. Functional studies further showed that E2F3 promotes S-phage entry, DNA replication, DNA damage repair and stem-like properties. Accordingly, E2F3 knockdown sensitizes breast cancer cells to DNA-damaging agents Adriamycin, Cisplatin, Olaparib and X-ray. Forkhead box M1 (FOXM1) is a downstream molecule of E2F3 signaling, mediating the effects of E2F3 on breast cancer cells. In an m6A methyltransferase METTL14-dependent manner, YTH RNA binding protein F2 (YTHDF2) increase E2F3 mRNA stability and expression, promotes DNA damage repair and induces therapy resistance. These data demonstrate that YTHDF2-E2F3 pathway is a novel target to overcome chemoresistance and radioresistance in breast cancer.

Exogenous pancreatic kininogenase protects against tacrolimus-induced renal injury by inhibiting PI3K/AKT signaling: The role of bradykinin receptors.

BACKGROUND: Tissue kallikrein offers a wide spectrum of biological activity in the protection against various types of injury. However, information on its role in tacrolimus (TAC)-induced renal injury is limited. OBJECTIVES: This study aimed to assess the beneficial effects of pancreatic kininogenase (PK) in a rat model of chronic TAC nephrotoxicity and in vitro. METHODS: Sprague Dawley rats were treated daily with either TAC or PK or a combination of the two for four weeks. The influence of PK on renal injury was examined in terms of renal function, histopathology, cytokine expression, oxidative stress, intracellular organelles, programmed cell death, and PI3K/AKT signaling. Human kidney proximal tubular (HK-2) cells and mouse mesangial (SV40 MES13) cells treated with TAC and PK were also studied. RESULTS: PK treatment improved renal function and histopathology. This effect was paralleled by downregulation of proinflammatory and profibrotic cytokine expression. TAC-induced oxidative stress was closely associated with endoplasmic reticulum stress and mitochondrial dysfunction, resulting in excessive programmed cell death (apoptosis and autophagy) that was significantly abrogated by concurrent PK interference with PI3K/AKT signaling. PK also stimulated bradykinin receptor 1 (B1R) and B2R mRNA synthesis and increased bioactive nitric oxide (NO) and cAMP concentrations in TAC-treated kidneys. Blockade of either B1R or B2R eliminated the renoprotective effects of PK. In HK-2 and SV40 MES13 cells, PK decreased TAC-induced overproduction of intracellular reactive oxygen species and inhibited apoptotic cells, whereas cell viability was improved. Moreover, activated PI3K/AKT signaling in HK-2 cells was inhibited by PK and the PI3K inhibitor, LY294002. CONCLUSIONS: These findings indicate that PK treatment protects against chronic TAC nephrotoxicity via inhibition of PI3K/AKT signaling.

WNT7B represses epithelial-mesenchymal transition and stem-like properties in bladder urothelial carcinoma.

BACKGROUND: Recurrence and metastasis are the major problems of bladder urothelial carcinoma, which mainly attribute to tumor cell stemness, epithelial-mesenchymal transition (EMT) and chemoresistance. METHODS: TCGA database was interrogated for gene mRNA expression in bladder urothelial carcinoma samples. CCLE database was interrogated for gene mRNA expression in bladder cancer cell lines. The correlation between two genes was analyzed by Pearson statistics. 37 human bladder urothelial carcinoma specimens were adopted for immunohistochemistry. Bladder cancer cells RT4, J82, and UM-UC-3 were used to carry out loss and gain of function studies. Kaplan-Meier method was performed to analyze the overall survival. FINDINGS: WNT7B is downregulated in high-grade bladder urothelial carcinomas. Low WNT7B expression is associated with unfavorable prognosis. Loss and gain of function studies showed that WNT7B inhibits bladder urothelial carcinoma cell EMT, stem-like properties and chemoresistance. FZD5, a specific receptor for WNT7B, mediates WNT7B signaling. ELF3 is a downstream component of WNT7B signaling, which transcriptionally modulates NOTCH1, a tumor suppressor in bladder urothelial carcinoma. INTERPRETATION: These data demonstrate that WNT7B/FZD5-ELF3-NOTCH1 signaling functions as a tumor-suppressing pathway in bladder urothelial carcinoma.

Higher circular RNA_0015278 correlates with absence of extrathyroidal invasion, lower pathological tumor stages, and prolonged disease-free survival in papillary thyroid carcinoma patients.

BACKGROUND: Circular RNA_0015278 (circ_0015278) inhibits the progression of several cancers and is greatly reduced in papillary thyroid carcinoma (PTC) tissues compared with benign thyroid lesions by microarray profiling. This study aimed to further investigate the correlation of circ_0015278 with tumor characteristics and prognosis in PTC patients. METHODS: Totally, 206 PTC patients who underwent tumor resection were retrospectively enrolled; subsequently, circ_0015278 expression in their tumor and adjacent tissues was detected by reverse transcriptional-quantitative polymerase chain reaction. Besides, disease-free survival (DFS) and overall survival (OS) were calculated. RESULTS: Circ_0015278 was reduced in tumor tissues compared with adjacent tissues (p < 0.001), and receiver operating characteristic analysis showed that it well discriminated tumor tissues from adjacent tissues (area under curve: 0.903, 95% confidence interval: 0.874-0.932). Besides, higher tumor circ_0015278 expression was correlated with absence of extrathyroidal invasion (p = 0.036), lower pathological tumor (pT) stage (p = 0.05), pathological node (pN) stage (p = 0.002), and pathological tumor-node-metastasis (pTNM) stage (p = 0.001). Moreover, higher tumor circ_0015278 expression was associated with a reduced relapse rate (p = 0.011), but not mortality rate (p = 0.110); meanwhile, it was also correlated with prolonged DFS (p = 0.017), but not OS (p = 0.136). Additionally, multivariate Cox's regression analyses showed that higher tumor circ_0015278 expression independently associated with favorable DFS (p = 0.026, hazard ratio = 0.529). CONCLUSION: Circ_0015278 is reduced in tumor tissues, while its' higher expression in tumor correlates with absence of extrathyroidal invasion, lower pT, pN, and pTNM stage, as well as prolonged DFS in PTC patients.

FZD5 prevents epithelial-mesenchymal transition in gastric cancer.

BACKGROUND: Frizzled (FZD) proteins function as receptors for WNT ligands. Members in FZD family including FZD2, FZD4, FZD7, FZD8 and FZD10 have been demonstrated to mediate cancer cell epithelial-mesenchymal transition (EMT). METHODS: CCLE and TCGA databases were interrogated to reveal the association of FZD5 with EMT. EMT was analyzed by investigating the alterations in CDH1 (E-cadherin), VIM (Vimentin) and ZEB1 expression, cell migration and cell morphology. Transcriptional modulation was determined by ChIP in combination with Real-time PCR. Survival was analyzed by Kaplan-Meier method. RESULTS: In contrast to other FZDs, FZD5 was identified to prevent EMT in gastric cancer. FZD5 maintains epithelial-like phenotype and is negatively modulated by transcription factors SNAI2 and TEAD1. Epithelial-specific factor ELF3 is a downstream effecter, and protein kinase C (PKC) links FZD5 to ELF3. ELF3 represses ZEB1 expression, further guarding against EMT. Moreover, FZD5 signaling requires its co-receptor LRP5 and WNT7B is a putative ligand for FZD5. FZD5 and ELF3 are associated with longer survival, whereas SNAI2 and TEAD1 are associated with shorter survival. CONCLUSIONS: Taken together, FZD5-ELF3 signaling blocks EMT, and plays a potential tumor-suppressing role in gastric cancer. Video Abstract.

FZD5 contributes to TNBC proliferation, DNA damage repair and stemness.

Chemotherapy currently remains the standard treatment for triple-negative breast cancer (TNBC). However, TNBC frequently develop chemoresistance, which is responsible for cancer recurrence and distal metastasis. Both DNA damage repair and stemness are related to chemoresistance. FZD5, a member in Frizzled family, was identified to be preferentially expressed in TNBC, and associated with unfavorable prognosis. Loss and gain of function studies revealed that FZD5 contributed to TNBC cell G1/S transition, DNA replication, DNA damage repair, survival, and stemness. Mechanistically, transcription factor FOXM1, which promoted BRCA1 and BIRC5 transcription, acted as a downstream effecter of FZD5 signaling. FOXM1 overexpression in FZD5-deficient/low TNBC cells induced FZD5-associated phenotype. Finally, Wnt7B, a specific ligand for FZD5, was shown to be involved in cell proliferation, DNA damage repair, and stemness. Taken together, FZD5 is a novel target for the development of therapeutic strategies to overcome chemoresistance and prevent recurrence in TNBC.

LGR4 maintains HGSOC cell epithelial phenotype and stem-like traits.

OBJECTIVE: High-grade serous ovarian cancer (HGSOC) is lethal mainly due to extensive metastasis. Cancer cell stem-like properties are responsible for HGSOC metastasis. LGR4, a G-protein-coupled receptor, is involved in the maintenance of stem cell self-renewal and activity in some human organs. METHODS: TCGA and CCLE databases were interrogated for gene mRNA in ovarian cancer tissues and cell lines. Gain and loss of functions of LGR4, ELF3, FZD5 and WNT7B were performed to identify their roles in ovarian cancer cell epithelial phenotype and stem-like properties. In vivo experiments were performed to observe the effect of LGR4 on ovarian cancer cell growth and peritoneal seeding. The binding of ELF3 to LGR4 gene promoter was investigated by dual-luciferase reporter assays and ChIP. RESULTS: LGR4 was shown to be overexpressed in HGSOCs and maintain the epithelial phenotype of HGSOC cells. LGR4 knockdown suppressed POU5F1, SOX2, PROM1 (CD133) and ALDH1A2 expression. Furthermore, LGR4 knockdown reduced CD133+ and ALDH+ subpopulations and impaired tumorisphere formation. To the contrary, LGR4 overexpression enhanced POU5F1 and SOX2 expression and tumorisphere formation capacity. LGR4 knockdown inhibited HGSOC cell growth and peritoneal seeding in xenograft models. Mechanistically, LGR4 and ELF3, an epithelium-specific transcription factor, formed a reciprocal regulatory loop, which was positively modulated by WNT7B/FZD5 ligand-receptor pair. Consistently, knockdown of ELF3, WNT7B, and FZD5, respectively, disrupted HGSOC cell epithelial phenotype and stem-like properties. CONCLUSION: Together, these data demonstrate that WNT7B/FZD5-LGR4/ELF3 axis maintains HGSOC cell epithelial phenotype and stem-like traits; targeting this axis may prevent HGSOC metastasis.

Non-canonical Fzd7 signaling contributes to breast cancer mesenchymal-like stemness involving Col6a1.

Mesenchymal-like stemness is characterized by epithelial-mesenchymal transition (EMT). Breast cancer (BC) cell mesenchymal-like stemness is responsible for distal lung metastasis. Interrogation of databases showed that Fzd7 was closely associated with a panel of mesenchymal-related genes and a panel of stemness-related genes. Fzd7 knockdown in mesenchymal-like MDA-MB-231 and Hs578T cells reduced expression of Vimentin, Slug and Zeb1, induced an epithelial-like morphology, inhibited cell motility, impaired mammosphere formation and decreased Lgr5+ subpopulation. In contrast, Fzd7 overexpression in MCF7 cells resulted in opposite changes. Fzd7 knockdown delayed xenograft tumor formation, suppressed tumor growth, and impaired lung metastasis. Mechanistically, Fzd7 combined with Wnt5a/b and modulated expression of phosphorylated Stat3 (p-STAT3), Smad3 and Yes-associated protein 1 (Yap1). Moreover, Fzd7-Wnt5b modulated expression of collagen, type VI, alpha 1 (Col6a1). Both Wnt5b knockdown and Col6a1 knockdown disrupted BC cell mesenchymal phenotype and stemness. Taken together, Fzd7 contributes to BC cell EMT and stemness, inducing tumorigenesis and metastasis, mainly through a non-canonical Wnt5b pathway. Col6a1 is implicated in Fzd7-Wnt5b signaling, and mediates Fzd7-Wnt5b -induced mesenchymal-like stemness. Video Abstract.

Identification of 9-Core Immune-Related Genes in Bladder Urothelial Carcinoma Prognosis.

Background: Immune microenvironment within tumors affects initiation, progression and clinical outcome of human cancers. Here we explored an immune-related gene signature associated with prognosis of patients with bladder urothelial carcinoma. Method: The Cancer Genome Atlas (TCGA) database was interrogated for expressions of immune-related genes in bladder urothelial carcinomas. Integrated bioinformatics analyses were performed to identify prognostic factors. Results: Twenty-seven immune-related genes were revealed significantly associated with patient's overall survival (OS) by univariate Cox proportional hazards regression analysis. Nine-core immune-related genes including MMP9, PDGFRA, AHNAK, OLR1, RAC3, IGF1, PGF, OAS1, and SH3BP2 were selected to construct a risk score model by multivariate Cox proportional hazards regression analysis. Bioinformatics analyses further validated that risk score could be used as an important independent factor in evaluating prognosis. Conclusion: We established a prognostic immune signature for patients with bladder urothelial carcinoma, which may provide novel targets for prediction and therapy of these patients.

Env diversity-dependent protection of the attenuated equine infectious anaemia virus vaccine.

Lentiviruses harbour high genetic variability for efficient evasion from host immunity. An attenuated equine infectious anaemia (EIA) vaccine was developed decades ago in China and presented remarkably robust protection against EIA. The vaccine was recently proven to have high genomic diversity, particular in env. However, how and to what extent the high env diversity relates to immune protection remains unclear. In this study, we compared immune protections and responses of three groups of horses stimulated by the high-diversity vaccine EIAV_HD, a single molecular clone of the vaccine EIAV_LD with low env diversity, as well as a constructed vaccine strain EIAV_MD with moderate env diversity. The disparity of virus-host interactions between three env diversity-varied groups (5 horses in each group) was evaluated using clinical manifestation, pathological scores, and env-specific antibody. We found the highest titres of env antibodies (Abs) or neutralizing Abs (nAbs) in the EIAV_HD group, followed by the EIAV_MD group, and the lowest titres in the EIAV_LD group (P<0.05). The occurrence of disease/death was different between EIAV_HD group (1/0), EIAV_MD (2/2), and EIAV_LD group (4/2). A similar env diversity-related linear relationship was observed in the clinical manifestations and pathological changes. This diversity-dependent disparity in changes between the three groups was more distinct after immunosuppression, suggesting that env diversity plays an important role in protection under low host immunocompetence. In summary, inoculation with vaccines with higher genetic diversity could present broader and more efficient protection. Our findings strongly suggest that an abundance of Env antigens are required for efficient protection against lentiviruses.

The N-glycosylation of Equine Tetherin Affects Antiviral Activity by Regulating Its Subcellular Localization.

Tetherin is an interferon-inducible type II transmembrane glycoprotein which inhibits the release of viruses, including retroviruses, through a "physical tethering" model. However, the role that the glycosylation of tetherin plays in its antiviral activity remains controversial. In this study, we found that mutation of N-glycosylation sites resulted in an attenuation of the antiviral activity of equine tetherin (eqTHN), as well as a reduction in the expression of eqTHN at the plasma membrane (PM). In addition, eqTHN N-glycosylation mutants colocalize obviously with ER, CD63, LAMP1 and endosomes, while WT eqTHN do not. Furthermore, we also found that N-glycosylation impacts the transport of eqTHN in the cell not by affecting the endocytosis, but rather by influencing the anterograde trafficking of the protein. These results suggest that the N-glycosylation of eqTHN is important for the antiviral activity of the protein through regulating its normal subcellular localization. This finding will enhance our understanding of the function of this important restriction factor.

Fzd2 Contributes to Breast Cancer Cell Mesenchymal-Like Stemness and Drug Resistance.

Cancer cell stemness is responsible for cancer relapse, distal metastasis, and drug resistance. Here we identified that Frizzled 2 (Fzd2), one member of Wnt receptor Frizzled family, induced human breast cancer (BC) cell stemness via noncanonical Wnt pathways. Fzd2 was overexpressed in human BC tissues, and Fzd2 overexpression was associated with an unfavorable outcome. Fzd2 knockdown (KD) disturbed the mesenchymal-like phenotype, migration, and invasion of BC cells. Moreover, Fzd2 KD impaired BC cell mammosphere formation, reduced Lgr5+ BC cell subpopulation, and enhanced sensitivity of BC cells to chemical agents. Mechanistically, Fzd2 modulated and bound with Wnt5a/b and Wnt3 to activate several oncogenic pathways such as interleukin-6 (IL-6)/Stat3, Yes-associated protein 1 (Yap1), and transforming growth factor-ß1 (TGF-ß1)/Smad3. These data indicate that Fzd2 contributes to BC cell mesenchymal-like stemness; targeting Fzd2 may inhibit BC recurrence, metastasis, and chemoresistance.

Lnc-MEG3 acts as a potential biomarker for predicting increased disease risk, systemic inflammation, disease severity, and poor prognosis of sepsis via interacting with miR-21.

BACKGROUND: This study aimed to investigate the correlations of long non-coding RNA maternally expressed gene 3 (lnc-MEG3), microRNA (miR)-21, and lnc-MEG3/miR-21 axis with disease risk, inflammation, disease severity, and 28-day mortality of sepsis. METHODS: Totally, 219 sepsis patients and 219 health controls (HCs) were enrolled. Plasma samples were obtained from sepsis patients within 24 hours after admission and from HCs on enrollment to detect lnc-MEG3 and miR-21 expressions by real-time quantitative polymerase chain reaction. RESULTS: The lnc-MEG3 expression and lnc-MEG3/miR-21 axis were increased, while miR-21 expression was decreased in sepsis patients compared with HCs. Lnc-MEG3 (area under the curve (AUC): 0.887, 95% confidence interval (CI): 0.856-0.917) and lnc-MEG3/miR-21 axis (AUC: 0.934, 95% CI: 0.909-0.958) had good values for predicting elevated sepsis risk, while miR-21 (AUC: 0.801, 95% CI: 0.758-0.844) presented a good predictive value for reduced sepsis risk. Furthermore, lnc-MEG3 expression and lnc-MEG3/miR-21 axis positively correlated with, whereas miR-21 expression negatively correlated with acute pathologic and chronic health evaluation II, sequential organ failure assessment score, serum creatinine, C-reactive protein, tumor necrosis factor-α, interleukin (IL)-1ß, IL-6, and IL-17 in sepsis patients. Additionally, lnc-MEG3 (AUC: 0.704, 95% CI: 0.626-0.783) and lnc-MEG3/miR-21 axis (AUC: 0.669, 95% CI: 0.589-0.750) exhibited acceptable values in predicting higher 28-day mortality risk, while miR-21 (AUC: 0.588, 95% CI: 0.505-0.672) presented a poor predictive value for lower 28-day mortality risk in sepsis patients. CONCLUSION: Lnc-MEG3 might serve as a potential biomarker for the development, progression, and prognosis prediction of sepsis via interacting with miR-21.

Ruxolitinib Alleviates Renal Interstitial Fibrosis in UUO Mice.

Ruxolitinib is a selective inhibitor of Jak1/2. Downstream signaling pathways of Jak, such as Stat3 and Akt/mTOR, are overactivated and contribute to renal interstitial fibrosis. Therefore, we explored the effect of Ruxolitinib on this pathological process. Unilateral ureteral obstruction (UUO) models and TGF-ß1-treated fibroblasts and renal tubular epithelial cells were adopted in this study. Ruxolitinib was administered to UUO mice and TGF-ß1-treated cells. Kidneys from UUO mice with Ruxolitinib treatment displayed less tubular injuries compared with those without Ruxolitinib treatment. Ruxolitinib treatment suppressed fibroblast activation and extracellular matrix (ECM) production in UUO kidneys and TGF-ß1-treated fibroblasts. Ruxolitinib treatment also blocked epithelial-mesenchymal transition (EMT) in UUO kidneys and TGF-ß 1-treated renal tubular epithelial cells. Moreover, Ruxolitinib treatment alleviated UUO-induced inflammation, oxidative stress and apoptosis. Mechanistically, Ruxolitinib treatment attenuated activation of both Stat3 and Akt/mTOR/Yap pathways. In conclusion, Ruxolitinib treatment can ameliorate UUO-induced renal interstitial fibrosis, suggesting that Ruxolitinib may be potentially used to treat fibrotic kidney disease.

The predictive value of microRNA-21 for sepsis risk and its correlation with disease severity, systemic inflammation, and 28-day mortality in sepsis patients.

BACKGROUND: This study aimed to investigate the value of microRNA (miR)-21 for predicting sepsis risk and its correlation with inflammation, disease severity as well as 28-day mortality in sepsis patients. METHODS: Totally, 219 sepsis patients and 219 healthy controls (HCs) were recruited. Plasma samples were obtained from sepsis patients within 24 hours after admission and from HCs at the enrollment to detect miR-21 expressions by real-time quantitative polymerase chain reaction. Besides, the clinical characteristics of sepsis patients were recorded and the 28-day mortality of sepsis patients was evaluated. RESULTS: MiR-21 expression was decreased in sepsis patients compared with HCs, and further receiver operating characteristic (ROC) curve analysis revealed that miR-21 was of a good value in predicting sepsis risk (area under the curve [AUC]: 0.801, 95% CI: 0.758-0.844). Besides, miR-21 expression was negatively associated with acute pathologic and chronic health evaluation II (APACHE II) and sequential organ failure assessment (SOFA) score in sepsis patients. Furthermore, miR-21 expression was negatively correlated with serum creatinine, C-reactive protein, tumor necrosis factor-α, interleukin (IL)-1ß, IL-6, and IL-17, while positively correlated with albumin in sepsis patients. However, there was no correlation of miR-21 expression with white blood cell, smoke, or comorbidities in sepsis patients. Additionally, ROC curve analysis displayed that miR-21 exhibited a poor predictive value for 28-day mortality risk in sepsis patients (AUC: 0.588, 95% CI: 0.505-0.672). CONCLUSION: MiR-21 might serve as a potential biomarker for the development and progression of sepsis, while not for prognosis prediction in sepsis patients.

Attenuation of Equine Lentivirus Alters Mitochondrial Protein Expression Profile from Inflammation to Apoptosis.

Equine infectious anemia virus (EIAV) is an equine lentivirus similar to HIV-1, targets host immune cells, and causes a life-long infection in horses. The Chinese live EIAV vaccine is attenuated from long-term passaging of a highly virulent strain in vitro The parent pathogenic strain (EIAVDLV34) induces a host inflammatory storm to cause severe pathological injury of animals. However, the vaccine strain (EIAVDLV121) induces a high level of apoptosis to eliminate infected cells. To investigate how these processes are regulated, we performed a comparative proteomics analysis and functional study in equine monocyte-derived macrophages (eMDMs) and found that the divergent mitochondrial protein expression profiles caused by EIAV strains with different virulence led to disparate mitochondrial function, morphology, and metabolism. This in turn promoted the distinct transformation of macrophage inflammatory polarization and intrinsic apoptosis. In EIAVDLV34-infected cells, a high level of glycolysis and increased mitochondrial fragmentation were induced, resulting in the M1-polarized proinflammatory-type transformation of macrophages and the subsequent production of a strong inflammatory response. Following infection with EIAVDLV121, the infected cells were transformed into M2-polarized anti-inflammatory macrophages by inhibition of glycolysis. In this case, a decrease in the mitochondrial membrane potential and impairment of the electron transport chain led to increased levels of apoptosis and reactive oxygen species. These results correlated with viral pathogenicity loss and may help provide an understanding of the key mechanism of lentiviral attenuation.IMPORTANCE Following viral infection, the working pattern and function of the cell can be transformed through the impact on mitochondria. It still unknown how the mitochondrial response changes in cells infected with viruses in the process of virulence attenuation. EIAVDLV121 is the only effective lentiviral vaccine for large-scale use in the world. EIAVDLV34 is the parent pathogenic strain. Unlike EIAVDLV34-induced inflammation storms, EIAVDLV121 can induce high levels of apoptosis. For the first time, we found that, after the mitochondrial protein expression profile is altered, EIAVDLV34-infected cells are transformed into M1-polarized-type macrophages and cause inflammatory injury and that the intrinsic apoptosis pathway is activated in EIAVDLV121-infected cells. These studies shed light on how the mitochondrial protein expression profile changes between cells infected by pathogenic lentivirus strains and cells infected by attenuated lentivirus strains to drive different cellular responses, especially from inflammation to apoptosis.

Equine Myxovirus Resistance Protein 2 Restricts Lentiviral Replication by Blocking Nuclear Uptake of Capsid Protein.

Human myxovirus resistance protein 2 (huMxB) has been shown to be a determinant type I interferon (IFN)-induced host factor involved in the inhibition of human immunodeficiency virus type 1 (HIV-1) as well as many other primate lentiviruses. This blocking occurs after the reverse transcription of viral RNA and ahead of integration into the host DNA, which is closely connected to the ability of the protein to bind the viral capsid. To date, Mx2s derived from nonprimate animals have shown no capacity for HIV-1 suppression. In this study, we examined the restrictive effect of equine Mx2 (eqMx2) on both equine infectious anemia virus (EIAV) and HIV-1 and investigated possible mechanisms for its specific function. We demonstrated that IFN-α/ß upregulates the expression of eqMx2 in equine monocyte-derived macrophages (eMDMs). The overexpression of eqMx2 significantly suppresses the replication of EIAV, HIV-1, and simian immunodeficiency viruses (SIVs) but not that of murine leukemia virus (MLV). The knockdown of eqMx2 transcription weakens the inhibition of EIAV replication by type I interferon. Interestingly, data from immunofluorescence assays suggest that the subcellular localization of eqMx2 changes following virus infection, from being dispersed in the cytoplasm to being accumulated at the nuclear envelope. Furthermore, eqMx2 blocks the nuclear uptake of the proviral genome by binding to the viral capsid. The N-terminally truncated mutant of eqMx2 lost the ability to bind the viral capsid as well as the restriction effect for lentiviruses. These results improve our understanding of the Mx2 protein in nonprimate animals.IMPORTANCE Previous research has shown that the antiviral ability of Mx2s is confined to primates, particularly humans. EIAV has been shown to be insensitive to restriction by human MxB. Here, we describe the function of equine Mx2. This protein plays an important role in the suppression of EIAV, HIV-1, and SIVs. The antiviral activity of eqMx2 depends on its subcellular location as well as its capsid binding capacity. Our results showed that following viral infection, eqMx2 changes its original cytoplasmic location and accumulates at the nuclear envelope, where it binds to the viral capsid and blocks the nuclear entry of reverse-transcribed proviral DNAs. In contrast, huMxB does not bind to the EIAV capsid and shows no EIAV restriction effect. These studies expand our understanding of the function of the equine Mx2 protein.

Double-stranded-RNA-specific adenosine deaminase 1 (ADAR1) is proposed to contribute to the adaptation of equine infectious anemia virus from horses to donkeys.

Equine infectious anemia virus (EIAV) is a member of the genus Lentivirus of the family Retroviridae. Horses are the most susceptible equids to EIAV infection and are therefore the primary hosts of this virus. In contrast, infected donkeys do not develop clinically active equine infectious anemia (EIA). This phenomenon is similar to what has been observed with HIV-1, which fails to induce AIDS in non-human primates. Interestingly, Shen et al. developed a donkey-tropic pathogenic virus strain (EIAVDV117, DV117) by serially passaging a horse-tropic pathogenic strain, EIAVLN40 (LN40), in donkeys. LN40, which was generated by passaging a field isolate in horses, displayed enhanced virulence in horses but caused no clinical symptoms in donkeys. Infection with DV117 induced acute EIA in nearly 100 % of donkeys. Genomic analysis of DV117 revealed a significantly higher frequency of A-to-G substitutions when compared to LN40. Furthermore, detailed analysis of dinucleotide editing showed that A-to-G mutations had a preference for 5'TpA and 5'ApA. These results strongly implicated the activity of the adenosine deaminase, ADAR1, in this type of mutation. Further investigation demonstrated that overexpression of donkey ADAR1 increased A-to-G mutations within the genome of EIAV. Together with our previous finding that multiple mutations in multiple genes are generated in DV117 during its adaptation from horses to donkeys, the present study suggests that ADAR1-induced A-to-G mutations occur during virus adaption to related new hosts contributing to the alteration of EIAV host tropism.

Double-stranded RNA-specific adenosine deaminase 1 (ADAR1) promotes EIAV replication and infectivity.

Adenosine deaminases that act on RNA (ADARs) have been reported to be functional on various viruses. ADAR1 may exhibit antiviral or proviral activity depending on the type of virus. Human immunodeficiency virus (HIV)-1 is the most well-studied lentivirus with respect to its interaction with ADAR1, and variable results have been reported. In this study, we demonstrated that equine ADAR1 (eADAR1) was a positive regulator of equine infectious anemia virus (EIAV), another lentivirus of the Retroviridae family. First, eADAR1 significantly promoted EIAV replication, and the enhancement of viral protein expression was associated with the long terminal repeat (LTR) and Rev response element (RRE) regions. Second, the RNA binding domain 1 of eADAR1 was essential only for enhancing LTR-mediated gene expression. Third, in contrast with APOBEC proteins, which have been shown to reduce lentiviral infectivity, eADAR1 increased the EIAV infectivity. This study indicated that eADAR1 was proviral rather than antiviral for EIAV.