Mechanisms of intestinal epithelial cell damage by Clostridiumperfringens.

Clostridium perfringens, a Gram-positive bacterium, causes intestinal diseases in humans and livestock through its toxins, related to alpha toxin (CPA), beta toxin (CPB), C. perfringens enterotoxin (CPE), epsilon toxin (ETX), Iota toxin (ITX), and necrotic enteritis B-like toxin (NetB). These toxins disrupt intestinal barrier, leading to various cell death mechanisms such as necrosis, apoptosis, and necroptosis. Additionally, non-toxin factors like adhesins and degradative enzymes contribute to virulence by enhancing colonization and survival of C. perfringens. A vicious cycle of intestinal barrier breach, misregulated cell death, and subsequent inflammation is at the heart of chronic inflammatory and infectious gastrointestinal diseases. Understanding these mechanisms is essential for developing targeted therapies against C. perfringens-associated intestinal diseases.

Ubiquitin E3 ligase SPOP is a host negative regulator of enterovirus 71-encoded 2A protease.

IMPORTANCE: EV71 poses a significant health threat to children aged 5 and below. The process of EV71 infection and replication is predominantly influenced by ubiquitination modifications. Our previous findings indicate that EV71 prompts the activation of host deubiquitinating enzymes, thereby impeding the host interferon signaling pathway as a means of evading the immune response. Nevertheless, the precise mechanisms by which the host employs ubiquitination modifications to hinder EV71 infection remain unclear. The present study demonstrated that the nonstructural protein 2Apro, which is encoded by EV71, exhibits ubiquitination and degradation mediated by the host E3 ubiquitin ligase SPOP. In addition, it is the first report, to our knowledge, that SPOP is involved in the host antiviral response.

Ceftazidime reduces cellular Skp2 to promote type-I interferon activity.

Skp2 plays multiple roles in malignant tumours. Here, we revealed that Skp2 negatively regulates type-I interferon (IFN-I)-mediated antiviral activity. We first noticed that Skp2 can promote virus infection in cells. Further studies demonstrated that Skp2 interacts with IFN-I receptor 2 (IFNAR2) and promotes K48-linked polyubiquitination of IFNAR2, which accelerates the degradation of IFNAR2 proteins. Skp2-mediated downregulation of IFNAR2 levels inhibits IFN-I signalling and IFN-I-induced antiviral activity. In addition, we uncovered for the first time that the antibiotic ceftazidime can act as a repressor of Skp2. Ceftazidime reduces cellular Skp2 levels, thus enhancing IFNAR2 stability and IFN-I antiviral activity. This study reveals a new role of Skp2 in regulating IFN-I signalling and IFN-I antiviral activity and reports the antibiotic ceftazidime as a potential repressor of Skp2.

Methotrexate and Triptolide regulate Notch signaling pathway by targeting the Nedd4-Numb axis.

Methotrexate (MTX) is used to treat rheumatoid arthritis, acute leukemia, and psoriasis. MTX can cause certain side effects, such as myelosuppression, while the exact mechanism of myelosuppression caused by MTX is unknown. Notch signaling pathway has been considered to be essential to regulate hematopoietic stem cell (HSC) regeneration and homeostasis, thus contributing to bone marrow hematopoiesis. However, whether MTX affects Notch signaling remains unexplored. Here, our study provides evidence that MTX strongly suppresses the Notch signaling pathway. We found that MTX inhibited the interaction between Nedd4 with Numb, thus restricting K48-linked polyubiquitination of Numb and stabilizing Numb proteins. This in turn inhibited the Notch signaling pathway by reducing Notch1 protein levels. Interestingly, we found that a monomeric drug, Triptolide, is capable of alleviating the inhibitory effect of MTX on Notch signaling pathway. This study promotes our understanding of MTX-mediated regulation of Notch signaling and could provide ideas to alleviate MTX-induced myelosuppression.

Targeting PARP11 to avert immunosuppression and improve CAR T therapy in solid tumors.

Evasion of antitumor immunity and resistance to therapies in solid tumors are aided by an immunosuppressive tumor microenvironment (TME). We found that TME factors, such as regulatory T cells and adenosine, downregulated type I interferon receptor IFNAR1 on CD8+ cytotoxic T lymphocytes (CTLs). These events relied upon poly-ADP ribose polymerase-11 (PARP11), which was induced in intratumoral CTLs and acted as a key regulator of the immunosuppressive TME. Ablation of PARP11 prevented loss of IFNAR1, increased CTL tumoricidal activity and inhibited tumor growth in an IFNAR1-dependent manner. Accordingly, genetic or pharmacologic inactivation of PARP11 augmented the therapeutic benefits of chimeric antigen receptor T cells. Chimeric antigen receptor CTLs engineered to inactivate PARP11 demonstrated a superior efficacy against solid tumors. These findings highlight the role of PARP11 in the immunosuppressive TME and provide a proof of principle for targeting this pathway to optimize immune therapies.

Selective elimination of CML stem/progenitor cells by picropodophyllin in vitro and in vivo is associated with p53 activation.

Chronic myeloid leukemia (CML) is a hematologic malignancy originating from BCR-ABL oncogene-transformed hematopoietic stem cells (HSCs) known as leukemia stem cells (LSCs). Therefore, targeting LSCs is of primary importance to eradicate CML. The present study demonstrates that picropodophyllin (PPP) effectively induces apoptosis and inhibits colony formation in CML stem/progenitor cells as well as quiescent CML progenitors resistant to imatinib therapy, while sparing normal hematopoietic cells in vitro. Administration of PPP in vivo markedly diminishes CML stem/progenitor cells in a transgenic mouse model of CML by inhibition of cell proliferation and enhancement of apoptosis in LSK cells, and significantly improves survival of CML mice. Furthermore, PPP treatment preferentially leads to transcriptional activation of p53 in CML but not normal CD34+ cells, upregulation of p53 protein in LSCs-enriched Sca-1+ cells from CML mice, and increased phosphorylation of p53 and upregulation of Bax protein in Ku812 cells. These results suggest that the inhibitory effects of PPP on CML stem/progenitor cells are associated with selective activation of p53 pathway and propose that PPP is a potent agent that selectively targets CML LSCs, and may be of value in the CML therapy.

HSV-1-encoded ICP0 degrades the host deubiquitinase BRCC36 to antagonize interferon antiviral response.

Type I interferon (IFN-I) plays pivotal roles in defense against viral infection. HSV-1 has evolved multiple strategies to evade IFN-I antiviral response. In this study, we revealed a new mechanism that HSV-1-encoded ICP0 regulates the host deubiquitinase BRCC36 to inhibit IFN-I antiviral response. We found that HSV-1 infection rapidly downregulates BRCC36 proteins at the early stage of infection. Further studies demonstrated that HSV-1-encoded ICP0 induces K48-linked polyubiquitination and degradation of BRCC36. Importantly, HSV-1-induced BRCC36 degradation promotes downmodulation of IFN-I receptor IFNAR1, thus restricting host IFN-I antiviral response to facilitate HSV-1 early infection. These findings uncover a novel immune evasion mechanism exploited by HSV-1 and could provide potential strategies for anti-HSV-1 therapy.

[Effect of Etoposide on Elimination of Chronic Myeloid Leukemia Stem Cells by Imatinib in Vivo].

OBJECTIVE: To investigate the effect of etoposide (ETO) on elimination of chronic myeloid leukemia (CML) stem cells by imatinib mesylate(IM) in vivo. METHODS: SCL-tTA/BCR-ABL mice were used as CML animal model. Flow cytometry was used to assess the effect of ETO alone or in combination with IM on the number of leukemia stem cell （LSC） in bone marrow and spleen, and peripheral blood neutrophils in CML mice and normal control FVB mice. RESULTS: The results showed that in CML mice, the number and proportion of LSC in bone marrow and the proportion of neutrophils in peripheral blood decreased significantly after ETO and IM combined treatment, and the degree of decrease was more significant than that of both alone. While in wild type FVB mice, the combination of ETO and IM showed no significant effect on the number and proportion of LSK cells in bone marrow and the proportion of neutrophils in spleen. CONCLUSION: ETO can selectively enhance elimination of CML LSC by IM in vivo.

Targeting UBE4A Revives Viperin Protein in Epithelium to Enhance Host Antiviral Defense.

Mutation and prevalence of pathogenic viruses prompt the development of broad-spectrum antiviral strategies. Viperin is a potent antiviral protein that inhibits a broad range of viruses. Unexpectedly, we found that Viperin protein production in epithelium is defective in response to both viruses and interferons (IFNs). We further revealed that viruses and IFNs stimulate expression of the acetyltransferase HAT1, which induces Lys197-acetylation on Viperin. Viperin acetylation in turn recruits UBE4A that stimulates K6-linked polyubiquitination at Lys206 of Viperin, leading to Viperin protein degradation. Importantly, UBE4A deficiency restores Viperin protein production in epithelium. We then designed interfering peptides (IPs) to inhibit UBE4A binding with Viperin. We found that VIP-IP3 rescues Viperin protein production in epithelium and therefore enhances cellular antiviral activity. VIP-IP3 renders mice more resistant to viral infection. These findings could provide strategies for both enhancing host broad-spectrum antiviral response and improving the efficacy of IFN-based antiviral therapy.

ADP-ribosyltransferase PARP11 modulates the interferon antiviral response by mono-ADP-ribosylating the ubiquitin E3 ligase ß-TrCP.

Outbreaks of viral infections are a global health burden. Although type I interferon (IFN-I) exerts broad-spectrum antiviral effects, its antiviral efficacy in host cells is largely restricted by viruses. How the antiviral efficacy of IFN-I can be improved remains to be explored. Here, we identified the ADP-ribosyltransferase poly(ADP-ribose) polymerase family member 11 (PARP11) as a potent regulator of IFN-I antiviral efficacy. PARP11 does not restrict IFN-I production induced by vesicular stomatitis virus or Sendai virus but inhibits the strength of IFN-I-activated signalling. Mechanistically, PARP11 mono-ADP-ribosylates the ubiquitin E3 ligase ß-transducin repeat-containing protein (ß-TrCP). Mono-ADP-ribosylation of ß-TrCP promotes IFNα/ß receptor subunit 1 (IFNAR1) ubiquitination and degradation. Moreover, PARP11 expression is upregulated by virus infections, including vesicular stomatitis virus, herpes simplex virus-1 and influenza A virus, thus promoting ADP-ribosylation-mediated viral evasion. We further highlight the potential for repurposing clinical ADP-ribosylation inhibitors. We found that rucaparib can target PARP11 to stabilize IFNAR1 and therefore exhibits efficient enhancement of IFN-I signalling and the host antiviral response. Consequently, rucaparib renders mice more resistant to viral infection. Our study updates the understanding of how ß-TrCP regulates its substrates and may provide a druggable target for improving IFN antiviral efficacy.

Expression of transmembrane protein 41A is associated with metastasis via the modulation of E­cadherin in radically resected gastric cancer.

Gastric cancer (GC) is one of the most commonly occurring malignancies worldwide, and metastasis is one of the key processes affecting the prognosis of GC. TMEM41A, which belongs to a group of transmembrane proteins that participate in signaling pathways and tumor development, is a 264­amino acid protein encoded by a gene mapped to human chromosome The exact role of TMEM41A in GC has not been determined to date. In the present study, the expression of TMEM41A in 147 cases of GC was analyzed with immunohistohemistry and the prognoses of these patients were analyzed. It was revealed that TMEM41A was highly expressed in GC tissues, and may be associated with the progression of GC and poor prognosis. The expression of TMEM41A was observed to be correlated with lymph node metastasis, distant metastasis and advanced tumor, node and metastasis stages. Knockdown of TMEM41A in vitro and in vivo decreased the GC cell migration ability by regulating epithelial­to­mesenchymal transition and cell autophagy, via the upregulation of E­cadherin and downregulating N­cadherin expression in GC cells by reverse transcription­quantitative polymerase chain reaction (PCR), semi­PCR and western blotting. Furthermore, TMEM41A upregulation was associated with the upregulation of p62 and altered the conversion of light chain (LC)3­1 into LC3­2 by western blotting. Knockdown of TMEM41A was also observed to affect tumor metastasis in nude mice. Therefore, TMEM41A may be considered as a novel therapeutic target for the treatment of GC­associated metastasis.

Gold nanoparticle-based miR155 antagonist macrophage delivery restores the cardiac function in ovariectomized diabetic mouse model.

Diabetic cardiomyopathy is a common disease in postmenopausal women, in whom the estrogen deficiency aggravates the pathology. In this study, we have found that estrogen deficiency due to ovariectomy aggravates the inflammation in the hearts of diabetic mice, as depicted by excessive proinflammatory type 1 macrophages (M1) over anti-inflammatory type 2 macrophages (M2). Accordingly, an additional increase of reactive oxygen species, cell apoptosis, cardiac hypertrophy, and fibrosis was observed in the hearts of ovariectomized diabetic mice, in comparison with the diabetes-only group. Significantly, miR155, a potent promoter of M1 polarization, was found to be additionally enhanced in the macrophages and hearts by ovariectomy. Tail vein injection of miR155-AuNP, in which thiol-modified antago-miR155 was covalently conjugated with gold nanoparticle (AuNP), preferentially delivered the nucleic acids into the macrophages via phagocytosis. Together with the increased M2 ratio and reduced inflammation, in vivo delivery of antago-miR155 reduced cell apoptosis and restored the cardiac function. The restoration efficacy of miR155-AuNP was much better than general macrophage depletion by clodrosome. In summary, we revealed that M1/M2 imbalance contributes to the aggravated cardiomyopathy in ovariectomized diabetic mice, and therapeutically reducing miR155 in macrophages by AuNP serves as a promising strategy in improving cardiac function.