Immune-checkpoint proteins, cytokines, and microbiome impact on patients with cervical insufficiency and preterm birth

BackgroundMicroenvironmental factors, including microbe-induced inflammation and immune-checkpoint proteins that modulate immune cells have been associated with both cervical insufficiency and preterm delivery. These factors are incompletely understood. This study aimed to explore and compare interactions among microbiome and inflammatory factors, such as cytokines and immune-checkpoint proteins, in patients with cervical insufficiency and preterm birth. In particular, factors related to predicting preterm birth were identified and the performance of the combination of these factors was evaluated.MethodsA total of 220 swab samples from 110 pregnant women, prospectively recruited at the High-Risk Maternal Neonatal Intensive Care Center, were collected between February 2020 and March 2021. This study included 63 patients with cervical insufficiency receiving cerclage and 47 control participants. Endo- and exocervical swabs and fluids were collected simultaneously. Shotgun metagenomic sequencing for the microbiome and the measurement of 34 immune-checkpoint proteins and inflammatory cytokines were performed.ResultsFirst, we demonstrated that immune-checkpoint proteins, the key immune-regulatory molecules, could be measured in endocervical and exocervical samples. Secondly, we identified significantly different microenvironments in cervical insufficiency and preterm birth, with precise cervical locations, to provide information about practically useful cervical locations in clinical settings. Finally, the presence of Moraxella osloensis (odds ratio = 14.785; P = 0.037) and chemokine CC motif ligand 2 levels higher than 73 pg/mL (odds ratio = 40.049; P = 0.005) in endocervical samples were associated with preterm birth. Combining M. osloensis and chemokine CC motif ligand 2 yielded excellent performance for predicting preterm birth (area under the receiver operating characteristic curve = 0.846, 95% confidence interval = 0.733-0.925).ConclusionMultiple relationships between microbiomes, immune-checkpoint proteins, and inflammatory cytokines in the cervical microenvironment were identified. We focus on these factors to aid in the comprehensive understanding and therapeutic modulation of local microbial and immunologic compositions for the management of cervical insufficiency and preterm birth

GSK-3β regulates the endothelial-to-mesenchymal transition via reciprocal crosstalk between NSCLC cells and HUVECs in multicellular tumor spheroid models

Abstract Background Chemotherapy used for patients with unresectable lung tumors remains largely palliative due to chemoresistance, which may be due to tumor heterogeneity. Recently, multiple studies on the crosstalk between lung cancer cells and their tumor microenvironment (TME) have been conducted to understand and overcome chemoresistance in lung cancer. Methods In this study, we investigated the effect of reciprocal crosstalk between lung cancer cells and vascular endothelial cells using multicellular tumor spheroids (MCTSs) containing lung cancer cells and HUVECs. Results Secretomes from lung cancer spheroids significantly triggered the endothelial-to-mesenchymal transition (EndMT) process in HUVECs, compared to secretomes from monolayer-cultured lung cancer cells. Interestingly, expression of GSK-3β-targeted genes was altered in MCTSs and inhibition of this activity by a GSK-3β inhibitor induced reversion of EndMT in lung tumor microenvironments. Furthermore, we observed that HUVECs in MCTSs significantly increased the compactness of the spheroids and exhibited strong resistance against Gefitinib and Cisplatin, relative to fibroblasts, by facilitating the EndMT process in HUVECs. Subsequently, EndMT reversion contributed to control of chemoresistance, regardless of the levels of soluble transforming growth factor (TGF)-β. Using the MCTS xenograft mouse model, we demonstrated that inhibition of GSK-3β reduces lung cancer volume, and in combination with Gefitinib, has a synergistic effect on lung cancer therapy. Conclusion In summary, these findings suggest that targeting EndMT through GSK-3β inhibition in HUVECs might represent a promising therapeutic strategy for lung cancer therapy

HO-1089 and HO-1197, Novel Herbal Formulas, Have Antitumor Effects via Suppression of PLK1 (Polo-like Kinase 1) Expression in Hepatocellular Carcinoma

The treatment for hepatocellular carcinoma (HCC), a severe cancer with a very high mortality rate, begins with the surgical resection of the primary tumor. For metastasis or for tumors that cannot be resected, sorafenib, a multi-tyrosine protein kinase inhibitor, is usually the drug of choice. However, typically, neither resection nor sorafenib provides a cure. The drug discovery strategy for HCC therapy is shifting from monotherapies to combination regimens that combine an immuno-oncology agent with an angiogenesis inhibitor. Herbal formulas can be included in the combinations used for this personalized medicine approach. In this study, we evaluated the HCC anticancer efficacy of the new herbal formula, HO-1089. Treatment with HO-1089 inhibited HCC tumor growth by inducing DNA damage-mediated apoptosis and by arresting HCC cell replication during the G2/M phase. HO-1089 also attenuated the migratory capacity of HCC cells via the inhibition of the expression of EMT-related proteins. Biological pathways involved in metabolism and the mitotic cell cycle were suppressed in HO-1089-treated HCC cells. HO-1089 attenuated expression of the G2/M phase regulatory protein, PLK1 (polo-like kinase 1), in HCC cells. HCC xenograft mouse models revealed that the daily oral administration of HO-1089 retarded tumor growth without systemic toxicity in vivo. The use of HO-1197, a novel herbal formula derived from HO-1089, resulted in statistically significant improved anticancer efficacy relative to HO-1089 in HCC. These results suggest that HO-1089 is a safe and potent integrated natural medicine for HCC therapy

Evolution of and Horizontal Gene Transfer in the <i>Endornavirus</i> Genus

<div><p>The transfer of genetic information between unrelated species is referred to as horizontal gene transfer. Previous studies have demonstrated that both retroviral and non-retroviral sequences have been integrated into eukaryotic genomes. Recently, we identified many non-retroviral sequences in plant genomes. In this study, we investigated the evolutionary origin and gene transfer of domains present in endornaviruses which are double-stranded RNA viruses. Using the available sequences for endornaviruses, we found that <i>Bell pepper endornavirus</i>-like sequences homologous to the glycosyltransferase 28 domain are present in plants, fungi, and bacteria. The phylogenetic analysis revealed the glycosyltransferase 28 domain of <i>Bell pepper endornavirus</i> may have originated from bacteria. In addition, two domains of <i>Oryza sativa endornavirus</i>, a glycosyltransferase sugar-binding domain and a capsular polysaccharide synthesis protein, also exhibited high similarity to those of bacteria. We found evidence that at least four independent horizontal gene transfer events for the glycosyltransferase 28 domain have occurred among plants, fungi, and bacteria. The glycosyltransferase sugar-binding domains of two proteobacteria may have been horizontally transferred to the genome of <i>Thalassiosira pseudonana</i>. Our study is the first to show that three glycome-related viral genes in the genus <i>Endornavirus</i> have been acquired from marine bacteria by horizontal gene transfer.</p></div

Phylogenetic relationships of the glycosyltransferase domains derived from BPEV, plants, fungi, and bacteria.

<p>The glycosyltransferase domains of various plants, fungi, and bacteria that are homologous to that of BPEV were identified by BLAST searching in several databases. The C-terminal regions of the aligned glycosyltransferase domains were used for the generation of the phylogenetic tree. A total of 93 amino acid sequences, including 54 plant sequences (in green), 21 fungal sequences (in brown), 17 bacterial sequences (in black) and the BPEV sequence (in red), were analyzed. The labels of the branches represent the aLRT values calculated using a SH-like method, and only values greater than 0.5 are displayed.</p

The phylogenetic relationships of two domains of OsEV and homologous proteins from other organisms.

<p>A phylogenetic tree based on the glycosyltransferase sugar-binding domain (approximately 70 amino acids) (A) and a phylogenetic tree based on the capsular polysaccharide synthesis proteins (approximately 90 amino acids) (B) derived from different organisms and OsEV. The branch colors indicate the kingdom of each organism (bacteria in black, fungi in brown, plants in green, and viruses in red). The aLRT values were calculated using a SH-like method, and values greater than 0.5 are shown on the branches.</p

Identification of plant sequences homologous to BPEV.

<p>(A) Schematic diagram of BPEV and the corresponding locations of the plant sequences homologous to BPEV. The black bar indicates the whole proteome of BPEV, and each domain of BPEV is indicated by a symbol of a different color. The small green fragments within dotted line boxes indicate the partial plant sequences homologous to BPEV with the respective names. The abbreviated protein names can be found in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0064270#pone-0064270-t001" target="_blank">Table 1</a>. (B) Amino acid alignment of the glycosyltransferase domains of BPEV and identified plant proteins using ClustalW. (C) Phylogenetic tree based on the glycosyltransferase domains of BPEV and 30 identified plant proteins constructed using the PhyML 3.0 server. The numbers on the branches are the aLRT values calculated using a SH-like method. Numbers greater than 0.5 are shown on each branch.</p

Endogenous EBPEs identified in fungi and bacteria.

<p>Endogenous EBPEs identified in fungi and bacteria.</p

Endogenous <i>Bell pepper endornavirus</i>-like sequences (EBPEs) identified in various plants.

<p>The accession number of each protein can be found in the peptide data for the individual plant species deposited in Phytozome. Abbreviation: endogenous <i>Bell pepper endornavirus</i>-like sequences (EBPEs). Each identified protein is named after the plant species. For example, EBPE1 from <i>Ricinus communis</i> is referred to as RcEBPE1. All identified EBPEs in plants are homologous to the sequence of the glycosyltransferase (GT) 28 domain in <i>Bell pepper endornavirus</i> (Accession No. NC_015781).</p