High-Throughput Discovery and Characterization of Viral Transcriptional Effectors in Human Cells

Viruses encode transcriptional regulatory proteins critical for controlling viral and host gene expression. Given their multifunctional nature and high sequence divergence, it is unclear which viral proteins can affect transcription and which specific sequences contribute to this function. Using a high-throughput assay, we measured the transcriptional regulatory potential of over 60,000 protein tiles across [~]1,500 proteins from 11 coronaviruses and all nine human herpesviruses. We discovered hundreds of new transcriptional effector domains, including a conserved repression domain in all coronavirus Spike homologs, dual activation-repression domains in VIRFs, and an activation domain in six herpesvirus homologs of the single-stranded DNA-binding protein that we show is important for viral replication and late gene expression in KSHV. For the effector domains we identified, we investigated their mechanisms via high-throughput sequence and chemical perturbations, pinpointing sequence motifs essential for function. This work massively expands viral protein annotations, serving as a springboard for studying their biological and health implications and providing new candidates for compact gene regulation tools.

El papel de los parámetros metabólicos de la 18F-FDG PET/TC en el linfoma linfoblástico pediátrico / The role of 18F-FDG PET/CT metabolic parameters in pediatric lymphoblastic lymphoma

Objetivo: Este estudio retrospectivo tuvo como objetivo evaluar el papel de los parámetros metabólicos de la 18F-FDG PET/TC en el linfoma linfoblástico pediátrico (LBL).Métodos: Treinta pacientes con LBL se sometieron a 42 exploraciones. Los parámetros metabólicos, que incluyeron el valor máximo de captación estandarizado (SUVmax), el volumen tumoral metabólico total (TMTV) y la glucólisis de lesión total (TLG), se midieron en la PET/TC basal. Se realizaron análisis univariantes y multivariantes de la supervivencia para evaluar su valor pronóstico. Doce pacientes se sometieron a PET/TC después del régimen de reinducción, y se calcularon la sensibilidad, la especificidad, el valor predictivo positivo (VPP), el valor predictivo negativo (VPN) y la precisión de la PET/TC para predecir la recaída.Resultados: Los pacientes con estadio IV tuvieron una TMTV más alta que los que tenían un estadio III (p=0,031). Además, los pacientes con T-LBL o afectación mediastínica tenían una TMTV y TLG altos (p<0,05). No hubo diferencias significativas en los parámetros metabólicos de la PET/TC entre los pacientes con diferente evolución (p>0,05). Los niños con una TMTV baja (<242,91cm3) tuvieron una mejor EFS a los 3 años comparados con aquellos con una TMTV elevada (88.9% vs. 56,3%; p=0,036). El SUVmax y el TLG no fueron predictivos de la EFS (p=0,874; p=0,152). Sin embargo, ninguno de los parámetros metabólicos de la PET/TC basales fueron factores pronósticos independientes para los resultados del LBL pediátrico. La PET/TC realizada después del régimen de reinducción presentó una mayor sensibilidad (50% vs. 0%) y VPN (90% vs. 83,3%) para predecir la recaída que la TC sola.Conclusiones: Los parámetros metabólicos de la PET/TC de referencia no fueron predictivos de los resultados en los niños con LBL. La PET/TC realizada después del régimen de reinducción tuvo una mejor sensibilidad y VPN que la TC sola, y una exploración

Objective: This retrospective study aimed to evaluate the role of metabolic parameters of 18F-FDG PET/CT in pediatric lymphoblastic lymphoma (LBL).Methods: Thirty patients with LBL underwent 42 scans. Metabolic parameters including maximum standardized uptake value (SUVmax), total metabolic tumor volume (TMTV), and total lesion glycolysis (TLG) were measured at baseline PET/CT. Univariate and multivariate analysis for survival were performed to assess their prognostic value. Twelve patients underwent PET/CT after reinduction regime, and the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and accuracy of PET/CT for predicting relapse were calculated.Results: Patients with stage IV had a higher TMTV than those with stage III (P=0.031). Besides, patients with T-LBL or mediastinal involvement had a high TMTV and TLG (P<0.05). There was no significant difference in PET/CT metabolic parameters between patients with different outcomes (P>0.05). Children with a low TMTV (<242.91cm3) had a better 3-year EFS compared with those with a high TMTV (88.9% vs. 56.3%; P=0.036). SUVmax and TLG were not predictive of EFS (P=0.874; P=0.152). However, none of the metabolic parameters of baseline PET/CT were independent prognostic factors for outcomes of pediatric LBL. PET/CT underwent after reinduction regime present with higher sensitivity (50% vs. 0%) and NPV (90% vs. 83.3%) for predicting relapse than CT alone.Conclusions: Metabolic parameters of baseline PET/CT were not predictive of outcomes in children with LBL. PET/CT done after the reinduction regime had better sensitivity and NPV than CT alone, and a negative scan could be a reliable indicator for sustained remission (AU)

SARS-CoV-2 Infects Syncytiotrophoblast and Activates Inflammatory Responses in the Placenta

SARS-CoV-2 infection during pregnancy leads to an increased risk of adverse pregnancy outcomes. Although the placenta itself can be a target of virus infection, most neonates are virus free and are born healthy or recover quickly. Here, we investigated the impact of SARS-CoV-2 infection on the placenta from a cohort of women who were infected late during pregnancy and had tested nasal swab positive for SARS-CoV-2 by qRT-PCR at delivery. SARS-CoV-2 genomic and subgenomic RNA was detected in 23 out of 54 placentas. Two placentas with high virus content were obtained from mothers who presented with severe COVID-19 and whose pregnancies resulted in adverse outcomes for the fetuses, including intrauterine fetal demise and a preterm delivered baby still in newborn intensive care. Examination of the placental samples with high virus content showed efficient SARS-CoV-2 infection, using RNA in situ hybridization to detect genomic and replicating viral RNA, and immunohistochemistry to detect SARS-CoV-2 nucleocapsid protein. Infection was restricted to syncytiotrophoblast cells that envelope the fetal chorionic villi and are in direct contact with maternal blood. The infected placentas displayed massive infiltration of maternal immune cells including macrophages into intervillous spaces, potentially contributing to inflammation of the tissue. Ex vivo infection of placental cultures with SARS-CoV-2 or with SARS-CoV-2 spike (S) protein pseudotyped lentivirus targeted mostly syncytiotrophoblast and in rare events endothelial cells. Infection was reduced by using blocking antibodies against ACE2 and against Neuropilin 1, suggesting that SARS-CoV-2 may utilize alternative receptors for entry into placental cells.

Short-term high-dose gavage of hydroxychloroquine changes gut microbiota but not the intestinal integrity and immunological responses in mice.

AIMS: Hydroxychloroquine (HCQ), a widely used antimalarial drug, is proposed to treat coronavirus disease 2019 (COVID-19). However, no report is currently available regarding the direct effects of HCQ on gut microbiota, which is associated with the outcomes of elderly patients with COVID-19. Here, we first investigated the effects of HCQ on intestinal microecology in mice. MAIN METHODS: Fifteen female C57BL/6J mice were randomly divided into two groups: HCQ group (n = 10) and control group (n = 5). Mice in the HCQ group were administered with HCQ at dose of 100 mg/kg by gavage daily for 14 days. The feces of mice were collected before and on the 7th and 14th days after HCQ challenge, and then analyzed by 16S rRNA amplicon sequencing. At the end of the experiment, the hematology, serum biochemistry and cytokines were determined, respectively. The mRNA expression of tight junction proteins in colonic tissues were also studied by RT-PCR. KEY FINDINGS: HCQ challenge had no effects on the counts of white blood cells, the levels of serum cytokines, and the gene expression of tight junction proteins in colon. HCQ also did not increase the content of serum d-lactate in mice. Notably, HCQ significantly decreased the diversity of gut microbiota, increased the relative abundance of phylum Bacteroidetes whereas decreased that of Firmicutes. SIGNIFICANCE: Short-term high dose HCQ challenge changes gut microbiota but not the intestinal integrity and immunological responses in mice. Special attention should be paid to the effects of HCQ on intestinal microecology in future clinical use.

Management of Clinical T1N0M0 Esophageal Cancer

BACKGROUND/AIMS: Endoscopic resection is a standard treatment for stage T1a esophageal cancer, with esophagectomy or radical radiation therapy (RT) performed for stage T1b lesions. This study aimed to compare treatment outcomes of each modality for clinical stage T1 esophageal cancer. METHODS: In total, 179 patients with clinical T1N0M0-stage esophageal cancer treated from 2006 to 2016 were retrospectively evaluated. Sixty-two patients with clinical T1a-stage cancer underwent endoscopic resection. Among 117 patients with clinical T1b-stage cancer, 82 underwent esophagectomy, and 35 received chemoradiotherapy or RT. We compared overall survival (OS) and recurrence-free survival (RFS) rates for each treatment modality. RESULTS: The median follow-up time was 32 months (range, 1 to 120 months). The 5-year OS and RFS rates for patients with stage T1a cancer receiving endoscopic resection were 100% and 85%, respectively. For patients with stage T1b, the 5-year OS and RFS rates were 78% and 77%, respectively, for the esophagectomy group; 80% and 44%, respectively, for the RT alone group; and 96% and 80%, respectively, for the chemoradiation group. The esophagectomy group showed significantly higher RFS than the RT alone group (p=0.04). There was no significant difference in RFS between the esophagectomy and chemoradiation groups (p=0.922). Grade 4 or higher treatment-related complications occurred in four patients who underwent esophagectomy. CONCLUSIONS: Endoscopic resection appeared to be an adequate treatment for patients with T1a-stage esophageal cancer. The multidisciplinary approach involving chemoradiation was comparable to esophagectomy in terms of survival outcome without serious complications for T1b-stage esophageal cancer.

Evaluation of the therapeutic effects and micro-structural influence of rTMS on esophageal cancer patients with depression by using DTI / 实用放射学杂志

Objective To investigate the therapeutic effects of repetitive transcranial magnetic stimulation (rTMS)on esophageal cancer patients with depression and the influence on the brain micro-structure by using the DTI technology.Methods Ten esophageal cancer patients with depression (6 male,4 female)were enrolled in this study according to the inclusion criteria.All patients received 1 0 days of rTMS treatment (stimulation frequency:10 Hz;stimulation site left:dorsolateral prefrontal cortex (DLPFC);stimulation intensity:1 10 % rest motor threshold).Before the first time and after the last time of the rTMS treatment,the DTI image acquisition and the coefficient assessment of hamilton depression scale(HAMD),self-rating depression scale(SDS),and self-rating anxiety scale (SAS)were conducted.Comparison of the mean fractional anisotropy (FA)of the depression related brain regions between pre-and post-rTMS was performed.Pearson correlation coefficient was calculated between the changes of FA value and the depression scale changes as well to understand their relationship.Results The HAMD,SAS and SDS were significantly decreased after pos-t rTMS (t＝7.69, P＝0.000;t＝12.86,P＝0.000;t＝10.51,P＝0.000)compared with pre-rTMS.Also,after rTMS depression patients showed significantly increased FA value in the bilateral hippocampus,left pallidum,bilateral thalamus,left middle frontal cortex,bilateral anterior cingulate cortex, and bilateral superior temporal cortex.Significant negative correlation was observed between the FA changes of the left pallidum and SAS(r＝－0.646,P＝0.044),and between the FA changes of right thalamus and HAMD (r＝－0.712,P＝0.021).Conclusion High frequency rTMS over the left DLPFC has significant antidepressant effect on esophageal cancer patients with depression.This may be related to the modulation of rTMS on the micro-structure of the left pallidum and right thalamus.

Next Generation Sequencing Identified a Novel Multi Exon Deletion of the NF1 Gene in a Chinese Pedigree with Neurofibromatosis Type 1.

Neurofibromatosis type 1 (NF1) is a genetic disease involving neurocutaneous abnormalities. Neurofibromatosis type 1 is an autosomal dominant disorder characterized by the neurofibromas and café-au-lait spots. Mutation in the NF 1 gene causes NF1. The NF 1 gene encodes neurofibromin. In this study, we found a 31-year-old Chinese boy with NF1. He presented only with café-au-lait spots over the whole body. The proband's mother had a severe phenotype with neurofibroma and café-au-lait macules over her whole body, mostly in the facial region. A novel multi exon deletion c.(4661+1_4662-1)_(5748+1_5749-1)del; [EX36_39DEL] on the NF 1 gene has been identified in the proband. Quantitative real-time polymerase chain reaction (qPCR) confirmed that this mutation is co-segregated well and was inherited from the proband's mother. The mutation was absent in the proband's father and normal individuals. The novel multi exon deletion results in the formation of a truncated NF1 protein that caused the NF1 phenotype in this family. Our present study also emphasized the significance of rapid, accurate and cost-effective screening for the patient with NF1 by next generation sequencing (NGS).

Retrospective study of combination chemotherapy with etoposide and ifosfamide in patients with heavily pretreated recurrent or persistent epithelial ovarian cancer

OBJECTIVE: This retrospective study is to evaluate the efficacy and toxicity of combination chemotherapy with etoposide and ifosfamide (ETI) in the management of pretreated recurrent or persistent epithelial ovarian cancer (EOC). METHODS: Patients with recurrent or persistent EOC who had measurable disease and at least one chemotherapy regimen were to receive etoposide at a dose of 100 mg/m²/day intravenous (IV) on days 1 to 3 in combination with ifosfamide 1 g/m²/day IV on days 1 to 5, every 21 days. RESULTS: From August 2008 to August 2016, 66 patients were treated with ETI regimen. Most patients were heavily pretreated prior to ETI: 53 (80.3%) patients had received 3 or more chemotherapy regimens. The response rate (RR) of ETI chemotherapy was 18.2% and median duration of response was 6.8 months (range, 0–30). Median survival of all patients was 5 months at a median follow up of 7.2 months. Platinum-free interval (PFI) more than 6 months prior to ETI has statistically significant correlation with overall survival (OS; 9.2 vs. 5.6 months; P=0.029) and RR (34.5% vs. 5.4%; P < 0.010). However, treatment free interval before ETI, number of prior chemotherapy regimen, and optimality of primary surgery did not show significant difference for RR or OS. Grade 3 or 4 hematologic toxicities were observed in 7 cases (3%) of the 232 cycles of ETI. CONCLUSION: The ETI combination regimen shows comparatively low toxicity and modest activity in heavily pretreated recurrent or persistent EOC patients with more than 6 months of PFI after last platinum treatment.

There Is No Advantage to Transpapillary Pancreatic Duct Stenting for the Transmural Endoscopic Drainage of Pancreatic Fluid Collections: A Meta-Analysis

BACKGROUND/AIMS: Options for the endoscopic management of symptomatic pancreatic fluid collections (PFCs) include transmural drainage (TM) alone, transpapillary drainage (TP) alone, or a combination of both drainage method (CD). There have been conflicting reports about the best method. This study performed a meta-analysis to determine whether CD presents an added clinical benefit over TM. METHODS: The included studies compared TM with CD and reported clinical success for both methods. A random-effects model was used to determine the pooled odds ratios (ORs) and the 95% confidence intervals (CIs) for the following outcomes: technical success, clinical success, complications, and recurrence. RESULTS: Nine studies involving a combined total of 604 drainage procedures—373 TMs (62%) and 231 CDs (38%)—were included. CD showed no additional benefit over TM in terms of technical success (OR, 1.12; 95% CI, 0.37–3.37; p=0.85), clinical success (OR, 1.11; 95% CI, 0.65–1.89; p=0.70), recurrence (OR, 1.49; 95% CI, 0.53–4.21; p=0.45), or complications (OR, 1.15; 95% CI, 0.61–2.18; p=0.67). CONCLUSIONS: Pancreatic duct (PD) stenting provides no additional clinical benefit for the TM of PFCs (particularly pseudocysts). Patients undergoing the TM of symptomatic pseudocysts may not require endoscopic retrograde pancreatography (ERP).

Advance in phage display technology for bioanalysis.

Phage display technology has emerged as a powerful tool for target gene expression and target-specific ligand selection. It is widely used to screen peptides, proteins and antibodies with the advantages of simplicity, high efficiency and low cost. A variety of targets, including ions, small molecules, inorganic materials, natural and biological polymers, nanostructures, cells, bacteria, and even tissues, have been demonstrated to generate specific binding ligands by phage display. Phages and target-specific ligands screened by phage display have been widely used as affinity reagents in therapeutics, diagnostics and biosensors. In this review, comparisons of different types of phage display systems are first presented. Particularly, microfluidic-based phage display, which enables screening with high throughput, high efficiency and integration, is highlighted. More importantly, we emphasize the advances in biosensors based on phages or phage-derived probes, including nonlytic phages, lytic phages, peptides or proteins screened by phage display, phage assemblies and phage-nanomaterial complexes. However, more efficient and higher throughput phage display methods are still needed to meet an explosion in demand for bioanalysis. Furthermore, screening of cyclic peptides and functional peptides will be the hotspot in bioanalysis.

A phosphorylation pattern-recognizing antibody specifically reacts to RNA polymerase II bound to exons

The C-terminal domain of RNA polymerase II is an unusual series of repeated residues appended to the C-terminus of the largest subunit and serves as a flexible binding scaffold for numerous nuclear factors. The binding of these factors is determined by the phosphorylation patterns on the repeats in the domain. In this study, we generated a synthetic antibody library by replacing the third heavy chain complementarity-determining region of an anti-HER2 (human epidermal growth factor receptor 2) antibody (trastuzumab) with artificial sequences of 7–18 amino-acid residues. From this library, antibodies were selected that were specific to serine phosphopeptides that represent typical phosphorylation patterns on the functional unit (YSPTSPS)₂ of the RNA polymerase II C-terminal domain (CTD). Antibody clones pCTD-1stS2 and pCTD-2ndS2 showed specificity for peptides with phosphoserine at the second residues of the first or second heptamer repeat, respectively. Additional clones specifically reacted to peptides with phosphoserine at the fifth serine of the first repeat (pCTD-1stS5), the seventh residue of the first repeat and fifth residue of the second repeat (pCTD-S7S5) or the seventh residue of either the first or second repeat (pCTD-S7). All of these antibody clones successfully reacted to RNA polymerase II in immunoblot analysis. Interestingly, pCTD-2ndS2 precipitated predominately RNA polymerase II from the exonic regions of genes in genome-wide chromatin immunoprecipitation sequencing analysis, which suggests that the phosphoserine at the second residue of the second repeat of the functional unit (YSPTSPS)2 is a mediator of exon definition.

Non-clear cell renal carcinoma: comparative analysis of the new and old histological classification in 79 cases / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To compare the old classification and 2004 WHO histological classification of renal cell carcinoma, summarize the differences and possible reasons, and correct the traditional pathological concepts of kidney cancer.</p><p><b>METHODS</b>Specimens of 79 cases histopathologically diagnosed as non-clear cell renal cell carcinomas after radical nephrectomy during 1998 to 2005 in Tianjin Medical University Cancer Hospital were reclassified according to the 2004 WHO renal cell carcinoma histological classification system.</p><p><b>RESULTS</b>After reclassification, there were 14 cases of clear cell renal cell carcinoma (CCRCC), 23 cases of papillary renal cell carcinoma (PRCC), 34 cases of chromophobe renal cell carcinoma (ChRCC), one collecting duct renal cell carcinoma, one unclassified renal cell carcinoma, 5 cases of mixed cell renal cell carcinoma (CCRCC + PRCC 2 cases, CCRCC + ChRCC 2 cases, PRCC + ChRCC 1 case), and one oncocytoma diagnosed.</p><p><b>CONCLUSIONS</b>Some chromophobe renal cell carcinomas and papillary renal cell carcinomas were easier to be diagnosed as granular cell renal cell carcinoma in the past. The eosinophilic cytoplasm similar to that in the granular cells, and some confusion between PRCC and ChRCC are the main reasons. The cellular characteristic features of granular renal cell carcinoma can be found in many types of renal tumors. Granular cell renal cell carcinoma is not an independent entity, therefore, it should be removed from the histological classification of renal cell carcinoma. The diagnosis standard of mixed renal cell carcinoma (MRCC) need to be determined and consummated.</p>

Biodistribution of Iodine-131-Iodomisonidazole and Imaging of Tumor Hypoxia in Mice bearing CT-26 Adenocarcinoma / 대한핵의학회잡지

urpose: Misonidazole is a radiosensitizer that binds in hypoxic cells. The purpose of this study was to find out the feasibility of I-131-Iodomisonidazole (IMISO) for imaging of tumor hypoxia. MATERIALS AND METHODS: Tosyl precursor was dissolved in acetonitrile and I-131-NaI was added to synthesize IMISO. Balb/c mice inoculated with CT-26 adenocarcinoma were injected with IMISO. Mice were sacrificed at 1,2,4,24 hr and % of injected dose per gram of tissue (%ID/g) was determined. For scintigraphy and MRI, mouse bearing CT-26 adenocarcinoma was administered with IMISO and imaging was performed 4 hr after. Then, mouse body was fixed and microtomized slice was placed on radiographic film for autoradiography. RESULTS: %ID/g of tumor was 1.64 (1h), 0.98 (2h), 0.85 (4h) and 0.20 (24h), respectively. At 24h, %ID/g of tumor was higher than that of all other tissues except thyroid. Tumor to muscle ratio increased with time and tumor to blood ratio also increased with time and reached 1.53 at 24 hr. On autoradiogram, tumor was well visualized as an increased activity in central hypoxic area of the tumor which corresponds to the area of high signal intensity on T2-weighted MR image. On scintigraphy, tumor uptake was visualized. CONCLUSION:: This RESULTS suggest that IMISO may have a potential for tumor hypoxia imaging in mouse model. However, further study is needed to improve it's localization in tumor tissue and to achieve acceptable images of tumor hypoxia.

Biodistribution and Scintigraphy of Iodine-131-Iododeoxyadenosine in Rats Bearing Breast Cancer / 대한핵의학회잡지

PURPOSE: I-131 labeled (2'-deoxy-2-iodo-p-D-arabinofuranosyl) adenine (IAD) may be involved in DNA synthesis during active proliferation of tumor cells. We conducted this study to find out the biodistribution of IAD and its feasibility for scintigraphic tumor imaging. MATERIALS AND METHODS: Tosyl acetyl-adenosine was dissolved in acetonitrile, and I-131-NaI was added and heated to synthesize IAD. Female Fisher 344 rats innoculated with breast tumor cells were injected witb 0.27 MBq of IAD. Rats were sacrificed at 0.5, 1, 2, 4, 24h and the % of injected dose per gram of tissue (%ID/g) was determined. For scintigraphy, rats bearing breast cancer were administered with 1.11 MBq of IAD and imaging was perforrned after 2 and 24h. Then, rat body was fixed and rnicrotomized slice was placed on radiographic film for autoradiography, RESULTS: %ID/g of tumor wa.' 0.74 (0.5h), 0.73 (1h), 0.55 (2h), 0.38 (4h), and 0.05 (24h), respectively. At 1h after injection, %ID/g of tumor was higher than that of heart (0.34), liver (0.42), spleen (0.47), kidney (0,69), muscle (0.14), bone (0.33) and intestine (0.51). However, %1D/g of tumor was lower than blood (1.06), lung (0.77), and thyroid (177.71). At 4h, %ID/g of tumor in comparison with other tissue did not change. Tumor contrast expressed by tumor to blood ratio was 0.69 and tumor to muscle ratio was 5.11 at 1h. However, these ratios did not improve through 24h. On autoradiogram and scintigraphy at 2 and 24 hour, the tumor was well visualized. CONCLUSION: This results suggest that Ial) may have a potential for tumor scintigraphy. However, further work is needed to improve localization in tumor tissue.