Power analysis becomes an inevitable step in experimental design of current biomedical research. Complex designs allowing diverse correlation structures are commonly used in RNA-Seq experiments. ...However, the field currently lacks statistical methods to calculate sample size and estimate power for RNA-Seq differential expression studies using such designs. To fill the gap, simulation based methods have a great advantage by providing numerical solutions, since theoretical distributions of test statistics are typically unavailable for such designs.
In this paper, we propose a novel simulation based procedure for power estimation of differential expression with the employment of generalized linear mixed effects models for correlated expression data. We also propose a new procedure for power estimation of differential expression with the use of a bivariate negative binomial distribution for paired designs. We compare the performance of both the likelihood ratio test and Wald test under a variety of simulation scenarios with the proposed procedures. The simulated distribution was used to estimate the null distribution of test statistics in order to achieve the desired false positive control and was compared to the asymptotic Chi-square distribution. In addition, we applied the procedure for paired designs to the TCGA breast cancer data set.
In summary, we provide a framework for power estimation of RNA-Seq differential expression under complex experimental designs. Simulation results demonstrate that both the proposed procedures properly control the false positive rate at the nominal level.
rQNestin34.5v.2 is an oncolytic herpes simplex virus 1 (oHSV) that retains expression of the neurovirulent ICP34.5 gene under glioma-selective transcriptional regulation. To prepare an ...investigational new drug (IND) application, we performed toxicology and efficacy studies of rQNestin34.5v.2 in mice in the presence or absence of the immunomodulating drug cyclophosphamide (CPA). ICP34.5 allows HSV1 to survive interferon and improves viral replication by dephosphorylation of the eIF-2α translation factor. rQNestin34.5v.2 dephosphorylated eIF-2α in human glioma cells, but not in human normal cells, resulting in significantly higher cytotoxicity and viral replication in the former compared to the latter. In vivo toxicity of rQNestin34.5v.2 was compared with that of wild-type F strain in immunocompetent BALB/c mice and athymic mice by multiple routes of administration in the presence or absence of CPA. A likely no observed adverse effect level (NOAEL) dose for intracranial rQNestin34.5v.2 was estimated, justifying a phase 1 clinical trial in recurrent glioma patients (ClinicalTrials.gov: NCT03152318), after successful submission of an IND.
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Sample size calculation and power estimation are essential components of experimental designs in biomedical research. It is very challenging to estimate power for RNA-Seq differential expression ...under complex experimental designs. Moreover, the dependency among genes should be taken into account in order to obtain accurate results.
In this paper, we propose a simulation based procedure for power estimation using the negative binomial distribution and assuming a generalized linear model (at the gene level) that considers the dependence between gene expression level and its variance (dispersion) and also allows equal or unequal dispersion across conditions. We compared the performance of both Wald test and likelihood ratio test under different scenarios. The null distribution of the test statistics was simulated for the desired false positive control to avoid excess false positives with the usage of an asymptotic chi-square distribution. We applied this method to the TCGA breast cancer data set.
We provide a framework for power estimation of RNA-Seq data. The proposed procedure is able to properly control the false positive error rate at the nominal level.
The role of the immune response to oncolytic Herpes simplex viral (oHSV) therapy for glioblastoma is controversial because it might enhance or inhibit efficacy. We found that within hours of oHSV ...infection of glioblastomas in mice, activated natural killer (NK) cells are recruited to the site of infection. This response substantially diminished the efficacy of glioblastoma virotherapy. oHSV-activated NK cells coordinated macrophage and microglia activation within tumors. In vitro, human NK cells preferentially lysed oHSV-infected human glioblastoma cell lines. This enhanced killing depended on the NK cell natural cytotoxicity receptors (NCRs) NKp30 and NKp46, whose ligands are upregulated in oHSV-infected glioblastoma cells. We found that HSV titers and oHSV efficacy are increased in Ncr1(-/-) mice and a Ncr1(-/-) NK cell adoptive transfer model of glioma, respectively. These results demonstrate that glioblastoma virotherapy is limited partially by an antiviral NK cell response involving specific NCRs, uncovering new potential targets to enhance cancer virotherapy.
The contribution of the tumor microenvironment to the development of pancreatic adenocarcinoma (PDAC) is unclear. The LSL-KrasG12D/+;LSL-p53R172H/+;Pdx-1-Cre (KPC) tumor model, which is widely ...utilized to faithfully recapitulate human pancreatic cancer, depends on Cre-mediated recombination in the epithelial lineage to drive tumorigenesis. Therefore, specific Cre-loxP recombination in stromal cells cannot be applied in this model, limiting the in vivo investigation of stromal genetics in tumor initiation and progression. To address this issue, we generated a new Pdx1FlpO knock-in mouse line, which represents the first mouse model to physiologically express FlpO recombinase in pancreatic epithelial cells. This mouse specifically recombines Frt loci in pancreatic epithelial cells, including acinar, ductal, and islet cells. When combined with the Frt-STOP-Frt KrasG12D and p53Frt mouse lines, simultaneous Pdx1FlpO activation of mutant Kras and deletion of p53 results in the spectrum of pathologic changes seen in PDAC, including PanIN lesions and ductal carcinoma. Combination of this KPF mouse model with any stroma-specific Cre can be used to conditionally modify target genes of interest. This will provide an excellent in vivo tool to study the roles of genes in different cell types and multiple cell compartments within the pancreatic tumor microenvironment.
Initially characterized as axon guidance factors, semaphorins also have been implicated to have critical roles in multiple physiological and developmental functions, including the regulation of ...immune responses, angiogenesis, organ formation, and the etiology of multiple forms of cancer. Moreover, their contribution in immunity and the regulation of tumour microenvironment is becoming increasingly recognized. Here, we provide a comprehensive analysis of class-3 semaphorins, the only secreted family of genes among veterbrate semaphorins, in terms of their expression profiles and their association with patient survival. We also relate their role with immune subtypes, tumour microenvironment, and drug sensitivity using a pan-cancer study.
Expression profiles of class-3 semaphorins (SEMA3s) and their association with patient survival and tumour microenvironment were studied in 31 cancer types using the TCGA pan-cancer data. The expression of SEMA3 family varies in different cancer types with striking inter- and intra- cancer heterogeneity. In general, our results show that SEMA3A, SEMA3C, and SEMA3F are primarily upregulated in cancer cells, while the rest of SEMA3s are mainly down-regulated in the tested tumours. The expression of SEMA3 family members was frequently associated with patient overall survival. However, the direction of the association varied with regards to the particular SEMA3 isoform queried and the specific cancer type tested. More specifically, SEMA3A and SEMA3E primarily associate with a poor prognosis of survival, while SEMA3G typically associates with survival advantage. The rest of SEMA3s show either survival advantage or disadvantage dependent on cancer type. In addition, all SEMA3 genes show significant association with immune infiltrate subtypes, and they also correlate with level of stromal cell infiltration and tumour cell stemness with various degrees. Finally, our study revealed that SEMA3 genes, especially SEMA3C and SEMA3F may contribute to drug induced cancer cell resistance.
Our systematic analysis of class-3 semaphorin gene expression and their association with immune infiltrates, tumour microenvironment and cancer patient outcomes highlights the need to study each SEMA3 member as a separate entity within each specific cancer type. Also our study validated the identification of class-3 semaphorin signals as promising therapeutic targets in cancer although further laboratory validation still needed.
Omicron subvariants continuingly challenge current vaccination strategies. Here, we demonstrate nearly complete escape of the XBB.1.5, CH.1.1, and CA.3.1 variants from neutralizing antibodies ...stimulated by three doses of mRNA vaccine or by BA.4/5 wave infection, but neutralization is rescued by a BA.5-containing bivalent booster. CH.1.1 and CA.3.1 show strong immune escape from monoclonal antibody S309. Additionally, XBB.1.5, CH.1.1, and CA.3.1 spike proteins exhibit increased fusogenicity and enhanced processing compared with BA.2. Homology modeling reveals the key roles of G252V and F486P in the neutralization resistance of XBB.1.5, with F486P also enhancing receptor binding. Further, K444T/M and L452R in CH.1.1 and CA.3.1 likely drive escape from class II neutralizing antibodies, whereas R346T and G339H mutations could confer the strong neutralization resistance of these two subvariants to S309-like antibodies. Overall, our results support the need for administration of the bivalent mRNA vaccine and continued surveillance of Omicron subvariants.
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•Bivalent booster induces 2- to 8-fold higher nAb titer than monovalent against XBB and XBB.1.5•CH.1.1 and CA.3.1 exhibit nearly complete escape of neutralization from bivalent booster•XBB.1.5, CH.1.1, and CA.3.1 show increased fusogenicity compared with BA.2•Homology modeling shows impacts of F486P mutation present in XBB.1.5 on ACE2 binding
Qu et al. show that bivalent booster recipients, compared with monovalent recipients, exhibit higher nAb titers against Omicron subvariants XBB, XBB.1, and XBB.1.5. The CH.1.1 and CA.3.1 variants show more substantial neutralization escape than the XBB variants. Further, structural modeling reveals that the F486P mutation in XBB.1.5 enhances ACE2 binding.
Oxidative stress plays a crucial role in neurodegenerative diseases like Parkinson's and Alzheimer's. Studies indicate the relationship between oxidative stress and the brain damage caused by a ...high‐fat diet. It is previously found that a lupin protein hydrolysate (LPH) has antioxidant effects on human leukocytes, as well as on the plasma and liver of Western diet (WD)‐fed ApoE−/− mice. Additionally, LPH shows anxiolytic effects in these mice. Given the connection between oxidative stress and anxiety, this study aimed to investigate the antioxidant effects of LPH on the brain of WD‐fed ApoE−/− mice. LPH (100 mg kg−1) or a vehicle is administered daily for 12 weeks. Peptide analysis of LPH identified 101 amino acid sequences (36.33%) with antioxidant motifs. Treatment with LPH palliated the decrease in total antioxidant activity caused by WD ingestion and regulated the nitric oxide synthesis pathway in the brain of the animals. Furthermore, LPH increased cerebral glutathione levels and the activity of catalase and glutathione reductase antioxidant enzymes and reduced the 8‐hydroxy‐2’‐deoxyguanosine levels, a DNA damage marker. These findings, for the first time, highlight the antioxidant activity of LPH in the brain. This hydrolysate could potentially be used in future nutraceutical therapies for neurodegenerative diseases.
A Lupinus angustifolius protein hydrolysate (LPH), containing 101 sequences with antioxidant activity, improves the antioxidant status in vitro and in the brain of mice after 12 weeks of treatment. Specifically, LPH decreases the inducible nitric oxide synthase (iNOS) gene expression, nitric oxide (NO) and 8‐hydroxy‐2’‐deoxyguanosine (8‐OHdG) production, and increases catalase (CAT) and glutathione reductase (GR) activities and glutathione (GSH) levels.
Gene expression profiling experiments with few replicates lead to great variability in the estimates of gene variances. Toward this end, several moderated t-test methods have been developed to reduce ...this variability and to increase power for testing differential expression. Most of these moderated methods are based on linear models with fixed effects where residual variances are smoothed under a hierarchical Bayes framework. However, they are inadequate for designs with complex correlation structures, therefore application of moderated methods to linear models with mixed effects are needed for differential expression analysis.
We demonstrated the implementation of the fully moderated t-statistic method for linear models with mixed effects, where both residual variances and variance estimates of random effects are smoothed under a hierarchical Bayes framework. We compared the proposed method with two current moderated methods and show that the proposed method can control the expected number of false positives at the nominal level, while the two current moderated methods fail.
We proposed an approach for testing differential expression under complex correlation structures while providing variance shrinkage. The proposed method is able to improve power by moderation and controls the expected number of false positives properly at the nominal level.
Correctly assessing the amount of blood loss is crucial in order to adequately treat postpartum haemorrhage (PPH) at an early stage and diminish any related symptoms and/or complications.
The aim of ...our study is to analyse correctness in visually estimated blood loss during labour and to measure the differences between subjectively measured and weighted blood losses (ml).
Cross-sectional study
A Swedish maternity unit with 6000 annual births
Midwives employed at a big maternity unit at a hospital in northern Stockholm, Sweden.
Midwives assisting 192 vaginal births were asked to visually estimate the blood loss from the assisted delivery. Coasters and sanitary pads were weighed following the birth. We analysed if there were any differences between subjective measured blood loss (ml) and weighted blood loss. These two methods were also compared to quantify concordance between estimated blood volume and the actual volume.
The number of overestimates of blood loss was 45.3 % (n=87) with an average of 72.9 ml; the number of underestimates was 49.4 % (n=95) with an average of 73.8 ml. Exact correct estimations of blood loss were done in 5.2 % of the cases (n=10).
The largest overestimation of a postpartum bleeding was by 520 ml; the largest underestimation was by 745 ml.
There was both underestimation and overestimation of blood loss. We found small but significant overestimates in PPH < 300 ml (16 ml). In PPH > 300 ml, there was a small but not significant underestimates (34 ml). Based upon our findings, we conclude that it is reasonable to start weighing blood loss when it exceeds 300 ml.
•Both underestimating and overestimating blood loss was done.•Exact correct estimates of PPH were done in 5.2 % of the cases (n=10).•In 83 % of the cases (n=160), the absolute estimation error is less than 120 ml.
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