SinCWIm: An imputation method for single-cell RNA sequence dropouts using weighted alternating least squares.

BACKGROUND AND OBJECTIVES: Single-cell RNA sequencing (scRNA-seq) provides a powerful tool for exploring cellular heterogeneity, discovering novel or rare cell types, distinguishing between tissue-specific cellular composition, and understanding cell differentiation during development. However, due to technological limitations, dropout events in scRNA-seq can mistakenly convert some entries in the real data to zero. This is equivalent to introducing noise into the data of cell gene expression entries. The data is contaminated, which affects the performance of downstream analyses, including clustering, cell annotation, differential gene expression analysis, and so on. Therefore, it is a crucial work to accurately determine which zeros are due to dropout events and perform imputation operations on them. METHODS: Considering the different confidence levels of different zeros in the gene expression matrix, this paper proposes a SinCWIm method for dropout events in scRNA-seq based on weighted alternating least squares (WALS). The method utilizes Pearson correlation coefficient and hierarchical clustering to quantify the confidence of zero entries. It is then combined with WALS for matrix decomposition. And the imputation result is made close to the actual number by outlier removal and data correction operations. RESULTS: A total of eight single-cell sequencing datasets were used for comparative experiments to demonstrate the overall superiority of SinCWIm over state-of-the-art models. SinCWIm was applied to cluster the data to obtain an adjusted RAND index evaluation, and the Usoskin, Pollen and Bladder datasets scored 94.46%, 96.48% and 76.74%, respectively. In addition, significant improvements were made in the retention of differential expression genes and visualization. CONCLUSIONS: SinCWIm provides a valuable imputation method for handling dropout events in single-cell sequencing data. In comparison to advanced methods, SinCWIm demonstrates excellent performance in clustering, visualization and other aspects. It is applicable to various single-cell sequencing datasets.

RPIPCM: A deep network model for predicting lncRNA-protein interaction based on sequence feature encoding.

LncRNA-protein interactionplays an important regulatory role in biological processes. In this paper, the proposed RPIPCM based on a novel deep network model uses the sequence feature encoding of both RNA and protein to predict lncRNA-protein interactions (LPIs). A negative sampling of sliding window method is proposed for solving the problem of unbalanced between positive and negative samples. The proposed negative sampling method is effective and helpful to solve the problem of data imbalance in the existing LPIs research by comparative experiments. Experimental results also show that the proposed sequence feature encoding method has good performance in predicting LPIs for different datasets of different sizes and types. In the RPI488 dataset related to animal, compared with the direct original sequence encoding model, the accuracy of sequence feature encoding model increased by 1.02%, the recall increased by 4.08%, and the value of MCC increased by 1.67%. In the case of the plant dataset ATH948, the sequence feature-based encoding demonstrated a 1.58% higher accuracy, a 1.53% higher recall, a 1.62% higher specificity, a 1.62% higher precision, and a 3.16% higher value of MCC compared to the direct original sequence-based encoding. Compared with the latest prediction work in the ZEA22133 dataset, RPIPCM is shown to be more effective with the accuracy increased by 2.23%, the recall increased by 1.78%, the specificity increased by 2.67%, the precision increased by 2.52%, and the value of MCC increased by 4.43%, which also proves the effectiveness and robustness of RPIPCM. In conclusion, RPIPCM of deep network model based on sequence feature encoding can automatically mine the hidden feature information of the sequence in the lncRNA-protein interaction without relying on external features or prior biomedical knowledge, and its low cost and high efficiency can provide a reference for biomedical researchers.

Comparison of NK cell subsets, receptors and functions induced by radiofrequency ablation and microwave ablation in HBV-associated primary hepatocellular carcinoma.

Background: Topical therapy has been shown to induce an immune response in patients with hepatocellular carcinoma (HCC). In this study, a prospective parallel group control experiment was conducted to compare the differences between radiofrequency ablation and microwave ablation in inducing the immune regulation of NK cells. Methods: Sixty patients with clinically and pathologically confirmed hepatitis B-associated hepatocellular carcinoma (HCC) were selected for thermal ablation. Patients were randomly assigned into the MWA group (n = 30) and the RFA group (n = 30). Patient's peripheral blood was isolated on days D0, D7, and month M1. NK cell subsets, receptors, and killing function were detected by flow cytometry and LDH. Student t test and rank sum test were used to compare the statistical differences between the RFA (radio frequency) and MWA (microwave) groups. The Kaplan-Meier curve and log-rank test were used to calculate the difference between the two survival curves. Results: Comparison of the frequency of CD3-CD56+ and CD3-CD56+CD16+ in NK cells between the RFA and WMA groups showed that there was no difference in the D0, D7, M1, D7-D0, M1-D0, and M1-D7 groups. The changes of the inhibitory NK cell receptor CD159A were significantly different at D7 (P<0.05). CD107a were compared between the RFA and WMA groups, indicating that CD107a changes induced by NK cells were significantly different at D7-D0 (P<0.05). Comparison of NK cell lysis activity of target K562 cells between the RFA and WMA groups showed that there was no difference at D0, D7, D7-D0. There was no difference in recurrence-free survival (RFS) between the RFA and WMA groups (P=0.11). Conclusions: The difference between MWA and RFA-induced NK cell changes was mainly manifested in the inhibitory receptors CD159a and CD107a 1 week after surgery, with microwave-induced changes being more severe. Comparison of the NK cell lysis activity of the target K562 cells between the RFA and WMA groups showed that there was no difference in D0, D7, D7- D0. Survival analysis showed that these differences did not affect the recurrence-free survival (RFS) in the two groups.

Dynamic changes of phenotype and function of natural killer cells in peripheral blood before and after thermal ablation of hepatitis B associated hepatocellular carcinoma and their correlation with tumor recurrence.

BACKGROUND: Thermal therapy induces an immune response in patients with hepatocellular carcinoma (HCC), but the dynamic characteristics of the natural killer (NK) cell immune response post-thermal ablation remain unclear. We conducted a prospective longitudinal cohort study to observe the dynamic changes of phenotype and function of NK cells in peripheral blood before and after thermal ablation of hepatitis B-associated HCC and their correlation with tumor recurrence. METHODS: Fifty-six patients clinically and pathologically confirmed with hepatitis B-associated HCC were selected for thermal ablation. Peripheral blood was collected on day 0, day 7, and month 1. NK cell subsets, receptors, and killing function were detected by flow cytometry, and the LDH levels were examined. Overall recurrence and associated variables were estimated using Kaplan-Meier, log-rank, and Cox proportional-hazards analyses. RESULTS: The frequency of CD3-CD56+ cells was increased on day 7 (P < 0.01) without significant differences between D0 and M1. NKG2D, NKp44, NKp30, CD159a, and CD158a expression was increased on M1 (all P < 0.05). The granzyme B and IFN-γ expression in NK cells were higher on M1 vs. D0 (P < 0.05). On day 7, the NK cell lysis activity of the target K562 cells was increased (P < 0.01) but decreased on M1 (P < 0.05). Survival analysis showed that CD158a expression and IFN-γ and perforin release on day 0 were associated with the risk of HCC recurrence. Cox regression analysis showed that the expression changes in CD56, NKp46, granzyme B, and perforin (D7-D0) induced by thermal ablation were associated with recurrence-free survival (RFS) of patients with HCC. CONCLUSION: Thermal ablation increased the frequency and function of CD3-CD56+ NK cells in the peripheral blood of patients with HCC. These cells tended to be more differentiated and activated. Notably, expression levels of NK cell receptors NKp46, perforin, and granzyme B were associated with RFS.

Identification of miRNA and mRNA expression profiles by PCR microarray in hepatitis B virus­associated hepatocellular carcinoma.

The present study aimed to identify differentially expressed microRNAs (miRNAs) and mRNAs in hepatitis B virus­associated hepatocellular carcinoma (HCC). A total of five HCC tissues and paired adjacent non­tumor tissues were screened to identify the differentially expressed miRNAs and target mRNAs using polymerase chain reaction microarrays. The interaction between differential miRNA and mRNA expression was concurrently analyzed using bioinformatics methods. A total of 32 differentially expressed miRNAs (four upregulated miRNAs and 28 downregulated miRNAs) and 16 differentially expressed mRNAs (11 upregulated mRNAs and five downregulated mRNAs) were identified. Among these, upregulated hsa­miRNA (miR)­96­5p and hsa­miR­18b­5p suppressed their target mRNAs forkhead box O1 and MET transcriptional regulator MACC1 (MACC1). Downregulation of hsa­miR­199a­5p led to upregulation of its target mRNAs, cyclin dependent kinase 4 and insulin like growth factor 2 (IGF2). The high­level expression of IGF2 mRNA and cyclin E1 mRNA was due to the low­level expression of hsa­miR­145­5p, hsa­miR­181a­5p, hsa­miR­199a­5p and hsa­miR­223a­3p, and hsa­miR­26a­5p and hsa­miR­26b­5p, respectively. The low­level expression of coronin 1A mRNA and MACC1 mRNA was due to overexpression of hsa­miR­517a­3p and hsa­miR­18a­5p, and hsa­miR­18b­5p, respectively. Numerous gene ontology terms were associated with oncogenesis. The most enriched pathways targeted by the dysregulated miRNAs and mRNAs were associated with cancer and oncogenesis pathways. The present data suggested that differential miRNA and mRNA expression is present in HCC. Thus, interactions between certain miRNAs and mRNAs may be involved in the pathogenesis of HCC.

C-arm Lipiodol CT in transcatheter arterial chemoembolization for small hepatocellular carcinoma.

AIM: To investigate the value of C-arm Lipiodol computed tomography (CT) for intra-procedural hepatocellular carcinoma (HCC) lesion detection during transcatheter arterial chemoembolization (TACE). METHODS: Forty patients (37 male, 3 female; mean age, 52.6 ± 12.5 years, age range: 25-82 years) diagnosed with HCC were enrolled in this study. All patients underwent 64-slice CT 1-2 wk before TACE. During the procedure, hepatic angiography was performed first. Following diagnostic embolization with Lipiodol injected into the hepatic artery, a C-arm CT scan was immediately conducted (C-arm Lipiodol CT). If new HCC lesions were confirmed, gelfoam particles were super-selectively injected into the tumor-nourishing blood vessel. A Lipiodol CT scan was performed 7-14 d after TACE. All images acquired from 64-slice CT, digital subtraction angiography (DSA), C-arm Lipiodol CT and Lipiodol CT were retrospectively reviewed by four radiologists and the number of detected lesions in each examination was counted, respectively. The results of Lipiodol CT were taken as the diagnostic reference. Alpha-fetoprotein values were examined both before and after TACE. This study only takes into account the lesions that were not found or were considered suspicious on 64-slice CT before TACE. RESULTS: Preprocedural 64-slice CT detected a total of 13 suspicious lesions in the 40 patients. DSA detected ten definite and four suspicious lesions. C-arm Lipiodol CT detected 71 lesions in total and Lipiodol CT confirmed 67 lesions with a diameter range of 3-12 mm. Four false-positive lesions, which were detected by C-arm Lipiodol CT, were considered to be hepatic artery-portal vein fistulas. The average alpha-fetoprotein values before and after TACE were significantly different (452.3 ± 192.6 ng/mL vs 223.8 ± 93.2 ng/mL; P = 0.039). CONCLUSION: C-arm Lipiodol CT has a higher diagnostic sensitivity for small HCC lesions. This technique may help physicians make intraprocedural decisions to provide patients with earlier treatment.

[Transgenic mouse models of the truncated platelet integrin ß3 cytoplasmic tail established by stem cell transplantation].

This study was purpose to establish the transgenic mouse models of the truncated platelet integrin ß3 by retrovirus-infected hematopoietic stem cells (HSCs) transplantation and to provide the basis for further study of the role of integrin ß3 cytoplasmic domain in platelet bi-directional signaling pathways. Wild-type ß3, ß3-Δ759 (R(760) GT(762) truncated ß3) and ß3-Δ754 (T(755) NITYRGT(762) truncated ß3) cDNAs were subcloned into MSCV MigR1 retroviral vector bearing a GFP gene and packaged into infective retrovirus with BOSC23 cell strain. The bone marrow HSCs of the ß3 deficient mice were infected by the retroviruses, and transplanted into lethally-irradiated wild type C57BL/6 mice. GFP positive rate and surface ß3 expression of the recipients' platelets at 6 to 8 weeks after transplantation were detected by flow cytometry to evaluate the transgenic efficiency. The results showed that four kinds of transgenic mouse models including vector, wild-type ß3, ß3-Δ759 and ß3-Δ754 were established successfully. GFP positive rates of transgenic mouse platelets ranged from 18% to 66% and the ß3 expression of transgenic mouse reached heterozygote (ß3(+/-) level of mouse). It is concluded that establishment of transgenic mouse models mediated by retrovirus-infected HSCs transplantation is a feasible, fast, and high throughput transgenic approach and laid a solid foundation for further research on the role of integrin ß3 cytoplasmic domain for bi-directional signaling of platelets in vivo, and for the gene therapy of platelet disorders.

Genetic analysis of liver metastatic cell lines with different metastatic potential.

Metastasis is the major feature of malignant tumors that causes 90% of cancer deaths. Our laboratory has already established liver metastatic clones with YCC-16, isolated from the blood of a gastric cancer patient and expanded in vitro culture using a repeated orthotopic implantation method, and had reported biologic behaviour of the parental YCC-16, the orthotopic primary S1L0, and S1L1, S2L2 and S3L3 liver metastatic clones. Here, using these cell lines, we screened from chromosomal abnormalities using karyotype analysis and micro-CGH matching. There were 31 genes screened using PCA method which were functionally related to cell adhesion. Also, there were 23 genes selected which were related to the liver specific metastasis but excluded genes related to adhesion. There were 4 genes which demonstrated reduced or increased expression stepwise with passage. In conclusion, our results should contribute to exploring the mechanisms of liver metastasis by gastric cancer.