Nanopore sequencing of infectious fluid is a promising supplement for gold-standard culture in real-world clinical scenario.

Introduction: Infectious diseases are major causes of morbidity and mortality worldwide, necessitating the rapid identification and accurate diagnosis of pathogens. While unbiased metagenomic next-generation sequencing (mNGS) has been extensively utilized in clinical pathogen identification and scientific microbiome detection, there is limited research about the application of nanopore platform-based mNGS in the diagnostic performance of various infectious fluid samples. Methods: In this study, we collected 297 suspected infectious fluids from 10 clinical centers and detected them with conventional microbiology culture and nanopore platform-based mNGS. The objective was to assess detective and diagnostic performance of nanopore-sequencing technology (NST) in real-world scenarios. Results: Combined with gold-standard culture and clinical adjudication, nanopore sequencing demonstrated nearly 100% positive predictive agreements in microbial-colonized sites, such as the respiratory and urinary tracts. For samples collected from initially sterile body sites, the detected microorganisms were highly suspected pathogens, and the negative predictive agreements were relatively higher than those in the microbial-colonized sites, particularly with 100% in abscess and 95.7% in cerebrospinal fluid. Furthermore, consistent performance was also observed in the identification of antimicrobial resistance genes and drug susceptibility testing of pathogenic strains of Escherichia coli, Staphylococcus aureus, and Acinetobacter baumannii. Discussion: Rapid NST is a promising clinical tool to supplement gold-standard culture, and it has the potential improve patient prognosis and facilitate clinical treatment of infectious diseases.

MicroRNA-15a inhibits hepatic stellate cell activation and proliferation via targeting SRY-box transcription factor 9.

Accumulating research have indicated that microRNAs are associated with the progression of hepatic fibrosis (HF). Nevertheless, the biological role and function of microRNA (miR)-15a in HF are still unknown. Our data revealed that miR-15a expression was decreased in TGF-ß1-treated LX-2 cells and CCl4-induced mouse model. Additionally, miR-15a could directly target the 3'­untranslated region of SRY-box transcription factor 9 (SOX9) to inhibit its expression. miR-15a overexpression attenuated the viability and invasion, but enhanced apoptosis in LX-2 cells. However, miR-15a knockdown had the opposite effects. Interestingly, SOX9 overexpression reversed the changes in cell viability, invasion and apoptosis mediated by miR-15a overexpression. Moreover, the miR-15a overexpression-mediated collagen I and alpha smooth muscle actin (a-SMA) downregulation were reversed by SOX9 overexpression. Overall, miR-15a could inhibit LX-2 cell viability and HF pathogenesis by targeting SOX9 in vitro and in vivo.

miR-219-3p regulates the occurrence of hepatic fibrosis by targeting Smad2.

Abnormal expression of microRNA (miR)-219-3p has been widely identified in different tumors. However, whether miR-219-3p is involved in the progression of hepatic fibrosis (HF) has never been explored. The present study showed that compared with healthy controls, the levels of miR-291-3p in peripheral blood were decreased in patients with HF. Furthermore, much lower levels of miR-291-3p were identified in fibrotic liver tissues compared with that of normal liver tissues. Receiver operating characteristic curve analysis showed that the levels of miR-291-3p in peripheral blood may screen patients with HF from healthy controls. Reverse transcription quantitative polymerase chain reaction analysis showed that overexpression of miR-291-3p significantly suppressed the mRNA levels of Snai1, vascular endothelial-specific cadherin (VE-cadherin), Vimentin, transforming growth factor (TGF)-ß1, and glial fibrillary acidic protein (GFAP). The protein levels of Snai1, VE-cadherin, Vimentin, TGF-ß1, and GFAP were also decreased in hepatic stellate cells transfected with miR-291-3p mimics. Further study indicated that mothers against decapentaplegic homolog 2 (Smad2) was a target gene of miR-291-3p. More importantly, silencing of Smad2 could abolish miR-291-3p inhibition-induced TGF-ß1 signaling activation. In summary, reduced peripheral blood miR-291-3p may be involved in the progression of HF via targeting Smad2.

High SHIP2 expression indicates poor survival in colorectal cancer.

SH2-containing inositol 5'-phosphatase 2 (SHIP2), which generally regulates insulin signaling, cytoskeleton remodeling, and receptor endocytosis, has been suggested to play a significant role in tumor development and progression. However, the associations between SHIP2 expression and the clinical features to evaluate its clinicopathologic significance in colorectal cancer (CRC) have not been determined yet. In the present study, one-step quantitative real-time polymerase chain reaction (qPCR) test and immunohistochemistry (IHC) analysis with CRC tissue microarrays (TMA) were employed to evaluate the mRNA and protein expression of SHIP2 in CRC. The results showed that SHIP2 expression in the mRNA and protein levels was significantly higher in CRC tissues than that in corresponding noncancerous tissues (both P < 0.05). The expression of SHIP2 protein in CRC was related to lymph node metastasis (P = 0.036), distant metastasis (P = 0.001), and overall survival (P = 0.009). Kaplan-Meier method and Cox multifactor analysis suggested that high SHIP2 protein level (P = 0.040) and positive distant metastasis (P = 0.048) were critically associated with the unfavorable survival of CRC patients. The findings suggested that SHIP2 may be identified as a useful prognostic marker in CRC and targeting CRC may provide novel strategy for CRC treatment.

High expression of MAGE-A9 correlates with unfavorable survival in hepatocellular carcinoma.

Melanoma-associated antigens (MAGE)-A9 has been reported to play important roles in the development of human cancers. However, the association between MAGE-A9 expression and the clinicopathological characteristics of hepatocellular carcinoma (HCC) is not well understood. The study was to detect the expression of MAGE-A9 in human HCC and investigate the association between its expression and the clinicopathological characteristics of HCC. Reverse transcription-polymerase chain reaction (RT-PCR), one-step quantitative -PCR (qPCR) and immunohistochemistry (IHC) analyses were performed to characterize the expression of MAGE-A9 in HCC cell lines and tissues. Kaplan-Meier survival and Cox regression analyses were employed to evaluate the prognosis of 100 HCC patients. The results showed that the expression of MAGE-A9 in HCC was significantly higher than that in non-cancerous cells and tissues. Moreover, the expression level of the MAGE-A9 protein in HCC was related to the pathological grade (p = 0.003), portal vein invasion (p = 0.001), distant metastasis (p = 0.022) and TNM stage (p = 0.005). Cox regression analysis further revealed that MAGE-A9 expression is an independent prognostic factor for disease-free survival (p = 0.006) and overall survival (p = 0.022). These data are the first to indicate that MAGE-A9 expression is a valuable prognostic biomarker for HCC and that high MAGE-A9 expression suggests unfavorable survival outcomes in HCC patients.

TIMP-3 expression associates with malignant behaviors and predicts favorable survival in HCC.

The tissue inhibitors of metalloproteinases (TIMPs) are proteins that specifically inhibit the proteolytic activity of the matrix metalloproteinases (MMPs). TIMP-3, the only member of the TIMPs that can tightly bind to the extracellular matrix, has been identified as a unique tumor suppressor that demonstrates the ability to inhibit tumor angiogenesis, invasion, and metastasis. This study aimed to detect the expression of TIMP-3 in hepatocellular carcinoma (HCC) and investigate the association between TIMP-3 expression and its clinicopathological significance in HCC patients. In the current study, reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting of HCC cell lines and one-step quantitative reverse transcription PCR (qPCR) and immunohistochemistry (IHC) analyses in HCC tissues were performed, to characterize the TIMP-3 expression. Kaplan-Meier survival and Cox regression analyses were utilized to evaluate the prognosis of 101 HCC patients. The results showed that the expression of TIMP-3 in HCC was significantly decreased relative to that of non-cancerous cells and tissues. Furthermore, the TIMP-3 expression was statistically associated with malignant behaviors of HCC, including portal vein invasion (p = 0.036) and lymph node metastasis (p = 0.030). Cox regression analysis revealed that TIMP-3 expression was an independent prognostic factor for disease-free survival (p = 0.039) and overall survival (p = 0.049). These data indicate that TIMP-3 expression is a valuable prognostic biomarker for HCC and that TIMP-3 expression suggests a favorable prognosis for HCC patients.

High expression of inositol polyphosphate phosphatase-like 1 associates with unfavorable survival in hepatocellular carcinoma.

Inositol polyphosphate phosphatase-like 1 (INPPL1), also known as SH2-containing inositol 5'-phosphatase 2 (SHIP2), has been suggested to act downstream of the PI3K/AKT pathway and play an important function in tumor development and progression. However, the associations between SHIP2 expression and the clinical features to determine its clinicopathologic significance in hepatocellular carcinoma (HCC) have not been investigated. In the present study, one-step quantitative PCR reverse transcription-polymerase chain reaction (qPCR) and immunohistochemistry (IHC) analysis with HCC tissue microarrays (TMA) were employed to evaluate the expression of SHIP2 in HCC. The results showed that SHIP2 expression in the mRNA and protein levels was significantly higher in HCC tissue than in corresponding non-cancerous tissue (p = 0.0014 and p < 0.001, respectively). The expression of SHIP2 protein in HCC was related to tumor differentiation, α-fetoprotein level, liver cirrhosis, and five-year survival rate (all p < 0.05). Kaplan-Meier method and log-rank test indicated that high expression of SHIP2 (p = 0.017) and tumor differentiation (p = 0.036) showed significant correlations with poor prognosis of HCC patients. The data indicate that SHIP2 expression is correlated with significant characteristics of HCC, and it may be useful as an unfavorable prognostic factor in HCC.

Elevated expression of SHIP2 correlates with poor prognosis in non-small cell lung cancer.

SH2-containing inositol 5'-phosphatase 2 (SHIP2) is a vital regulator of phosphoinositide pools in metabolic pathways and is considered to downregulate phosphatidylinositol 3'-kinase signaling, which underlies the development of several kinds of human cancers. However, SHIP2 expression in non-small cell lung cancer (NSCLC) and its relationship with the clinical characteristics of NSCLC remain poorly understood. In this study, one-step quantitative reverse transcription-polymerase chain reaction and immunohistochemistry analysis with tissue microarray was used to evaluate SHIP2 expression in NSCLC and to investigate the relationship of this expression to NSCLC prognosis. Results showed that the expression of SHIP2 messenger RNA and protein was significantly higher in NSCLC than in corresponding non-cancerous tissues (both p < 0.05). SHIP2 protein expression in NSCLC was related to lymph node metastasis (p = 0.042), TNM stage (p = 0.036), and 5-year survival rate (p = 0.046). The Kaplan-Meier method and log-rank test suggested that high SHIP2 expression, tobacco consumption, and advanced tumor stage were significantly associated with low survival of NSCLC patients. The results of this research suggested that SHIP2 expression was correlated with malignant phenotypes of NSCLC and may thus serve as a poor prognostic factor and valuable oncogene for NSCLC.