Association between C-reactive protein and all-cause mortality among critically ill patients with acute kidney injury.

AIM: To explore the relationship between C-reactive protein (CRP) and mortality in critically ill patients with acute kidney injury (AKI). MATERIALS AND METHODS: A total of 580 patients diagnosed with AKI within 48 hours of ICU admission between September 2017 and August 2019 were enrolled. Patients were followed for all-cause mortality in-hospital and then up to 2 years after discharge. We performed two multivariate regression analysis to assess the association between CRP and mortality, and conducted stratified analysis to assess whether the effect of the CRP differed across subgroups. RESULTS: According to initial CRP quartiles, patients were divided into 4 groups (quartile 1, CRP ≤ 2.87 mg/L; quartile 2, CRP: 2.87 - 25.95 mg/L; quartile 3, CRP: 25.95 - 111.51 mg/L; quartile 4, CRP > 111.51 mg/L). Patients with high CRP levels have higher APACHE-II score, longer length of stay in the ICU, and higher mortality. In multivariate regression analysis, high CRP was associated with the increased risk of in-hospital mortality after adjusting for age, gender, surgical grade, heart rate, serum potassium, serum chloride, coronary heart disease, and atherosclerotic cerebral infarction (quartile 4 vs. quartile 1, OR: 3.810, 95% CI: 2.081 - 6.973). For 2-year mortality, the increased trend was still significant with the OR (95% CI) of the quartile 4 group of 5.117 (2.678 - 9.780) after adjusting for confounders. Subgroup analyses detected in each group showed that the in-hospital and 2-year risk of mortality increased with higher CRP levels. CONCLUSION: Higher CRP level was associated with the increased risk of mortality in critically ill patients with AKI.

Evaluation of immature granulocyte parameters in myeloid neoplasms assayed by Sysmex XN hematology analyzer.

Immature granulocytes (IGs) have significance for the diagnosis of myeloid neoplasms (MNs). The current study aims to use a hematology analyzer to evaluate the accuracy of IG parameters in MNs. Blood specimens from 388 patients with MN, 524 with non-hematological neoplasms (non-HNs), including 109 patients with inflammation and 68 undergoing G-CSF administration, and 500 healthy control subjects were analyzed. IG parameters was assayed by Sysmex XN-9000 (XN) and compared with manual assessments. A high level of agreement between IG% derived from XN and manual measurements for MN patients (r = 0.828, p < 0.0001) was revealed but only a moderate correlation for acute myeloid leukemia patients (AML; r = 0.597; p < 0.0001). Bland-Altman bias analysis was conducted, and the results showed that differences in IG% from XN and manual analysis for MN patients were considered clinically insignificant. ROC analysis demonstrated a good performance of IG# (AUC = 0.842) and IG% (AUC = 0.885) assessed by XN for MN patients with cut-off values of 0.200 × 109/L and 1.95%, respectively. IG parameters from Sysmex XN analyzer are helpful for screening of MNs even though granulocyte morphological abnormalities may interfere with IG parameter accuracy.

Red Blood Cell Classification Based on Attention Residual Feature Pyramid Network.

Clinically, red blood cell abnormalities are closely related to tumor diseases, red blood cell diseases, internal medicine, and other diseases. Red blood cell classification is the key to detecting red blood cell abnormalities. Traditional red blood cell classification is done manually by doctors, which requires a lot of manpower produces subjective results. This paper proposes an Attention-based Residual Feature Pyramid Network (ARFPN) to classify 14 types of red blood cells to assist the diagnosis of related diseases. The model performs classification directly on the entire red blood cell image. Meanwhile, a spatial attention mechanism and channel attention mechanism are combined with residual units to improve the expression of category-related features and achieve accurate extraction of features. Besides, the RoI align method is used to reduce the loss of spatial symmetry and improve classification accuracy. Five hundred and eighty eight red blood cell images are used to train and verify the effectiveness of the proposed method. The Channel Attention Residual Feature Pyramid Network (C-ARFPN) model achieves an mAP of 86%; the Channel and Spatial Attention Residual Feature Pyramid Network (CS-ARFPN) model achieves an mAP of 86.9%. The experimental results indicate that our method can classify more red blood cell types and better adapt to the needs of doctors, thus reducing the doctor's time and improving the diagnosis efficiency.

Attention-Aware Residual Network Based Manifold Learning for White Blood Cells Classification.

The classification of six types of white blood cells (WBCs) is considered essential for leukemia diagnosis, while the classification is labor-intensive and strict with the clinical experience. To relieve the complicated process with an efficient and automatic method, we propose the Attention-aware Residual Network based Manifold Learning model (ARML) to classify WBCs. The proposed ARML model leverages the adaptive attention-aware residual learning to exploit the category-relevant image-level features and strengthen the first-order feature representation ability. To learn more discriminatory information than the first-order ones, the second-order features are characterized. Afterwards, ARML encodes both the first- and second-order features with Gaussian embedding into the Riemannian manifold to learn the underlying non-linear structure of the features for classification. ARML can be trained in an end-to-end fashion, and the learnable parameters are iteratively optimized. 10800 WBCs images (1800 images for each type) is collected, 9000 images and five-fold cross-validation are used for training and validation of the model, while additional 1800 images for testing. The results show that ARML achieving average classification accuracy of 0.953 outperforms other state-of-the-art methods with fewer trainable parameters. In the ablation study, ARML achieves improved accuracy against its three variants: without manifold learning (AR), without attention-aware learning (RML), and AR without attention-aware learning. The t-SNE results illustrate that ARML has learned more distinguishable features than the comparison methods, which benefits the WBCs classification. ARML provides a clinically feasible WBCs classification solution for leukemia diagnose with an efficient manner.

Hybrid adversarial-discriminative network for leukocyte classification in leukemia.

PURPOSE: Leukemia is a lethal disease that is harmful to bone marrow and overall blood health. The classification of white blood cell images is crucial for leukemia diagnosis. The purpose of this study is to classify white blood cells by extracting discriminative information from cell segmentation and combining it with the fine-grained features. We propose a hybrid adversarial residual network with support vector machine (SVM), which utilizes the extracted features to improve the classification accuracy for human peripheral white cells. METHODS: Firstly, we segment the cell and nucleus by utilizing an adversarial residual network, which contains a segmentation network and a discriminator network. To extract features that can handle the inter-class consistency problem effectively, we introduce the adversarial residual network. Then, we utilize convolutional neural network (CNN) features and histogram of oriented gradient (HOG) features, which can extract discriminative features from images of segmented cell nuclei. To utilize the representative features fully, a discriminative network is introduced to deal with neighboring information at different scales. Finally, we combine the vectors of HOG features with those of CNN features and feed them into a linear SVM to classify white blood cells into six types. RESULTS: We used three methods to evaluate the effect of leukocyte classification based on 5000 leukocyte images acquired from a local hospital. The first approach is to use the CNN features as the input of SVM to classify leukocytes, which achieved 94.23% specificity, 95.10% sensitivity, and 94.41% accuracy. The use of the HOG features for SVM achieved 83.50% specificity, 87.50% sensitivity, and 85.00% accuracy. The use of combined CNN and HOG features achieved 94.57% specificity, 96.11% sensitivity, and 95.93% accuracy. CONCLUSIONS: We propose a novel hybrid adversarial-discriminative network for the classification of microscopic leukocyte images. It improves the accuracy of cell classification, reduces the difficulty and time pressure of doctors' work, and economizes the valuable time of doctors in daily clinical diagnosis.

CD4+CD25+Foxp3+ regulatory T cells suppress NKG2D-mediated NK cell cytotoxicity in peripheral blood.

BACKGROUND: Studies have shown that CD4CD25Foxp3Treg cells suppress NKG2D expression on NK cells via a cell contact-dependent mechanism and increased TGF-ß and IL-10 production in some cancer models. We herein aimed to explore whether CD4CD25Foxp3Tregs suppress NKG2D-mediated NK cell cytotoxicity in peripheral blood and elucidate the exact mechanism underlying this phenomenon. METHODS: To explore the function of NKG2D, NK cell cultures were treated with an NKG2D-blocking antibody to block these receptors. Additionally, TGF-ß- and IL-10-blocking antibodies were added to NK and CD4CD25Foxp3Treg cell cocultures to evaluate whether the latter cells suppress NKG2D expression of NK cells via increasing the production of TGF-ß and IL-10. The expression of NKG2D on NK cells was detected by 3-color flow cytometry, and NK cell activity was assessed by 3 assays: a nonradioactive cytotoxicity assay, an ELISA measuring IFN-γ production and a flow cytometry assay to evaluate CD107a expression. RESULTS: Blocking NKG2D decreased NK cell cytotoxicity, IFN-γ production and CD107a expression. Moreover, blocking TGF-ß and IL-10 substantially increased the NKG2D expression in NK and CD4CD25Foxp3Treg cell cocultures. Similarly, blocking TGF-ß and IL-10 enhanced NK cell cytotoxicity, IFN-γ production and CD107a expression; Transwell insert assays also revealed increased IFN-γ production and CD107a and NKG2D expression. CONCLUSION: CD4CD25Foxp3Tregs suppress NKG2D-mediated NK cell cytotoxicity in peripheral blood via a cell contact-dependent mechanism and increased TGF-ß and IL-10 production.

The miR-599 promotes non-small cell lung cancer cell invasion via SATB2.

MicroRNAs (miRNAs) play important roles in the pathogenesis of many types of cancers by negatively regulating gene expression at posttranscriptional level. Here, we identified that miR-599 is up-regulated in non-small cell lung cancer (NSCLC) patients. It promoted NSCLC cell proliferation by negatively regulating SATB2. In NSCLC cell lines, CCK-8 proliferation assay indicated that the cell proliferation is promoted by miR-599 mimics. Transwell assay showed that miR-599 mimics promoted the invasion and migration of NSCLC cells. Luciferase assays confirmed that miR-599 directly binds to the 3'untranslated region of SATB2, and western blotting showed that miR-599 suppresses the expression of SATB2 at the protein level. This study indicates that miR-599 promotes proliferation and invasion of NSCLC cell lines via SATB2. The miR-599 may represent a potential therapeutic target for NSCLC treatment.

Visfatin Triggers the Cell Motility of Non-Small Cell Lung Cancer via Up-Regulation of Matrix Metalloproteinases.

High levels of visfatin are correlated with worse clinical prognosis of various cancers. Still, the effects and mechanisms of visfatin on progression of non-small cell lung cancer (NSCLC) remain unclear. This study revealed that plasma levels of visfatin in patients with NSCLC (585 ± 287 pg/ml) were significantly (p < 0.01) higher than those in healthy people (142 ± 61.1 pg/ml). The high level of plasma visfatin was found to be significantly (p < 0.05) correlated with TNM stage, lymph node metastasis and distant metastasis. Visfatin treatment can increase the migration and invasion of NSCLC cells via up-regulation of metalloproteinase-2 (MMP-2) and MMP-9. Both si-MMP-2 and si-MMP-9 attenuated visfatin-induced migration of NSCLC cells. The inhibitor of NF-κB, while not ERK1/2, p38-MAPK or PI3K/Akt, can significantly abolish visfatin-induced migration of A549 cells and up-regulation of MMP-2 and MMP-9. Furthermore, visfatin can increase the phosphorylation of IκBα and p65 and the transcription activities of NF-κB in NSCLC cells. ACHP, the inhibitor of IKK-ß, blocked visfatin-induced activation of p65 and up-regulation of MMP-2 and MMP-9. Collectively, our data revealed that visfatin can trigger the in vitro migration and invasion of NSCLC cells via up-regulation of MMPs through activation of NF-κB.

Myeloid-derived suppressor cells regulate T cell and B cell responses during autoimmune disease.

MDSCs are a heterogeneous group of myeloid cells that suppress T cell activity in cancer and autoimmune disease. The effect of MDSCs on B cell function is not clear. Using the CIA model of autoimmune disease, we found an increase in M-MDSCs in the periphery of WT mice with CIA compared with naïve mice. These MDSCs were absent from the periphery of CCR2(-/-) mice that developed exacerbated disease. M-MDSCs, isolated from immunized mice, inhibited autologous CD4(+) T cell proliferation. The M-MDSC-mediated suppression of T cell proliferation was NO and IFN-γ dependent but IL-17 independent. Furthermore, we demonstrated for the first time that M-MDSCs from CIA mice also inhibited autologous B cell proliferation and antibody production. The suppression of B cells by M-MDSCs was dependent on the production of NO and PGE2 and required cell-cell contact. Administration of M-MDSCs rescued CCR2(-/-) mice from the exacerbated CIA phenotype and ameliorated disease in WT mice. Furthermore, adoptive transfer of M-MDSCs reduced autoantibody production by CCR2(-/-) and WT mice. In summary, M-MDSCs inhibit T cell and B cell function in CIA and may serve as a therapeutic approach in the treatment of autoimmune arthritis.

Interleukin-18 synergism with interleukin-2 in cytotoxicity and NKG2D expression of human natural killer cells.

Natural killer (NK) cells play an important role in anti-tumor immunity. Interleukin (IL)-18 is an immunoregulatory cytokine that induces potent NK cell-dependent anti-tumor responses when administrated with other cytokines. In this study, we explored the effects of combining IL-18 and IL-2 on NK cytotoxicity as well as expression levels of the NK cell receptor NKG2D in vitro. Freshly isolated PBMCs were incubated for 48 h with IL-18 and IL-2, then CD107a expression on CD3-CD56+ NK cells was determined by three-colour flow cytometry to evaluate the cytotoxicity of NK cells against human erythroleukemia K562 cells and human colon carcinoma HT29 cells. Flow cytometric analysis was also employed to determine NKG2D expression on NK cells. The combined use of IL-18 and IL-2 significantly increased CD107a expression on NK cells compared with using IL-18 or IL-2 alone, suggesting that the combination of these two cytokines exerted synergistic enhancement of NK cytotoxicity. IL-18 also enhanced NKG2D expression on NK cells when administered with IL-2. In addition, blockade of NKG2D signaling with NKG2D-blocking antibody attenuated the up-regulatory effect of combining IL-18 and IL-2 on NK cytolysis. Our data revealed that IL-18 synergized with IL-2 to dramatically enhance the cytolytic activity of human NK cells in a NKG2D-dependent manner. The results appear encouraging for the use of combined IL-18 and IL-2 in tumor immunotherapy.

Peripheral Foxp3+ regulatory T cells and natural killer group 2, member D expression levels in natural killer cells of patients with colorectal cancer.

Foxp3+ regulatory T cells (Tregs) and natural killer group 2, member D (NKG2D)-positive natural killer (NK) cells are considered to be important in the immune escape of colorectal cancer (CRC). However, the association between these two variables remains obscure. Therefore, in the present study, the levels of peripheral Tregs and NKG2D expression in NK cells and the associations in CRC patients were investigated. A total of 35 CRC patients and 16 healthy controls were enrolled in this study. Flow cytometry was performed to assay Treg numbers and NKG2D expression levels in NK cells in peripheral blood samples. Serum carcino-embryonic antigen (CEA) protein was assayed by electrochemiluminescence. Peripheral Treg numbers were significantly increased (P<0.05), while NKG2D expression levels in NK cells were significantly reduced (P<0.01) in CRC patients compared with healthy controls. However, no significant differences were identified in Treg numbers between CRC patients with and without lymph node metastases and between CRC patients with different clinical stages of CRC. Similarly, no significant differences were detected in NKG2D expression levels in NK cells between the different patient groups. Statistical analysis revealed that increased Treg numbers were not correlated with reduced NKG2D expression levels in NK cells from CRC patients. In addition, no statistical correlation was observed between Treg numbers and serum CEA protein in CRC patients. In conclusion, the upregulation of Tregs was not significantly correlated with the downregulation of NKG2D expression in NK cells in peripheral blood from CRC patients.

Killer cell immunoglobulin-like receptor genotypes and haplotypes with susceptibility to pulmonary tuberculosis infection.

BACKGROUND: Killer cell immunoglobulin-like receptors (KIRs) are expressed on natural killer (NK) cells and T cells and organized in highly polymorphic families. Genetic diversity is an important characteristic of KIR genes. The aim of the study was to investigate the influence of KIR genotypes and halotypes on the risk of pulmonary tuberculosis (PTB). METHODS: A sequence specific primer polymerase chain reaction (SSP-PCR) was employed to amplify the KIR genes and pseudogenes in 139 pulmonary tuberculosis (PTB) patients and 30 healthy controls. The innovative point of our study was the subdivision of the patient group according to sputum smear test (positive and negative). KIR genotype and haplotype frequencies were compared between the PTB group and the control group by Chi-square test, and p < 0.05 was regarded as statistically significant. RESULTS: The genotype AH and FZ14 may be associated with the clearance of Mycobacterium. In addition, haplotype B may be the susceptive haplotype that facilitated the clearance of Mycobacterium and haplotype A may be protective haplotype of PTB. CONCLUSIONS: Therefore, the diversity of genotypes and haplotypes induced an inflammatory reaction that resulted in continuous infection.

The Tim-3/galectin-9 pathway involves in the homeostasis of hepatic Tregs in a mouse model of concanavalin A-induced hepatitis.

T cell immunoglobulin- and mucin-domain-containing molecule-3 (Tim-3) is a negative regulator of interferon (IFN)-γ-secreting CD4(+) Th1 cells and plays a key role in autoimmune diseases. Here, we report that galectin-9 expression was increased in hepatic CD4(+)CD25(+) T cells in a mouse model of concanavalin A (Con A)-induced hepatitis. Moreover, Tim-3 showed increased levels in CD4(+)CD25(+) Foxp3(+) regulatory T cells (Tregs). Further analyses showed that blocking the Tim-3/galectin-9 pathway resulted in the suppression of Tregs in vitro, thereby significantly increasing interferon (IFN)-γ production from hepatic Teffs. Moreover, blockade of Tim-3 in vivo with an anti-Tim-3 antibody exacerbated the acute hepatitis, possibly by increased IFN-γ production. Furthermore, we found that in vitro activation of CD4(+)CD25(-) T cells with the T cell receptor (TCR) plus interleukin 2 (IL-2) up-regulated Tim-3 expression. And the induced Tim-3 interacted with galectin-9 to induce CD4(+) T cell apoptosis which could be partly reversed by blocking Tim-3 signaling. Our results suggested that the Tim-3/galectin-9 pathway plays a critical role in the homeostasis of hepatic Tregs through the elimination induction in Teffs and the inhibition of IFN-γ release, which contributes to the pathogenesis of liver damage and constitutes at least part of the mechanism underlying the induction of hepatitis by Con A.

Urinary podocalyxin positive-element occurs in the early stage of diabetic nephropathy and is correlated with a clinical diagnosis of diabetic nephropathy.

AIMS: We tested whether urinary podocalyxin-positive element (PCX+EL) can be a marker of early stage of diabetic nephropathy (DN). METHODS: DN patients (n=68) and health controls (n=28) were enrolled in this study. Patients were classified into three groups: normoalbuminuria, microalbuminuria and macroalbuminuria. Urinary PCX+EL, serum cystatin C (Scys C) and serum creatinine (SCr) were quantified, and correlations between urinary PCX+EL and urinary albumin, Scys C and SCr were examined. The comparison of diagnosis efficiency among urinary PCX+EL, Scys C and SCr was made by receiver operating characteristic (ROC) analysis. RESULTS: Urinary PCX+EL, Scys C and SCr significantly increased in DN patients compared with controls. Urinary PCX+EL increased significantly in all three patients groups compared with controls. However, the concentration of Scys C and SCr did not increase in normoalbuminuria group. Urinary albumin, Scys C and SCr correlated with urinary PCX+EL. ROC curve analysis indicated that area under the curve (AUC) of urinary PCX+EL (0.966) is higher than that of Scys C (0.857) and SCr (0.726) for discriminating nephropathy between DN patients and controls. CONCLUSION: Urinary PCX+EL may be a noninvasive marker for the early stage of DN.

Elevated plasma level of HMGB1 is associated with disease activity and combined alterations with IFN-α and TNF-α in systemic lupus erythematosus.

Recent studies indicate that high-mobility group box protein 1 (HMGB1) contributes to the pathogenesis of diverse autoimmune disorders. It induces the production of interferon-alpha (IFN-alpha) and tumor necrosis factor alpha (TNF-alpha) in vitro. In the present study, plasma HMGB1, TNF-alpha, and IFN-alpha were determined with ELISA in 37 patients with systemic lupus erythematosus (SLE) and 39 age- and sex-matched healthy controls (HC). The possible associations of these cytokines with disease activities, autoantibodies, and certain laboratory parameters were also explored. The plasma levels of HMGB1, TNF-alpha, and IFN-alpha were increased in SLE patients compared with those of HC (P < 0.05). Moreover, the levels of HMGB1 and TNF-alpha in the active SLE patients were elevated compared with those in inactive patients and HC. Additionally, plasma HMGB1 was positively correlated with peripheral neutrophils, and plasma TNF-alpha was positively correlated with anti-Sm, ESR and CRP, while plasma IFN-alpha was inversely correlated with the age and platelet level in SLE patients. Our data indicated that increased plasma HMGB1 was associated with disease activity in SLE, which was similar to TNF-alpha. High level of plasma IFN-alpha may be related to nephritis and thrombocytopenia in SLE.

Possible association of decreased NKG2D expression levels and suppression of the activity of natural killer cells in patients with colorectal cancer.

Natural-killer group 2 (NKG2), a natural killer (NK) cell receptor, plays a critical role in regulating NK cytotoxicity. In this study, we investigated the expression levels of natural killer group 2 member A (NKG2A) and natural killer group 2 member D (NKG2D) in NK cells as well as the regulatory function of NKG2D in patients with colorectal cancer (CRC). Sixty-two CRC patients and 32 healthy controls were enrolled in this study. The expression levels of NKG2A and NKG2D mRNA in peripheral blood mononuclear cells (PBMCs) were investigated using real-time PCR. Flow cytometry was performed to assay the levels of NKG2A and NKG2D proteins in NK cells. The levels of NKG2D mRNA in PBMCs in the patients were significantly lower than those in the controls [mean ± SD, 1.11 ± 0.60 (CRC patients) vs. 1.65 ± 0.71 (healthy controls); p < 0.01], whereas the 2 groups showed no apparent difference in the levels of NKG2A mRNA (p>0.05). In addition, the patients showed significantly lower NKG2D levels in NK cells than the controls did (71.23% ± 8.31% [CRC patients] vs. 79.39% ± 5.58% [healthy controls]; p < 0.01). However, we observed no distinct difference in the NKG2A expression levels in NK cells between the 2 groups (p> 0.05). Notably, blockage of NKG2D signaling with anti-NKG2D antibodies ex vivo resulted in decreased cytotoxicity and CD107a degranulation. Our data revealed that the decrease in NKG2D expression levels may have been associated with suppression of NK cell activity in CRC patients.