Streptavidin Peroxidase Research Articles

Tetrahedral DNA nanostructures-assisted electrochemical assay for detecting circulating tumor DNA by combining a masking tactic with 3D-hybridization chain reactions

Circulating tumor DNA (ctDNA) is a remarkable noninvasive tumor marker that plays a crucial role in tumor diagnosis, prognosis and treatment. However, detecting low-abundance ctDNA from a substantial amount of nucleic acids originating from healthy cells is challenging. Herein, we proposed a tetrahedral DNA nanostructures (TDNs)-assisted electrochemical biosensor for ctDNA detection. This biosensor combines a masking tactic with 3D-hybridization chain reactions. Masking hairpins (MHs) were initially introduced to prevent interference from wild-type (WT) DNA. Then, the initiator sequence was transferred to the electrode surface modified with TDNs by the target ctDNA. The initiator sequence triggers the 3D self-assembly of hairpin strands, leading to the formation of DNA networks or even DNA hydrogels (long reaction time). This process generates numerous evenly distributed biotin molecules that can bind to streptavidin peroxidase to considerably amplify the signal. This method exhibits high sensitivity (the minimum concentration for detecting ctDNA is 1 aM, which corresponds to 60 ctDNA molecules in 100 μl sample) and excellent specificity (single mismatch). More importantly, this high-performance sensor can detect ctDNA with other mutation sites and their mixtures by modifying the corresponding capture probes on the TDNs. Furthermore, this ultrasensitive sensor effectively detects target ctDNA (0.001%) at high levels of WT DNA and in complex matrices such as serum. These findings suggest that the sensor has promising potential as a noninvasive tool for early tumor diagnosis.

El efecto protector del extracto de Granada contra la úlcera gástrica experimental inducida por etanol en ratas

The Pomegranate (Punica granatum) is a commonly found fruit in the Mediterranean and Iran, which has a variety of uses including medicinal purposes, cosmetics, and as a spice in culinary applications. Pharmacological functions of Pomegranate include antioxidation, anti–tumor, anti–hepatotoxicity, anti–lipoperoxidation and antibacterial properties. The aim of this study was to evaluate the therapeutic efficacy of Pomegranate extract by utilizing its antioxidant activity in an experimental rat model of gastritis induced by ethanol. In the study, 24 female Wistar albino rats (180–200 g) were used. Gastritis in rats was induced using Ethanol. In experimental groups, Tumor necrosis factor–alpha, Myloperoxidase, Superoxide Dismutase and Malondialdehyde were examined for biochemical analyzes. Streptavidin peroxidase immunohistochemistry method was applied to gastric tissues with gastritis. A statistically significant difference was observed between Superoxide Dismutase and Meloperoxidase levels. CD8 and CD68 immunoreactivity was higher in the Ethanol group compared to the other groups. A decrease was observed in CD8 and CD68 positive immunoreactivity in Ethanol+Pomegranate extract group compared to Ethanol group. The study found that the immunoreactivity of MHC–I and MHC–II was found in specific locations, namely intraepithelial lymphocytes located in the epithelium, some capillary vessel endothelium, and connective tissue. Changes in anti–oxidative stress markers such as Superoxide Dismutase and Myloperoxidase contributed to the mucosal protective effect of Pomegranate extract in Ethanol–induced gastritis.

Expression and clinical significance of SYNE3 in non-small cell lung cancer.

To detect the expression of Spectrin Repeat Containing Nuclear Envelope Family Member 3 (SYNE3) and Cluster of Differentiation 34 (CD34) in non-small cell lung cancer (NSCLC). It also aimed to explore the relationship between SYNE3 and NSCLC angiogenesis and clinicopathologic features to identify new biomarkers for NSCLC. Forty-five NSCLC stage IA-IVB tissue specimens from patients diagnosed at Bazhong Central Hospital were collected from January to September 2022, along with 45 para-cancerous normal lung tissues as controls. None of the NSCLC patients had received anti-tumor therapies, including radiotherapy, chemotherapy, targeted therapy, immunotherapy, or traditional Chinese medicine. All specimens were stained for SYNE3 and CD34 using the Streptavidin-Peroxidase (SP) method. The expression levels of SYNE3 and CD34 in NSCLC tissues and para-cancerous tissues were detected, and a correlation analysis between SYNE3 and clinicopathological features was performed. The number of CD34-labeled microvessels was counted using the Microvessel density (MVD) method. The relationship between SYNE3 and NSCLC angiogenesis was examined through the correlation between CD34-MVD and NSCLC clinicopathologic features. The expression of SYNE3 in NSCLC was significantly lower than that in para-cancerous normal lung tissues, while the expression of CD34 in NSCLC was significantly higher than in para-cancerous normal lung tissues (P=0.037). SYNE3 expression in NSCLC was negatively correlated with tumor diameter and was lower in male patients with a smoking history compared to female patients without a smoking history. CD34 expression was positively correlated with Tumor, Node, Metastasis staging and lymph node metastasis. There was a significant correlation between the expression of SYNE3 and CD34 in NSCLC (r=0.450, P=0.000). SYNE3 was lowly expressed and negatively correlated with tumor size in NSCLC, whereas CD34 was highly expressed and positively correlated with TNM stage and lymph node metastasis. The significant correlation between the expressions of SYNE3 and CD34 suggests that SYNE3 may play a key role in NSCLC angiogenesis and progression.

Multiplex Microarrays in 96-Well Plates Photoactivated with 4-Azidotetrafluorobenzaldehyde for the Identification and Quantification of β-Lactamase Genes and Their RNA Transcripts.

Antibiotic-resistant bacteria represent a global issue that calls for novel approaches to diagnosis and treatment. Given the variety of genetic factors that determine resistance, multiplex methods hold promise in this area. We developed a novel method to covalently attach oligonucleotide probes to the wells of polystyrene plates using photoactivation with 4-azidotetrafluorobenzaldehyde. Then, it was used to develop the technique of microarrays in the wells. It consists of the following steps: activating polystyrene, hybridizing the probes with biotinylated target DNA, and developing the result using a streptavidin-peroxidase conjugate with colorimetric detection. The first microarray was designed to identify 11 different gene types and 16 single-nucleotide polymorphisms (SNPs) of clinically relevant ESBLs and carbapenemases, which confer Gram-negative bacteria resistance to β-lactam antibiotics. The detection of bla genes in 65 clinical isolates of Enterobacteriaceae demonstrated the high sensitivity and reproducibility of the technique. The highly reproducible spot staining of colorimetric microarrays allowed us to design a second microarray that was intended to quantify four different types of bla mRNAs in order to ascertain their expressions. The combination of reliable performance, high throughput in standard 96-well plates, and inexpensive colorimetric detection makes the microarrays suitable for routine clinical application and for the study of multi-drug resistant bacteria.

Expression of Notch1 in Ameloblastoma and Correlation with CBCT Imaging Subtypes

Objective: Ameloblastoma (AM) is a prevalent benign odontogenic tumor affecting the jaw bone. Its potential for recurrence and malignant transformation is closely associated with the regulation of signaling pathways. The objective of this study was to examine the expression and clinical significance of Notch1 in AM, along with exploring the correlation between Notch1 expression and AM imaging patterns. Methods: The streptavidin-peroxidase (S-P) immunohistochemical technique was employed to detect the expression of Notch1 protein in the oral mucosa of 70 AM patients and 25 normal controls. Additionally, the preoperative cone-beam computed tomography (CBCT) images of the patients were classified, and the results were analyzed in relation to the expression of Notch1 protein in AM. Results: The positive expression rate of Notch1 in the 70 AM patients was 74.29% (52/70), while the Notch1 expression rate in 26 oral mucosa patients was 19.23% (5/26), indicating a statistically significant difference (P < 0.05). The expression rate of Notch1 was 58.06% in single-room AM and lower in multi-room AM (85.29%) and honeycomb AM (100%) (P < 0.01). However, there was no significant difference in the expression rate of Notch1 between multi-room AM and honeycomb AM (P > 0.05). Conclusion: Notch1 may serve as an important diagnostic indicator in AM, and the apoptosis inhibition ability and cell proliferation ability of single-room AM are lower compared to those of the room and honeycomb types

Effect of paclitaxel octreotide conjugate on human ovarian paclitaxel-resistant cell xenograft tumor model and the mechanism underlying reversal of paclitaxel resistance

Purpose: To determine the effect of paclitaxel octreotide conjugate (POC) on human ovarian paclitaxelresistant cell xenograft tumor model and the mechanism underlying reversal of paclitaxel resistance.
 Methods: Forty female BALB/c-nu/nu mice were subcutaneously inoculated with 106 paclitaxel-resistant cells (a2780/taxol) per mouse during the logarithmic growth phase of ovarian cancer. They were randomly divided into four groups (control, octreotide, paclitaxel and POC). Immunohistochemical streptavidin-peroxidase (SP) method was used to determine expression of nuclear proliferation antigen (PCNA) while TUNEL method was used to assess apoptosis of human ovarian cancer metastasis. Realtime polymerase chain reaction (PCR) was used to assay mRNA expression levels of somatostatin receptor 2 (SSTR2), multidrug-resistant gene (MDR1), vascular endothelial growth factor (VEGF), matrix metalloproteinase-9 (MMP-9), and acetylated tubulin (α-tubulin and β-III-tubulin), while the corresponding protein expressions were assayed using western blotting.
 Results: Immunohistochemical SP showed significantly lower PCNA levels in octreotide, paclitaxel and POC groups than in control mice, but that of POC mice was significantly reduced, relative to those of octreotide and paclitaxel groups (p < 0.05). There were significantly higher expression levels of SSTR2 mRNA and protein in octreotide, paclitaxel and POC groups than in control mice, but they were significantly higher in POC group than in octreotide and paclitaxel groups (p < 0.05). The mRNA and protein expressions of other factors in POC mice were significantly lower than those in both octreotide and paclitaxel groups (p < 0.05).
 Conclusion: Paclitaxel-octreotide conjugate effectively inhibits the growth of a2780/taxol xenografts in nude mice, induces tumor cell apoptosis, and suppresses tumor cell growth via mechanism involving enhancement of SSTR2 expression, and decreases in levels of acetylated tubulin, matrix metalloproteinase-9, and vascular endothelial growth factor.

Correlations of special AT-rich sequence binding protein 2 and chitinase-3-like protein-1 with sensitivity to paclitaxel chemotherapy for gastric cancer

Abstract Background: Our objective was to examine the associations between special AT-rich sequence binding protein 2 (SATB2) and chitinase-3-like protein-1 (CHI3L1) and the responsiveness to paclitaxel treatment in individuals with gastric cancer. Methods: From March 2018 to October 2020, our hospital collected gastric cancer samples along with adjacent gastric mucosal tissues located more than 5 cm away from the cancerous margin. These samples were obtained from 90 patients who underwent chemotherapy regimens containing paclitaxel. To assess the rates of positive expression of CHI3L1 and SATB2 in gastric cancer and adjacent tissues, the immunohistochemical streptavidin-peroxidase (SP) technique was utilized. Results: The positive expression rate of CHI3L1 was higher in gastric cancer tissues compared to adjacent tissues, while the positive expression rate of SATB2 was lower (P<0.05). Risk factors that influenced the positive expression of CHI3L1 in gastric cancer tissues included the level of differentiation, tumor-node-metastasis (TNM) stage, and the presence of lymph node metastasis (OR>1, P<0.05). Additionally, the positive expression of SATB2 was also affected by TNM stage and lymph node metastasis, which were identified as risk factors (OR>1, P<0.05). In gastric cancer tissues, there was a negative correlation observed between the expressions of CHI3L1 and SATB2 (r<0, P<0.05). According to the analysis results of Kendall’s tau-b (K), it was found that the presence of CHI3L1 had an inverse relationship with the responsiveness to paclitaxel-based chemotherapy in gastric cancer (r=-0.498, P=0.000), while SATB2 exhibited a positive correlation with the sensitivity (r=0.513, P=0.000). During the 3-year follow-up after chemotherapy, the survival rate was 55.55% (50/90). Conclusions: The findings indicate a strong correlation between SATB2 and CHI3L1 with the TNM stage, lymph node metastasis, response to paclitaxel-based chemotherapy, and the overall survival rate of individuals.

Ozone-Induced Lung and Bronchial Injury: A Mouse Model Study

Ozone pollution is a prominent public health issue, but there are few studies on the effect of ozone on the ultrastructure of respiratory system; we conducted this research. Exposed to 1.1 ppm O3 4 h per day, the mice lungs and bronchi were taken on the 15th or 30th day. The sections stained with HE and immunohistochemical streptavidin–peroxidase methods for NQO1, Nrf2, and Keap1 were observed and measured under the optical microscope. TEM was used for ultrastructure observation. The animals’ serums were detected for CRP and IL-6 levels. The HE-stained sections showed no obvious micromorphological changes in the O3 exposure, but the NQO1 average optical density was higher than the control on the 15th day (p < 0.05). The ultrastructural changes were found in the O3 exposure group, such as bulges and vacuoles in type I alveolar cells, the increased evacuation of substance from lamellar bodies in the type II alveolar cells, the increased space around the goblet nucleus, binuclear goblet, and columnar cells. CRP and IL-6 levels increased compared with the control (p < 0.05). Although inhaling 1.1 ppm O3 had no significant effect on the micromorphology of the mice lungs and bronchi, it did affect the ultrastructure with oxidative stress and inflammatory responses.

EGF/EGFR Promotes Salivary Adenoid Cystic Carcinoma Cell Malignant Neural Invasion via Activation of PI3K/AKT and MEK/ERK Signaling.

Salivary adenoid cystic carcinoma (SACC) is one of the most common malignant cancers of the salivary gland, and 32.4-72.0% of SACC cases exhibit neural invasion (NI); however, the molecular mechanism underlying the high invasion potential of SACC remains unclear. The present study investigated the role of epidermal growth factor receptor (EGFR) in the AKT inhibition- or mitogen-activated protein kinase kinase (MEK)-induced NI and epithelialmesenchymal transition (EMT) in SACC cells using EGFR, PI3K, and MEK inhibitors. SACC-83 cell viability was assessed using an MTT assay, and a wound healing assay was performed to evaluate cell migration. Immunohistochemical staining with streptavidin peroxidase was used to detect the positive expression rate of EMT, AKT, phosphorylated (p)-AKT, ERK, and p-ERK proteins. The impact of EGFR, PI3K, and MEK inhibitors on tumor growth and NI was examined in a xenograft model in nude mice. EGF and EGFR are effective in increasing cell viability, migration, and invasion. SACC metastasis is affected by the PI3K/AKT and MEK/ERK pathways, both of which are initiated by EGF/EGFR. The EMT and NI are regulated by the EGF/EGFR, PI3K/AKT, and MEK/ERK pathways. The present findings demonstrate the importance of suppressed EGFR/AKT/MEK signaling in NI in SACC by neural-tumor co-culture in vitro. Furthermore, our preclinical experiment provides solid evidence that injection of EGFR, PI3K, and MEK inhibitors suppressed the tumor growth and NI of SACC cells in nude mice. It was identified that inhibitors of EGFR, PI3K/AKT or MEK/ERK suppressed the proliferation, migration, and NI of SACC-83 cells via downregulation of the PI3K/AKT or MEK/ERK pathways. It was also demonstrated that inhibition of EGFR abolishes EMT in SACC by inhibiting the signaling of PI3K/AKT and MEK/ERK. The present results suggest the potential effectiveness of targeting multiple oncogenes associated with downstream pathways of EGF/EGFR, as well as potential therapeutic targets to limit NI in SACC by PI3K/AKT or MEK/ERK inhibition.

Immunohistochemical Study of Cancer Stem Cell marker, Tight Junction Protein, and Lymphatic Density in Malignant Salivary Gland Tumors

Background: CD117/c-kit, is a powerful stem cell marker for malignant salivary gland tumors in which dysregulation of c-kit is closely associated with impairment of cell adhesion molecules and cancer metastasis.
 Objective: The main purpose of this work is to evaluate the immunohistochemical expression of c-kit, and claudin-1 and measure the density of lymph vessels (LVD) in common malignant salivary gland tumors by using podoplanin (D2-40) antibody.
 Materials and Methods: Immunohistochemical staining with streptavidin peroxidase was used to analysis the expression of c-kit, claudin-1 and stained podoplanin (D2-40) lymphatic vessels on fifty archival paraffin blocks of malignant salivary gland tumor (MSGTs) cases included 20 cases of AdCC, 11 cases of MEC, 10 cases of CXPA, 6 cases of AcCC, and 3 cases of PAC.
 Results: The immunopositivity of c-kit (CD117) was detected in 44/50 (88%) of studied cases, whereas, claudin-1 protein was observed in 35 (70%) of our specimens of MSGTs. Count down of stained lymph vessels between examined cases was, MEC on the top, followed by CXPA, AdCC, PAC and AcCC. A direct correlation was observed between c-kit and lymphatic density, on the other hand, the inverse correlation was found d between c-kit and cld-1, as well as, between cld-1 and lymphatic density
 Conclusion: Up regulation of cancer stem cell marker c-kit (CD117) expression is associated with decrease of tight junction protein cld-1 and increase the density of stained lymphatic vessels by podoplanin (D2-40) antibody which confirms the using of c-kit inhibitor to improve treatment strategy of malignant salivary gland tumors.

KDF1 Promoted Proliferation and Metastasis of Epithelial Ovarian Cancer via Wnt/Beta-Catenin Pathway: TCGA-Based Data Mining and Experimental Validation.

Objectives: It has been reported that keratinocyte differentiation factor 1 (KDF1) was related to proliferation, differentiation, and cell cycle. However, the role of KDF1 has not been reported in ovarian cancer. The present study investigated the function and the potential mechanism of KDF1 in ovarian cancer. Methods: We evaluated the prognostic value in ovarian cancer based on data from the Cancer Genome Atlas (TCGA) database. The Kruskal–Wallis test, Wilcoxon signed-rank test, and logistic regression were used to evaluate the relationship between KDF1 expression and clinicopathologic features. The Cox regression and the Kaplan–Meier method were adopted to evaluate prognosis-related factors. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) gene enrichment analysis, and Gene Set Enrichment Analysis (GSEA) were performed to identify the key biological process related to KDF1. Then the expression of KDF1 in ovarian cancer tissues was validated by streptavidin–peroxidase (SP) immunohistochemistry. The proliferation and invasion ability of KDF1 were determined by EdU and Transwell assay, respectively, with KDF1 gene silencing and overexpression. The mRNA expression of KDF1 was determined by qPCR. The protein expression of KDF1 was determined using the Western blot. Methods: By performing differential expression analysis on the ovarian cancer data of the TCGA database, it was found that KDF1 is highly expressed in ovarian cancer patients and associated with poorer overall survival (OS) and progression-free survival (PFS) of ovarian cancer patients. The highly expressed KDF1 may reduce cell adhesion according to GO, KEGG, and GSEA results. After analysis combining the relevant clinical features, we found that the high expression of KDF1 is an independent prognostic factor of ovarian cancer and associated with platinum resistance and tumor metastasis in ovarian cancer. At the same time, the BioGRID database showed that there might be protein–protein interaction between KDF1 and E-cadherin. Then we further validated that the high expression of KDF1 had a close correlation with the stage and grade of ovarian cancer in ovarian cancer tissue chips. Silencing KDF1 inhibited the proliferation and invasion ability of SKOV3 cells. By contrast, ectopic expression of KDF1 promoted the proliferation and invasion ability of A2780 cells. We also found that KDF1 can interact with E-cadherin and regulate the expression of Wnt5A and β-catenin, hence activating Wnt/β-catenin pathway via in vitro and vivo experiments. Conclusions: Based on the bioinformatics analysis, in vitro experiments, and an in vivo study, it is indicated that KDF1 played an important role in ovarian cancer progression and might be a therapeutic target for patients with ovarian cancer.

MORPHOLOGICAL AND IMMUNOHISTOCHEMICAL FEATURES OF PLACENTAL DAMAGE DUE TO THE INCORPORATION OF 137Cs.

to investigate the morphological and immunohistochemical features of placental damage due to theincorporation of 137Cs depending on the scenario of pregnancy completion. The study material consisted of placentas from 60 women with reproductive losses inanamnesis and signs of termination of the current pregnancy (first group) and placental samples from 30 women with an uncomplicated gestation and an unencumbered anamnesis (control group). The detailed study required the distribution of placental samples from the first group into subgroups. Subgroup 1a included 38 placentas from women who gave birth at 37-40 weeks, despite signs of termination of the current pregnancy. Subgroup 1b - placentas of 13 women who gave birth at a gestation period of 28-36 weeks + 6 days. Subgroup 1c - 9 placental samples from women who gave birth at a gestation period of 22-27 weeks + 6 days. The volumetric activity of the 137Cs in the placentas was measured using β-spectrometer. The histology of the placenta was studied using a standard technique. The following expressions were studied in placenta: CD31 / PECAM-1, CD45 / T200 / LCA, CD56 / NCAM-1, CEA / CD66e Ab-2, Vimentin, using indirect streptavidin peroxidase detection method. Placentas accumulate 137Cs. The different volumetric activity of the isotope correlates with scenarios of pregnancy. Due to the action of incorporated 137Cs with a specific mass of more than 1.1 Bq/kg, placental dysfunction develops. The consequences of placental dysfunction depend on the volumetric activity of the 137Cs and the preservation of adaptive and compensatory reactions in the placenta. Morphological and immunohistochemical features of placental damage to incorporated 137Cs were established, depending on the scenario of completion of pregnancy. A marker of unfavorable completion of pregnancy is the expression of a carcinoembryonic antigen (CEA) in the placenta. Premature termination of pregnancy (PTP) is a multifactorial pathology associated with pathological changes in immune and neuroendocrine regulation and hereditary, infectious, and environmental factors that disrupt the adaptation mechanisms in the mother-placenta-fetus system. Intraplacental irradiation of 137Cs is one of the factors in the multifactorial nature of reproductive losses. As a result of intraplacental irradiation of 137Cs, the architecture of the placenta is disturbed, the activity of pro-inflammatory cytokines CD45 and CD56 increases, and the coagulation cascade is activated. Extreme effects depend on the volumetric activity of the isotope incorporated in the placenta and the organ's compensatory capacity. Accumulation of up to 1.0 Bq/kg 137Cs does not affect the course of gestation. Internal irradiation with an activity of 4.5-10.4 Bq/kg 137Cs triggers late preterm labor. The nature of the damages corresponds to the category of «lesion of the maternal stroma» of the placenta. The volumetric activity of 137Cs over 10.4 Bq/kg is a probable cause of early preterm labor and antenatal fetal death. At the same time, the maternal and fetal structures of the placenta suffer damage. Expression of vimentin is a marker of placental destruction due to internal irradiation of 137Cs with a specific gravity of more than 4.5 Bq/kg. Expression of CEA in the structures of the placenta of women with PTP is a unique find and marker of premature birth and antenatal fetal death with intraplacental irradiation of 137Cs with an activity of more than 4.5 Bq/kg.

Mechanism of improving skin barrier function in a mouse model of atopic dermatitis using Mahuang Lianqiao Chixiaodou decoction and its disassembled formula

To reveal the mechanism of Mahuang Lianqiao Chixiaodou decoction and its disassembled formula for improving the skin barrier function in a mouse model of atopic dermatitis (AD). Sixty specific-pathogen free male BALB/c mice were randomly divided into the control group, model group, whole formula group (WF), exterior-releasing formula group (ERF), interior-clearing formula group (ICF), and positive control group (PC). A mouse model of AD was established using the semi-antigen 2,4-dinitrofluorobenzene induction method. The lesion scores, transepidermal water loss and pH, and skin histopathology of mice in each group were observed. The expressions of filaggrin, loricrin, and involucrin were detected by the streptavidin peroxidase immunohistochemical method and western blotting, and their mRNA expressions were detected by quantitative polymerase chain reaction. Mice in the WF, ERF, ICF, and PC groups showed reduced skin lesion performance, improved histopathology, decreased skin lesion score, transepidermal water loss and pH, and upregulated expressions of proteins including filaggrin, loricrin, and involucrin, and their mRNAs. The most obvious regulatory effect was observed in the WF group, followed by the ICF, ERF, and PC groups, accordingly. Mahuang Lianqiao Chixiaodou decoction and its disassembled formula can improve the skin barrier function in a mouse model of AD by upregulating filaggrin, loricrin, and involucrin, and their mRNA expressions, and the most optimal effect was noted in the WF group, followed by the ICF and ERF groups, which suggests that the effect of clearing heat and resolving dampness in improving the skin barrier function of AD is more obvious and is one of the key treatments for AD.

Electrochemical Immunosensing of ST2: A Checkpoint Target in Cancer Diseases

A magnetic beads (MB)-involved amperometric immunosensor for the determination of ST2, a member of the IL1 receptor family, is reported in this work. The method utilizes a sandwich immunoassay and disposable screen-printed carbon electrodes (SPCEs). Magnetic immunoconjugates built on the surface of carboxylic acid-microsized magnetic particles (HOOC-MBs) were used to selectively capture ST2. A biotinylated secondary antibody further conjugated with a streptavidin peroxidase conjugate (Strep-HRP) was used to accomplish the sandwiching of the target protein. The immune platform exhibits great selectivity and a low limit of detection (39.6 pg mL−1) for ST2, allowing the determination of soluble ST2 (sST2) in plasma samples from healthy individuals and patients diagnosed with pancreatic ductal adenocarcinoma (PDAC) in only 45 min once the immunoconjugates have been prepared. The good correlation of the obtained results with those provided by an ELISA kit performed using the same immunoreagents demonstrates the potential of the developed strategy for early diagnosis and/or prognosis of the fatal PDAC disease.

The significance of detecting glucose transporter 1 and calretinin in serous effusions to differentiate between carcinoma cells and reactive mesothelial cells.

The cytologic evaluation of serous effusions to distinguish malignant cells from reactive mesothelial cells (RMCs)was an enormous challenge. The purpose of this study was to investigate the diagnostic value of glucose transporter 1 (GLUT1) and calretinin (CR) in serous effusions of patients with malignant and in order to significantly ameliorate the diagnostic accuracy. The expressions of GLUT1 and CR were measured by streptavidin-peroxidase (S-P) immunocytochemical technique in serous effusions of 183 patients with malignant and in 95 patients with benign diseases. The positive ratio of GLUT1 was 91.8% (168/183) in serous effusions from patients with malignant and 5.3% (5/95) in benign diseases, they had a significant difference (P < .01). CR was expressed 89.5% (85/95) in benign diseases and 6.6% (12/183) in malignant, it also showed an important difference (P < 0.01). The combination of GLUT1 + CR revealed the best efficiency: the sensitivity and specificity were 100% and 98.9%, respectively. Immunocytochemical staining for GLUT1 and CR may be used as a complementary tool for the detection of malignant effusions and help to make a distinction between cancer cells and RMCs. The combination of GLUT1 and CR with immunocytochemistry stained can be achieved a higher diagnostic performance.

Endoglin Expression as a Measure of Microvessel Density in Oral Head and Neck Cancer by Immunohistochemical Study.

Abstract Background and Aim: Tumor growth is limited by the balance between the need for oxygen and nutrients and diffusion of these substances from vessels around tumors. Therefore, angiogenesis, development of new blood vessels from previous blood vessels, is necessary for tumor growth and metastasis. This study was carried out with the aim of evaluating the angiogenesis in oral cancer by assessing microvessel density (MVD) using Endoglin (CD105). This work also correlated the expressions of marker with histopathological grades of oral squamous cell carcinoma. Material and Methods: Immunohistochemical staining with Streptavidin peroxidase was used to analyze the CD105 expression in 25 cases of oral squamous cell carcinoma. Results: All studied cases showed positive of CD105 immunoreactivity. The study revealed a highly significant increase of CD105-MVD in poorly differentiated OSSC than well differentiated OSCC. Conclusion: Increased expression of CD 105 in high grade tumors suggests that Endoglin is likely to contribute significantly to resistance in response to therapy.

Expression of CD44 and IL-10 in normotensive and preeclamptic placental tissue.

We aimed to demonstrate cell-cell adhesion and apoptotic changes in preeclamptic placentas by examining the expression of CD44 and IL-10. Placenta samples of 15 preeclamptic and 15 healthy 35-38th week-pregnant women were involved in the study. Tissue samples were taken only from the maternal side of the placenta and fixed in 10% formaldehyde, then blocked in paraffin wax and 5 μm-thick sections were cut and stained with Masson Trichrome. Antigen retrieval was performed for sections, incubated with CD44 antibody and anti-IL-10 antibody. After the application of streptavidin peroxidase followed by AEC chromogen solution, sections were counterstained with Mayer hematoxylin. In the preeclampsia group, increased CD44 positive expression was observed in maternal decidua cells and fibroblast cells close to root villi. CD44 was positively expressed in muscle cells around the blood vessels, mucosal connective tissue areas, syncytial nodes, and syncytial bridges. In the preeclampsia group, significant increased IL-10 expression was seen in subendothelial layers of the medium-sized vessels in the maternal region. IL-10 was also positively expressed in decidua cells outside the vessels, and inflamed connective tissue areas, chorionic villus cells with intense inflammation in intervillous spaces. CD44 was found to be an essential molecule in the regulation of vascular permeability, inflammatory response, activation of the cells, cell-to-cell interaction, and the signaling pathways to which they are associated. Since IL-10 regulates appropriate pregnancy outcomes and contributes to the balance of anti-inflammatory signals via both paracrine and autocrine regulators of trophoblast activity, we proposed that it might be a key to elucidate the etiology of preeclampsia with CD44 receptor.

The Effect of Metformin on the Clinicopathological Features of Breast Cancer With Type 2 Diabetes.

BackgroundThe present study aimed to review the use of hypoglycemic drugs and clinicopathological data in breast cancer patients with type 2 diabetes mellitus (T2DM), and to investigate the effect of metformin on the clinicopathological features of breast cancer in patient with T2DM.MethodsEighty-nine patients with breast cancer hospitalized in the Second Affiliated Hospital of Xi’an Jiaotong University from January 2012 to December 2014 were included. Thirty-three patients were on metformin (metformin group) and 56 patients were on control group. Streptavidin-peroxidase (SP) method was used to quantify protein expression of molecular markers (estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor-2 (HER-2)), molecular markers of proliferation (Ki-67 and epidermal growth factor receptor (EGFR)) and epithelial-mesenchymal transition (EMT) molecular markers (matrix metalloproteinase-2 (MMP-2), E-cadherin and downstream N-cadherin). Fluorescence in situ hybridization was used to detect HER-2 (+ and ++).ResultsThe rate of lymph node metastasis and the level of Ki-67/MMP-2 in the metformin group were significantly lower than those in the control group (P < 0.05). The ratio of luminal pattern in metformin group was higher than that in the control group (P < 0.05). However, there were no differences in the parameters of age, duration of diabetes, body mass index, tumor size, histological grade of cancer and clinical pathological features between the two groups. No significant difference was observed in the expressions of ER, PR, HER-2, EGFR, E-cadherin, N-cadherin and the recurrence rate between two groups.ConclusionsMetformin is associated with luminal breast cancer and can inhibit breast cancer invasion and metastasis in some cases. It may be associated with EMT and is beneficial to the prognosis of breast cancer.

Magnetic beads-based electrochemical immunosensing of HIF-1α, a biomarker of tumoral hypoxia

Hypoxia-inducible factor-1 alpha (HIF-1α) is a transcription factor involved in tumor growth and metastasis by regulating genes involved in response to hypoxia. This work reports the first magnetic beads (MBs)-based electrochemical immunoassay for the determination of HIF-1α. The design involves a sandwich immunoassay using MBs and amperometric detection at disposable screen-printed carbon electrodes (SPCEs). Magnetic immunoconjugates of carboxylic acid-microsized magnetic particles (HOOC-MBs) modified with a specific antibody were used to selectively capture the target protein which was sandwiched with a biotinylated secondary antibody further conjugated with a streptavidin peroxidase conjugate (Strep-HRP). The great analytical performance of the immunoassay in terms of selectivity and low limit of detection (76 pg mL−1) for HIF-1α allows the determination of the endogenous amount of this biomarker in 0.5 μg raw cancer cells lysates cultured in normoxia or induced-hypoxia with CoCl2. In addition, the developed immunosensing approach was employed to analyze HIF-1α in saliva samples with results that agree with those provided by an ELISA kit involving the same immunoreagents.

Effect of minimally invasive drilling closed drainage on the expression of HIF-1α, Claudin-5, TGF- β and Smad2/3 in patients with chronic subdural hematoma

Objective To investigate the effects of single-hole and double-hole drilling and closed drainage on the expression of Hypoxia inducible factor-1α (HIF-1α), Claudin-5, transforming growth factor-β (TGF-β) and Smad2/3 in the epidermis of patients with chronic subdural hematoma. Methods 100 patients with chronic subdural hematoma were randomly divided into single hole and double hole drainage group according to random number table. Immunohistochemical streptavidin-peroxidase (SP) was used to detect the expression of TGF- β protein, and Western blot was used to detect the expression of HIF-1α, Claudin-5 and Smad2/3 in the epihematoma of the two groups before and after operation. Results Compared with pre-operation, the expression of HIF-1α, TGF-β and Smad2/3 protein in adventitia of hematoma in both groups decreased after operation, especially in the double-hole group (P<0.05). The expression of Claudin-5 protein in adventitia of hematoma was significantly increased in both groups, especially in the double-hole group, with statistically significant difference (P<0.05). Conclusions Single-hole and double-hole drilling and sealing drainage is an effective method to treat chronic subdural hematoma. It can reduce the expression of HIF-1α, TGF-β and Smad2/3 protein in the epidermis of hematoma, and significantly increase the expression of Claudin-5, and the effect of double-hole drilling and sealing drainage is more significant. Key words: Drainage; Hematoma, subdural, chronic; Hypoxia-inducible factor 1, alpha subunit; Claudin-5; Transforming growth factor beta; Smad proteins, receptor-regulated