A novel method for simultaneously measuring boronophenylalanine uptake in brain tumor cells and number of cells using inductively coupled plasma atomic emission spectroscopy.

Boron neutron capture therapy (BNCT) combines neutron irradiation with boron compounds that are selectively uptaken by tumor cells. Boronophenylalanine (BPA) is a boron compound used to treat malignant brain tumors. The determination of boron concentration in cells is of great relevance to the field of BNCT. This study was designed to develop a novel method for simultaneously measuring the uptake of BPA by U87 and U251 cells (two brain tumor cell lines) and number of cells using inductively coupled plasma atomic emission spectroscopy (ICP-AES). The results revealed a linear correlation between phosphorus intensity and the numbers of U87 and U251 cells, with correlation coefficients (R2) of 0.9995 and 0.9994, respectively. High accuracy and reliability of phosphorus concentration standard curve were also found. Using this new method, we found that BPA had no significant effect on phosphorus concentration in either U87 or U251 cells. However, BPA increased the boron concentration in U87 and U251 cells in a concentration-dependent manner, with the boron concentration in U87 cells being higher than that in U251 cells. In both U87 and U251 cells, boron was mainly distributed in the cytoplasm and nucleus, accounting for 85% and 13% of the total boron uptake by U87 cells and 86% and 11% of the total boron uptake by U251 cells, respectively. In the U87 and U251 cell-derived xenograft (CDX) animal model, tumor exhibited higher boron concentration values than blood, heart, liver, lung, and brain, with a tumor/blood ratio of 2.87 for U87 cells and 3.11 for U251 cells, respectively. These results suggest that the phosphorus concentration in U87 and U251 cells can represent the number of cells and BPA is easily uptaken by tumor cells as well as in tumor tissue.

Supplement of exogenous inorganic pyrophosphate inhibits atheromatous calcification in Apolipoprotein E knockout mice.

Inorganic pyrophosphate (PPi) is the endogenous inhibitor for vascular calcification (VC). The present study was to investigate the effects of adenosine disodium triphosphate (ADTP) and alendronate sodium (AL), two exogenous PPi sources, on the atheromatous calcification (AC) in Apolipoprotein E knockout (ApoE KO) mice. ApoE KO mice were randomly divided into five groups: ApoE KO group, ApoE KO + ADTP (Low) group, ApoE KO + ADTP (High) group, ApoE KO + AL (Low) group and ApoE KO + AL (High) group. The mice in ApoE KO + ADTP (Low) group and ApoE KO + ADTP (High) group were intraperitoneally injected with ADTP with dose of 0.5 and 1.0 mg/kg/day for 2 months respectively. The mice in ApoE KO + AL (Low) group and ApoE KO + AL (High) group were intraperitoneally injected with AL with dose of 0.6 and 1.2 mg/kg/day for 2 months respectively. The age matched C57 mice were used as control group. All ApoE KO and C57 mice were fed with normal chow throughout the experiment. The calcification was evaluated using von Kossa method. The contents of PPi, triglyceride (TG), total cholesterol (TC), high density lipoprotein (HDL) and low density lipoprotein (LDL), tumor necrosis factor α (TNF-α), interleukin-6 (IL-6), interferon-γ (IFN-γ) and interleukin-10 (IL-10) as well as the activity of alkaline phosphatase (ALP) in serum were measured. The results showed that compared with C57 mice, ApoE KO mice developed severe AC accompanied with high levels of TC, TG, LDL, IL-6, TNF-α and IFN-γ in serum and with low levels of PPi and IL-10 in serum. Both ADTP and AL dose-dependently reduced the AC in ApoE KO mice compared with that of ApoE mice, without affecting the contents of lipid profiles. In addition, ADTP and AL increased the contents of PPi and IL-10 while decreased the contents of TNF-α, IL-6 and IFN-γ in serum of ApoE KO mice, having no affection on ALP activity. The results suggested that ADTP and AL reduced AC in ApoE KO mice by increasing the PPi level and regulating the inflammation.

Organoids: Construction and Application in Gastric Cancer.

Gastric organoids are biological models constructed in vitro using stem cell culture and 3D cell culture techniques, which are the latest research hotspots. The proliferation of stem cells in vitro is the key to gastric organoid models, making the cell subsets within the models more similar to in vivo tissues. Meanwhile, the 3D culture technology also provides a more suitable microenvironment for the cells. Therefore, the gastric organoid models can largely restore the growth condition of cells in terms of morphology and function in vivo. As the most classic organoid models, patient-derived organoids use the patient's own tissues for in vitro culture. This kind of model is responsive to the 'disease information' of a specific patient and has great effect on evaluating the strategies of individualized treatment. Herein, we review the current literature on the establishment of organoid cultures, and also explore organoid translational applications.

Evaluation of Pharmacokinetics of Boronophenylalanine and Its Uptakes in Gastric Cancer.

Boron neutron capture therapy (BNCT), a cellular-level particle radiation therapy, combines boron compounds selectively delivered to tumor tissue with neutron irradiation. Boronophenylalanine (BPA) is a boron compound widely used in malignant melanoma, malignant brain tumors, and recurrent head and neck cancer. However, neither basic nor clinical research was reported for the treatment of gastric cancer using BPA. Selective distribution of boron in tumors rather than that in blood or normal tissue prior to neutron irradiation is required for the successful treatment of BNCT. This study evaluated the pharmacokinetics and safety of 10B-labeled BPA (10B-BPA, abbreviated as BPA) and its uptakes in gastric cancer. Pharmacokinetics and safety were evaluated in Sprague-Dawley (SD) rats intravenously injected with BPA. The uptakes of boron in gastric cancer cell line MKN45 and in cell-derived xenografts (CDX) and patient-derived xenografts (PDX) animal models were measured. The results showed that the boron concentration in the blood of rats decreased fast in the first 30 min followed by a steady decrease following the observation time, having a half-life of 44.11 ± 8.90 min and an AUC-last of 815.05 ± 62.09 min×µg/ml. The distribution of boron in different tissues (heart, liver, lung, stomach, and small intestine) of rats revealed a similar pattern in blood except for that in the brain, kidney, and bladder. In MKN45 cells, boron concentration increased in a time- and concentration-dependent manner. In both CDX and PDX animal models, the boron is preferentially distributed in tumor tissue rather than in blood or normal tissues. In addition, BPA had no significant adverse effects in rats. Taken together, the results suggested that BPA revealed a fast decrease in boron concentration in rats and is more likely to distribute in tumor cells and tissue.

Calpain inhibitor prevents atherosclerosis in apolipoprotein E knockout mice by regulating mRNA expression of genes related to cholesterol uptake and efflux.

PURPOSE: We previously reported that a calpain inhibitor (CAI) prevents the development of atherosclerosis in rats. This study aimed to investigate the effects of CAI (1 mg/kg) on atherosclerosis in apolipoprotein E knockout (ApoE KO) mice that were fed a high-fat diet (HFD) and explore the underlying mechanism by analyzing the expression of genes related to the uptake and efflux of cholesterol. METHODS: Atherosclerotic plaques were evaluated. The activity of calpain in the aorta and that of superoxide dismutase (SOD) in the serum were assessed. Lipid profiles in the serum and liver were examined. Serum oxidized low-density lipoprotein (oxLDL), malondialdehyde (MDA), tumor necrosis factor (TNF-α), and interleukin-6 (IL-6) levels were measured. The mRNA expressions of CD68, TNF-α, IL-6, CD36, scavenger receptor (SR-A), peroxisome proliferator-activated receptor gamma (PPAR-γ), liver-x-receptor alpha (LXR-α), and ATP-binding cassette transporter class A1 (ABCA1) in the aorta and peritoneal macrophages were also evaluated. RESULTS: CAI reduced calpain activity in the aorta. CAI also impeded atherosclerotic lesion formation and mRNA expression of CD68 in the aorta and peritoneal macrophages of ApoE KO mice compared with those of mice receiving HFD. However, CAI had no effect on body weight and lipid levels in both the serum and liver. CAI significantly decreased MDA, oxLDL, TNF-α, and IL-6 levels and increased SOD activity in the serum. Moreover, CAI significantly inhibited the mRNA expression of TNF-α and IL-6 genes in the aorta and peritoneal macrophages. In addition, CAI significantly downregulated the mRNA expression of scavenger receptors CD36 and SR-A and upregulated the expression of genes involved in the cholesterol efflux pathway, i.e., PPAR-γ, LXR-α, and ABCA1 in the aorta and peritoneal macrophages. CONCLUSIONS: CAI inhibited the development of atherosclerotic lesions in ApoE KO mice, and this effect might be related to the reduction of oxidative stress and inflammation and the improvement of cholesterol intake and efflux pathways.

The Reliability and Validity of Post Stroke Depression Scale in Different Type of Post Stroke Depression Patients.

OBJECTIVE: A self-rating post stroke depression scale (PSDS) showed a good reliability and validity to assess severity of depressive symptoms among stroke patients. This study aimed to retest the psychometric properties of PSDS in different types of post-stroke depression (PSD). MATERIALS AND METHODS: A total of 170 stroke patients were recruited in the study. 82 and 25 patients were respectively diagnosed as PSD symptoms disorder (PSDSD) and PSD disorder (PSDD) patients according to their respective diagnostic criteria. The PSDS and the 9-item Patient Health Questionnaire (PHQ-9) were used to assess the severity of depression. Cronbach α, Spearman rank coefficient and independent sample t-test were conducted to examine reliability, internal consistency and discriminate validity. Then the receiver operating characteristic curve and Youden index were used to performance evaluation and cut-off value respectively in different subtypes of PSD patients. RESULTS: The Cronbach α of PSDS was 0.857, indicting a good reliability. The Spearman correlation coefficient between PSDS and PHQ-9 was 0.942 (P<0.001). The discriminate validity displayed significant difference between PSDSD as well as PSDD and no depression patients (all P<0.001). 5/24 and 10/24 were the cut-off value for PSDSD and PSDD patients. CONCLUSIONS: PSDS is a useful screen tool with an acceptable psychometric properties for estimation of different subtypes of PSD patients.

New opinion on the subtypes of poststroke depression in Chinese stroke survivors.

AIM: Poststroke depression (PSD) is the most common complication of stroke. However, some stroke survivors with depression cannot meet the diagnostic criteria of PSD. The aim of this study was to propose the new conception of stroke patients with depression and then make them to receive reasonable diagnosis and treatment. METHODS: We first put forward the opinion that the general PSD should consist of PSD disorder (PSDD) and PSD symptoms (PSDS) according to the Diagnostic and Statistical Manual of Mental Disorder - Fifth Edition (DSM-5) and ZhongDa diagnostic criteria - first edition (ZD-1), respectively. The ZD-1 was established based on the suggestions of 65 Chinese chief doctors considering that the symptoms of PSDS might be different from those of PSDD and the duration of DSM-5 was too strict. Then, 166 stroke inpatients were recruited, and the study was conducted using the diagnosis and classification of PSD to verify the new concept. RESULTS: A total of 24 (14.46%) and 80 (48.19%) stroke patients were diagnosed with PSDD and PSDS, respectively, according to individual diagnosis criteria. Moreover, patients meeting the diagnostic criteria of PSDD should satisfy the criteria of PSDS first. The distribution frequencies of depressive symptoms were different, which suggested that there might be discrepant depressive symptoms between PSDS and PSDD. CONCLUSION: The present study proposes new opinion about the classification and diagnosis of depression in stroke survivors. The definition and criteria of PSDS are beneficial to explore phenomenological consistency and provide useful information for early recognition and appropriate interventions.

Nonadherent culture method downregulates stem cell antigen-1 expression in mouse bone marrow mesenchymal stem cells.

Mesenchymal stem cells (MSCs) are primarily isolated by their adherence to plastic and their in vitro growth characteristics. Expansion of these cells from an adherent culture is the only method to obtain a sufficient number of cells for use in clinical practice and research. However, little is known with regard to the effect of adherence to plastic on the phenotype of the cells. In the present study, bone marrow CD45-CD31-CD44- stem cell antigen (Sca)-1+ MSCs were sorted by flow cytometry and expanded in adherent cultures. The expression levels of the adhesion molecule, Sca-1, in the adherent cultures were compared with those from nonadherent cultures at different time points. The flow cytometry results indicated that the expression levels of Sca-1 decreased in the MSCs in the nonadherent cultures grown in ultra-low-adherent plates. Furthermore, the result was confirmed by quantitative polymerase chain reaction at the same time points. Therefore, the results demonstrated that the loss of plastic adherence downregulated the expression of Sca-1. The observations may provide novel insights into the molecular mechanisms underlying plastic adherent culture.

Reliability and validity of a new post-stroke depression scale in Chinese population.

BACKGROUND: Nowadays there is still a lack of effective method to evaluate post-stroke depression. To distinguish patients with and without depression after stroke reliably, this study proposes a new Post-Stroke Depression Scale (PSDS). METHODS: PSDS was developed based on various depression scales and clinician experiences. 158 stroke patients who were able to finish PSDS and Hamilton Depression Rating Scale (HDRS) were recruited. Cronbach α, Spearman rank coefficient and Kruskal-Wallis test were respectively used to examine reliability, internal consistency and discriminate validity. Then the Receiver Operating Characteristic (ROC) curve was used to determine the ability of scale and categorized scales to the range of depression. Finally, the factors of the PSDS were classified by average clustering analysis. RESULTS: The Cronbach α of PSDS was 0.797 (95% CI) indicted a good reliability. The Spearman correlation coefficient between PSDS and HDRS was 0.822 (P<0.001) showed an excellent congruent validity. The discriminate validity displayed significant difference between patients with and without depression (P<0.001). 6/24 was set to be the cut-off value by ROC analysis. Moreover, the different severity was distinguished by the value 6/24, 15/24 and 17/24. LIMITATIONS: The small sample size maybe the main limitation, the larger sample used in different fields according sex, age and side-lesion was needed to verity the results. The cut off value calculated by ROC curve maybe react the severity of the disease to some extent, but it is not absolute. CONCLUSIONS: PSDS is a valid, reliable and specific tool for evaluating post-stroke depression patients and can be conveniently utilized.

Expression of miR-100 and RBSP3 in FTC-133 cells after exposure to 131I.

BACKGROUND: Sodium iodide ((131)I) therapy for the management of differentiated thyroid cancer is based on the deposition of certain doses of ionizing radiation, which can modulate microRNA (miRNA, miR) expression. Recent studies have suggested that miR-100 is significantly differentially expressed between benign and malignant thyroid tissue samples and modulates retinoblastoma 1 serine phosphates from human chromosome 3 (RBSP3), which is involved in the regulation of cell growth and differentiation. Therefore, the authors tested the hypothesis that a potential mechanism of (131)I treatment affects miR-100, which in turn regulates RBSP3 to modulate cell proliferation in thyroid cancer in vitro. MATERIALS AND METHODS: A follicular thyroid carcinoma cell line (FTC-133) was treated with (131)I or transfected with an oligonucleotide (miR-100 mimics, inhibitor, or negative control). Real-time quantitative PCR was used to confirm the expression levels of the miR-100 and RBSP3 mRNAs. Western blot analysis was performed to detect the levels of the RBSP3 protein. The cell cycle was analyzed on a cytofluorimeter by fluorescence-activated cell sorting analysis. RESULTS: RBSP3 protein expression was detected in FTC-133 cells. (131)I treatment inhibited the expression of miR-100 in FTC cells, as assessed by real-time quantitative PCR analysis, whereas it upregulated the RBSP3 mRNA and protein. Overexpression and knockdown experiments indicated that miR-100 repressed the expression of the RBSP3 mRNA by blocking its translation. Overexpression of miR-100 led to the downregulation of the RBSP3 protein and promoted the transition of FTC cells from the G1 to the S phase, as assessed using FACS analysis. CONCLUSION: (131)I treatment inhibited the expression of miR-100, which modulated RBSP3 in FTC cells. The new mechanism of suppression of the proliferation of FTC cells by I described here might occur through the downregulation of miR-100.

Different expression of NOD2 in decidual stromal cells between normal and unexplained recurrent spontaneous abortion women during first trimester gestation.

The NOD2 gene, encoding intracellular paternal recognition receptor (PRR) also called caspase activation and recruitment domain 15 (CARD15), is mutated in Crohn's disease, an autoimmune-disorder. Unexplained recurrent spontaneous abortion (URSA) involved in complex auto-immune disorder. However, little is known about the expression of NOD2 protein at maternal-fetal interface with URSA patients. Our aim was to compare the expression levels of NOD2 in the decidual stromal cells (DSCs) from patients with normal pregnancy to those with unexplained recurrent spontaneous abortion (URSA) during first trimester pregnancy. Tissues and DSCs were collected from 12 patients with URSA and 26 patients with normal pregnancies that required abortion. DSCs in the normal pregnancy group showed significantly higher mRNA and protein levels of NOD2 than those isolated from the URSA group using real time PCR and in cell western. The appropriate expression of NOD2 in normal DSCs suggests that this protein may be required to sustain normal pregnancy.