Fluorine-18 prostate-specific membrane antigen-1007-avid indeterminate bone lesions in prostate cancer: clinical and PET/CT features to predict outcomes and prognosis.

AIM: To determine clinical and fluorine-18 prostate-specific membrane antigen-1007 (18F-PSMA-1007) integrated positron-emission tomography (PET)/computed tomography (CT) features that could be used to interpret indeterminate bone lesions (IBLs) and assess the prognosis of prostate cancer (PCa) in patients with IBLs. MATERIALS AND METHODS: Consecutive patients who underwent PSMA PET/CT were analysed retrospectively. IBLs were identified as benign or malignant based on follow-up imaging and clinical management. Lesion- and patient-based assessments were performed to define features predictive of bone lesion results and determine clinical risk. Patients' prognosis was analysed based on clinical characteristics, including prostate-specific antigen (PSA) and alkaline phosphatase (ALP), respectively. RESULTS: A total of 105 patients (mean age ± SD, 72.1 ± 8 years) were evaluated and 158 IBLs were identified. Fifty-three (33.5%), 36 (22.8%), and 69 (43.7%) IBLs were benign, malignant, and equivocal, respectively. Variables including location, maximum standard uptake value (SUVmax), and lymph node metastases (LNM) were related to the benignancy or malignancy of IBLs (p=0.046, p<0.001 and p<0.001, respectively). Regression analysis indicated that LNM, SUVmax, and location of IBLs could be predictors of lesion interpretation (p<0.001, p=0.002 and p=0.035). Patients with benign IBLs experienced the most considerable decreases in PSA and ALP levels. CONCLUSIONS: LNM, SUVmax, and location may contribute to IBL interpretation. A rapid decrease in PSA and ALP levels might suggest a better prognosis for patients with benign IBLs.

Olfactory Three-Needle Electroacupuncture Improved Synaptic Plasticity and Gut Microbiota of SAMP8 Mice by Stimulating Olfactory Nerve.

OBJECTIVE: To investigate the effects and mechanisms of olfactory three-needle (OTN) electroacupuncture (EA) stimulation of the olfactory system on cognitive dysfunction, synaptic plasticity, and the gut microbiota in senescence-accelerated mouse prone 8 (SAMP8) mice. METHODS: Thirty-six SAMP8 mice were randomly divided into the SAMP8 (P8), SAMP8+OTN (P8-OT), and SAMP8+nerve transection+OTN (P8-N-OT) groups according to a random number table (n=12 per group), and 12 accelerated senescence-resistant (SAMR1) mice were used as the control (R1) group. EA was performed at the Yintang (GV 29) and bilateral Yingxiang (LI 20) acupoints of SAMP8 mice for 4 weeks. The Morris water maze test, transmission electron microscopy, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining, Nissl staining, Golgi staining, Western blot, and 16S rRNA sequencing were performed, respectively. RESULTS: Compared with the P8 group, OTN improved the cognitive behavior of SAMP8 mice, inhibited neuronal apoptosis, increased neuronal activity, and attenuated hippocampal synaptic dysfunction (P<0.05 or P<0.01). Moreover, the expression levels of synaptic plasticity-related proteins N-methyl-D-aspartate receptor 1 (NMDAR1), NMDAR2B, synaptophysin (SYN), and postsynaptic density protein-95 (PSD95) in hippocampus were increased by OTN treatment (P<0.05 or P<0.01). Furthermore, OTN greatly enhanced the brain-derived neurotrophic factor (BDNF)/cAMP-response element binding (CREB) signaling and phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/AKT/mTOR) signaling compared with the P8 group (P<0.05 or P<0.01). However, the neuroprotective effect of OTN was attenuated by olfactory nerve truncation. Compared with the P8 group, OTN had a very limited effect on the fecal microbial structure and composition of SAMP8 mice, while specifically increased the genera Oscillospira and Sutterella (P<0.05). Interestingly, the P8-N-OT group showed an abnormal fecal microbiota with higher microbial α-diversity, Firmicutes/Bacteroidetes ratio and pathogenic bacteria (P<0.05 or P<0.01). CONCLUSIONS: OTN improved cognitive deficits and hippocampal synaptic plasticity by stimulating the olfactory nerve and activating the BDNF/CREB and PI3K/AKT/mTOR signaling pathways. Although the gut microbiota was not the main therapeutic target of OTN for Alzheimer's disease, the olfactory nerve was essential to maintain the homeostasis of gut microbiota.

Resolvin E1 heals injured cardiomyocytes: Therapeutic implications and H-FABP as a readout for cardiovascular disease & systemic inflammation.

The purpose of this study is to investigate heart-fatty acid binding protein (H-FABP) leakage from cardiomyocytes as a quantitative measure of cell membrane damage and to test healing by Resolvin E1 (RVE1) as a potential therapeutic for patients with inflammatory diseases (cardiovascular disease and comorbidities) with high morbidity and mortality. Our quantitative ELISA assays demonstrated H-FABP as a sensitive and reliable biomarker for measuring cardiomyocyte damage induced by lipopolysaccharide (LPS) and healing by RvE1, a specialized pro-resolving mediator (SPM) derived from the Omega-3 fatty acid, eicosapentaenoic acid (EPA), a dietary nutrient that balances inflammation to restore homeostasis. RvE1 reduced leakage of H-FABP by up to 86%, which supports our hypothesis that inflammation as a mechanism of injury can be targeted for therapy. H-FABP as a blood biomarker was tested in 40 patients admitted to Boston Medical Center for respiratory distress, (20 patients with and 20 patients without COVID infection). High levels of H-FABP correlated with clinically diagnosed CVD, diabetes, and end-stage renal disease (ESRD) in both patient groups. The level of H-FABP indicates not only CVD damage but is a valuable measure for patients with increased inflammation disease comorbidities.

[Characteristics of serum autoantibodies in patients with lupus nephritis and tubulointerstitial damage].

OBJECTIVE: To observe the tubulointerstitial damage (TID) in lupus nephritis (LN) and investigate the relationship between autoantibodies and TID in lupus nephritis. METHODS: This cross-sectional study was conducted in a comprehensive tertiary hospital in Peking University Shenzhen Hospital. From March 2012 to July 2021, LN patients who performed renal biopsy were enrolled in the study. Clinical, laboratory and pathology data were collected. We classified the patients into none-or-mild group and moderate-to-severe groups according to the severity of interstitial fibrosis (IF) /tubular atrophy (TA) or tubulointerstitial inflammation (TII). The t test, U test and Chi-square test were used for statistical analysis as appropriate. RESULTS: A total of 226 patients were included, of who 190 (84%) were female with a median age of 32 (26, 39) years. 89% (201/226) of the patients who pathologically proved to be proliferative LN by renal biopsy. The frequency of moderate-to-severe TII and moderate-to-severe IF/TA was 30% (67/226) and 34% (76/226) respectively. For autoantibodies, the patients with moderate-to-severe TII had a lower rate of positive serum anti-ribonucleoprotein (anti-RNP) antibodies than the patients with none-or-mild TII (34% vs. 51%), and moderate-to-severe IF/TA had a lower rate of positive anti-ribosomal P protein (anti-P) antibodies than patients with none-or-mild IF/TA (19% vs. 33%). For other clinical indicators, the patients with moderate-to-severe TII and moderate-to-severe IF/TA were more often combined with proliferative LN, hypertension and anemia than the patients with none-or-mild TII and none-or-mild IF/TA, respectively. The patients with moderate-to-severe TII had higher serum creatinine values and lower glomerular filtration rates than the patients with none-or-mild TII. The patients with moderate-to-severe IF/TA had higher serum creatinine values, and lower glomerular filtration rates than the patients with none-or-mild IF/TA. CONCLUSION: In patients with LN in Southern China, anti-RNP antibodies and anti-P antibodies may be potential protective factors for TII and IF/TA, respectively. More studies are needed to identify the risk factors of lupus patients with TID and investigate the correlation between autoantibodies and TID, which are critical for developing better preventive and therapeutic strategies to improve the survival rate of LN.

Inline amplification of mid-infrared intrapulse difference frequency generation.

We demonstrate an ultrafast mid-infrared source architecture that implements both intrapulse difference frequency generation (iDFG) and further optical parametric amplification (OPA), in an all-inline configuration. The source is driven by a nonlinearly compressed high-energy Yb-doped-fiber amplifier delivering 7.4 fs pulses at a central wavelength of 1030 nm, at a repetition rate of 250 kHz. It delivers 1 µJ, 73 fs pulses at a central wavelength of 8 µm, tunable over more than one octave. By enrolling all the pump photons in the iDFG process and recycling the long wavelength pump photons amplified in the iDFG in the subsequent OPA, we obtain an unprecedented overall optical efficiency of 2%. These performances, combining high energy and repetition rate in a very simple all-inline setup, make this technique ideally suited for a growing number of applications, such as high harmonic generation in solids or two-dimensional infrared spectroscopy experiments.

A resonance suppression method in platform style inertial reference unit via particle swarm optimization notch filter.

With the development of laser communication, remote sensing imaging, and other technologies, an inertial reference unit (IRU) plays an essential part in the line-of-sight (LOS) stabilization system used for acquiring, pointing, and tracking targets. The IRU provides a stable reference beam to realize accurate LOS pointing under external disturbances. Compared with the frame style IRU, the platform style IRU (PIRU) can achieve a higher bandwidth and better precision. However, mechanical resonance is introduced by a flexure hinge inevitably in the PIRU, which affects the performance of the LOS stabilization system. In this paper, an open-loop dynamic model of PIRU is established. Identification experiments are carried out with results indicating a 28.7 dB resonance peak at 27.07 Hz in the x axis and a 30.3 dB resonance peak at 26.59 Hz in the y axis. An asymmetric notch filter is used to suppress the resonance peak to achieve a higher control bandwidth. A fitness function is designed to represent the effect of resonance suppression. A particle swarm optimization algorithm is used to search for an optimal solution of the fitness function to obtain the parameters of the asymmetric notch filter. Experimental results show that the resonance peak is reduced by 97.88% and the system bandwidth reaches 159.31 Hz.

[Consistency of peripheral whole blood and venous serum procalcitonin in children: a multicenter parallel controlled study].

Objective: To explore the consistency of peripheral whole blood and venous serum procalcitonin (PCT) levels, and the value of peripheral whole blood PCT in evaluating pediatric bacterial infection. Methods: This multicenter cross-sectional parallel control study was conducted in 11 children's hospital. All the 1 898 patients older than 28 days admitted to these hospitals from March 2018 to February 2019 had their peripheral whole blood and venous serum PCT detected simultaneously with unified equipment, reagent and method. According to the venous serum PCT level, the patients were stratified to subgroups. Analysis of variance and chi-square test were used to compare the demographic characteristics among groups. And the correlation between the peripheral blood and venous serum PCT level was investigated by quantitative Pearson correlation analysis.The PCT resultes were also converted into ranked data to further test the consistency between the two sampling methods by Spearman's rank correlation test. Furthermore, the ranked data were converted into binary data to evaluate the consistency and investigate the best cut-off of peripheral blood PCT level in predicting bacterial infection. Results: A total of 1 898 valid samples were included (1 098 males, 800 females),age 27.4(12.2,56.7) months. There was a good correlation between PCT values of peripheral whole blood and venous serum (r=0.97, P<0.01). The linear regression equation was PCTvenous serum=0.135+0.929×PCTperipheral whole blood. However, when stratified to 5 levels, PCT results showed diverse and unsatisfied consistency between the two sampling methods (r=0.51-0.92, all P<0.01). But after PCT was converted to ordinal categorical variables, the stratified analysis showed that the coincidence rate of the measured values by the two sampling methods in each boundary area was 84.9%-97.1%. The dichotomous variables also showed a good consistency (coincidence rate 96.8%-99.3%, Youden index 0.82-0.89). According to the severity of disease, the serum PCT value was classified into 4 intervals（<0.5、0.5-<2.0、2.0-<10.0、≥10.0 µg/L）, and the peripheral blood PCT value also showed a good predictive value (AUC value was 0.991 2-0.997 9). The optimal cut points of peripheral whole blood PCT value 0.5、1.0、2.0、10.0 µg/L corresponding to venous serum PCT values were 0.395, 0.595, 1.175 and 3.545 µg/L, respectively. Conclusions: There is a good correlation between peripheral whole blood PCT value and the venous serum PCT value, which means that the peripheral whole blood PCT could facilitate the identification of infection and clinical severity. Besides, the sampling of peripheral whole blood is simple and easy to repeat.

Down-regulation of betatrophin enhances insulin sensitivity in type 2 diabetes mellitus through activation of the GSK-3ß/PGC-1α signaling pathway.

OBJECTIVE: The incidence of type 2 diabetes mellitus (T2DM) among children and adolescents has been rising. Accumulating evidences have noted the significant role of betatrophin in the regulation of lipid metabolism and glucose homeostasis. In our study, we tried to figure out the underlying mechanism of betatrophin in insulin resistance (IR) in type 2 diabetes mellitus (T2DM). METHODS: First, fasting serum betatrophin, fasting blood glucose (FBG), insulin, total cholesterol (TC), triglyceride (TG), and low-density lipoprotein cholesterol (LDL-C) were detected in T2DM children. The homeostasis model assessment of insulin resistance (HOMA-IR), Gutt insulin sensitivity index (ISIG) and Matsuda insulin sensitivity index (ISIM) were calculated. A T2DM-IR mouse model was induced by high-fat diet, with the expression of GSK-3ß and PGC-1α detected. Besides, HepG2 cells were induced by a high concentration of insulin to establish an IR cell model (HepG2-IR). The cell viability, glucose consumption, liver glycogen content, inflammation, and fluorescence level of GSK-3ß and PGC-1α were analyzed. RESULTS: Betatrophin was highly expressed in serum of T2DM children and was positively correlated with FBG, insulin, TC, TG, LDL-C and HOMA-IR, while negatively correlated with ISIG and ISIM. Betatrophin and GSK-3ß in the liver tissues of T2DM-IR mice were increased, while the PGC-1α expression was decreased. Betatrophin expression was negatively correlated with PGC-1α and positively correlated with GSK-3ß. Silencing of betatrophin enhanced insulin sensitivity through the activation of GSK-3ß/PGC-1α signaling pathway. In vitro experiments also found that silencing of betatrophin promoted glucose consumption and glycogen synthesis while inhibited inflammation. CONCLUSION: Our findings concluded that silencing of betatrophin could enhance insulin sensitivity and improve histopathological morphology through the activation of GSK-3ß/PGC-1α signaling pathway.

Risk factors for cerebrospinal fluid shunt infections during an outbreak: a case-control study.

BACKGROUND: There are few published reports of cerebrospinal fluid (CSF) shunt infection outbreaks. In 2017-2018, British Columbia Children's Hospital (BCCH) experienced an increase in CSF shunt infections co-incident with a move to new operating rooms and a change in shunt catheters used. AIMS: To describe how an outbreak was detected, investigations were undertaken to determine the cause, risk factors associated with CSF shunt infection during the outbreak, and changes implemented to attempt to control the outbreak. METHODS: Retrospective case-control study. Population included patients who underwent new shunt insertion or revision. Univariate logistic regression models were fitted for each of the variables. Associations with P-values <0.2 were considered of potential interest for further investigation. FINDINGS: There were six cases of CSF shunt infection and 19 controls. The causative organism was different in each case. The only risk factors that met the criteria for further investigation were being a neonate at the time of surgery [odds ratio (OR) 9.0, 95% confidence interval (CI) 0.7-125.3, P=0.10] and the presence of gastrointestinal disease (OR 3.8, 95% CI 0.5-26.2, P=0.18). No association was found with the operating room used or the surgical staff. In response to the outbreak, human traffic through the operating rooms was limited, rigid adherence to the wearing of surgical masks was enforced, and return to the previous CSF shunt catheters used was implemented. CONCLUSION: No modifiable risk factors were associated with CSF shunt infection. After implementation of surgical protocol changes, no further cases of CSF shunt infection linked to the outbreak were identified.

Discovery of novel 9-heterocyclyl substituted 9H-purines as L858R/T790M/C797S mutant EGFR tyrosine kinase inhibitors.

Targeting L858R/T790M/C797S mutant EGFR is a major challenge in the new-generation EGFR tyrosine kinase inhibitors development for conquering drug resistant NSCLC. In this study, a series of novel 9-heterocyclyl substituted 9H-purine derivatives were designed as EGFRL858 R/T790 M/C797S tyrosine kinase inhibitors. Among these compounds, D4, D9, D11 and D12 showed significantly potent anti-proliferation and EGFRL858 R/T790 M/C797S inhibition activity. In particular, the most potent compound D9 showed anti-proliferation against HCC827 and H1975 cell lines with the IC50 values of 0.00088 and 0.20 µM, respectively. And D9 inhibited the EGFRL858R/T790M/C797S with an IC50 value of 18 nM. Furtherly, D9 could significantly suppress the EGFR phosphorylation, induce the apoptosis, arrest cell cycle at G0/G1, and inhibit colony formation in HCC827 cell line by a concentration-dependent manner. Molecular docking indicated that the introduction of a cyclopropylsulfonamide group in D9 led to the formation of additional two hydrogen bonds with mutant Ser797 which played key roles in generating efficient EGFRL858 R/T790 M/C797S inhibitory activity. These findings strongly indicated that 9-heterocyclyl substituted 9H-purine derivatives were promising L858R/T790M/C797S mutant EGFR-TKIs. The introduction of extra hydrogen bond interaction with mutant Ser797 is efficient method for the design of the fourth-generation EGFR-TKIs.

LncRNA SNHG1 promotes cell proliferation in laryngeal cancer via Notch1 signaling pathway.

OBJECTIVE: We aimed at elucidating the potential function of long noncoding ribonucleic acids (lncRNAs) small nucleolar RNA host gene 1 (SNHG1) in the progression of laryngeal cancer (LC) and its underlying mechanism. PATIENTS AND METHODS: Relative level of SNHG1 in LC tissues and controls was determined by quantitative real-time polymerase chain reaction (qRT-PCR). Its expression in LC patients with different tumor stages and statues of lymph node metastasis was examined as well. Correlation between SNHG1 expression and prognosis of LC patients was evaluated by the Kaplan-Meier method. SNHG1 siRNA (si-SNHG1) was constructed for downregulation of SNHG1 expression. Potential effects of downregulated SNHG1 on viability and proliferation of LC cells were detected by cell counting kit-8 (CCK-8) and colony formation assay, respectively. After knockdown of SNHG1, relative levels of Notch1 and hairy, and enhancer of split homolog-1 (Hes1) were determined by qRT-PCR and Western blot. Regulatory effects of SNHG1/Notch1 axis on biological behaviors of LC were finally evaluated. RESULTS: SNHG1 was upregulated in LC tissues than that of controls. Besides, its level was higher in LC with T3-T4 relative to those of T1-T2. Higher abundance of SNHG1 was identified in LC patients with lymph node metastasis compared with those non-metastatic patients. Survival analysis indicated that LC patients with high-level SNHG1 had worse overall survival. Knockdown of SNHG1 in Tu212 and Hep2 cells downregulated relative levels of Notch1 and Hes1. Moreover, SNHG1 knockdown resulted in decreased viability and proliferative ability of LC cells. Notch1 overexpression could reverse the regulatory effects of SNHG1 on viability and proliferation of LC cells. CONCLUSIONS: LncRNA SNHG1 is highly expressed in LC tissues. It promotes the proliferation of LC cells by inhibiting Notch1 pathway, thereby promoting the progression of LC.

The Function of Wnt Ligands on Osteocyte and Bone Remodeling.

Bone homeostasis is continually maintained by the process of bone remodeling throughout life. Recent studies have demonstrated that Wnt signaling pathways play a fundamental role in the process of bone homeostasis and remodeling. Intracellular Wnt signaling cascades are initially triggered by a Wnt ligand-receptor complex formation. In previous studies, the blocking of Wnt ligands from different osteoblastic differentiation stages could cause defective bone development at an early stage. Osteocytes, the most abundant and long-lived type of bone cell, are a crucial orchestrator of bone remodeling. However, the role of Wnt ligands on osteocyte and bone remodeling remains unclear. In our present study, we found that, besides osteoblasts, osteocytes also express multiple Wnt ligands in the bone environment. Then, we used a Dmp1-Cre mouse line, in which there is expression in a subset of osteoblasts but mainly osteocytes, to study the function of Wnt ligands on osteocyte and bone remodeling in vivo. Furthermore, we explored the role of Wnt ligands on osteocytic mineralization ability, as well as the regulatory function of osteocytes on the process of osteoblastic differentiation and osteoclastic migration and maturity in vitro. We concluded that Wnt proteins play an important regulatory role in 1) the process of perilacunar/canalicular remodeling, as mediated by osteocytes, and 2) the balance of osteogenesis and bone resorption at the bone surface, as mediated by osteoblasts and osteoclasts, at least partly through the canonical Wnt/ß-catenin signaling pathway and the OPG/RANKL signaling pathway.

ß-Casomorphin increases fat deposition in broiler chickens by modulating expression of lipid metabolism genes.

ß-Casomorphin is an opioid-like bioactive peptide derived from ß-casein of milk that plays a crucial role in modulating animal's feed intake, growth, nutrient utilization and immunity. However, the effect of ß-casomorphin on lipid metabolism in chickens and its mechanism remain unclear. The aim of this study was to investigate the effects of ß-casomorphin on fat deposition in broiler chickens and explore its mechanism of action. A total of 120 21-day-old Arbor Acres male broilers (747.94±8.85 g) was chosen and randomly divided into four groups with six replicates of five birds per replicate. Three groups of broilers were injected with 0.1, 0.5 or 1.0 mg/kg BW of ß-casomorphin in 1 ml saline for 7 days, whereas the control group received 1 ml saline only. The results showed that subcutaneous administration of ß-casomorphin to broiler chickens increased average daily gain, average daily feed intake and fat deposition, and decreased feed : gain ratio (P<0.05). The activity of malate dehydrogenase in the pectoral muscle, liver and abdominal adipose tissue was also increased along with the concentrations of insulin, very-low-density lipoprotein and triglyceride in the plasma (P<0.05). The activity of hormone-sensitive lipase in the liver and abdominal adipose tissue and the concentration of glucagon in the plasma were decreased by injection with ß-casomorphin (P<0.05). Affymetrix gene chip analysis revealed that administering 1.0 mg/kg BW ß-casomorphin caused differential expression of 168 genes in the liver with a minimum of fourfold difference. Of those, 37 genes are directly involved in lipid metabolism with 18 up-regulated genes such as very low density lipoprotein receptor gene and fatty acid synthase gene, and 19 down-regulated genes such as lipoprotein lipase gene and low density lipoprotein receptor gene. In conclusion, ß-casomorphin increased growth performance and fat deposition of broilers. Regulation of fat deposition by ß-casomorphin appears to take place through changes in hormone secretion and enzyme activities by controlling the gene expression of lipid metabolism and feed intake, increasing fat synthesis and deposition.

Isolation and identification of pathogenic Aeromonas veronii from a dead Yangtze finless porpoise.

Diseases caused by Aeromonas veronii in freshwater fish have been widely reported, but other species such as aquatic mammals have probably been overlooked. Here, we identified one isolate of A. veronii from a Yangtze finless porpoise Neophocaena asiaeorientalis asiaeorientalis exhibiting disease and mortality, and subsequently confirmed its virulence in artificial infection of BALB/c mice. The bacterial isolate was identified as A. veronii based on physiological, biochemical, and phenotypic features, and homology of the 16S rRNA, cpn60, rpoB, dnaJ and gyrB genes. Our results expand the known host spectrum of A. veronii, which is of great importance for the etiology of porpoise, dolphin, and other cetacean diseases.

[Homocysteine induces calcium overload in neonatal rat atrial cells through activation of sodium current and CaMK⠡δ].

Objective: To investigate the effect and related mechanism of homocysteine (Hcy) on calcium overload in neonatal rat atrial cells (NRICs). Methods: NRICs were assigned to 9 groups after culture for 3 days: (1) control group; (2) Hcy group (0, 50, 100, 200, 500 µmol/L for 48 hours); (3) antioxidant group (NAC, 10 µmol/L for 24 hours); (4) Hcy+NAC group (500 µmol/L Hcy for 48 hours, then treated with 10 µmol/L NAC for 24 hours); (5) calcium/calmodulin dependent protein kinase Ⅱδ (CaMKⅡδ) inhibitor group (KN-93, 3 µmol/L KN-93 for 5 hours); (6) specific sodium current inhibitor group (ELE, 1 µmol/L ELE for 5 hours); (7) Hcy+KN-93 group (500 µmol/L Hcy for 48 hours, then treated with 3 µmol/L KN-93 for 5 hours); (8) Hcy+ELE group (500 µmol/L Hcy for 48 hours, then treated with 1 µmol/L ELE for 5 hours; (9) Hcy+KN-93+ELE group (500 µmol/L Hcy for 48 hours, then treated with 3 µmol/L KN-93 and 1 µmol/L ELE for 5 hours). Moreover, NRICs were also treated with CaMKⅡδ-siRNA lentivirus, and Nav1.5-siRNA lentivirus, negative lentivirus carrier containing green fluorescent protein (GFP) for 24 hours. The MOI values of the three groups were 10. Infection efficiency of lentivirus was determined by observing the percentage of GFP fluorescence under inverted fluorescence microscope after transfection for 24 hours, and cultured regularly with simultaneous Puro screening, then cells were grouped as Hcy+CaMKⅡδ-siRNA group, Hcy+Nav1.5-siRNA group and Hcy+negative group. The concentration of Ca(2+) in NRICs ([Ca(2+)]i) of various groups was detected through Fluo-4/AM fluorescence probe, then 2', 7'- two chlorofluorescein diacetate (DCFH-DA) was used as a probe to detect reactive oxygen species (ROS) in NRICs by flow cytometry. The malondialdehyde (MDA) was detected by the activity of superoxide dismutase (SOD) and xanthine oxidase was detected by thiobarbituric acid colorimetry. The protein and mRNA expression level of CaMKⅡδ and Nav1.5 in NRICs were detected by Western blot and quantitative real-time PCR. Results: (1) ROS, MDA and SOD were similar between NAC group and control group, ROS and MDA were significantly increased, while SOD was significantly reduced in Hcy group in a concentration-dependent manner. (2) [Ca(2+)]i: The level of [Ca(2+)]i was (155.57+7.25), (187.43+13.07), (248.98+27.22) and (307.36+15.09) nmol/L in 50, 100, 200 and 500 µmol/L Hcy groups, which was significantly higher than that in the control group ((123.18+7.24) nmol/L, P<0.01). In addition, the level of [Ca(2+)]i in Hcy+NAC group ((222.87+23.71)nmol/L) was significantly lower than that in Hcy 500 µmol/L group ((305.15+39.45) nmol/L, P<0.05), while [Ca(2+)]i level was similar between NAC group and the control group. (3) The protein expression of CaMKⅡδ and Nav1.5 was significantly upregulated in Hcy groups than in the control group. The protein expression level of CaMKⅡδ-Thr287 was significantly lower in NAC group than in Hcy 500 µmol/L group (P<0.01), however, there was no significant difference on the protein expression levels of CaMKⅡδ-Thr287 and Nav1.5 between NAC group and control group (all P>0.05). (4) The protein expression levels of CaMKⅡδ-Thr287 and the concentration of [Ca(2+)]i were significantly lower in Hcy+KN-93 group and Hcy+KN-93+ELE group than in Hcy 500 µmol/L group (P<0.05). [Ca(2+)]i concentration was significantly lower in Hcy+KN-93 group, Hcy+ELE group and KN-93+ELE+Hcy group than in Hcy 500 µmol/L group (P<0.05). (5) The mRNA and protein expression levels of CaMKⅡδ and Nav1.5 in each group infected with lentivirus: the GFP expression was ideal post lentivirus transfection for 24 hours (up to 90%), which was significantly lower in the CaMKⅡδ-siRNA group and Nav1.5-siRNA group than in the negative infection group (all P<0.05), which was similar between negative infection group and control group (P>0.05). Moreover, the mRNA and protein expression levels of CaMKⅡδ and CaMKⅡδ-Thr287 was significantly lower in Hcy+Nav1.5-siRNA group than in Hcy+negative infection group (P<0.05). The protein and mRNA levels of Nav1.5 were similar between Hcy+CaMKⅡδ-siRNA group and Hcy+negative infection group (P>0.05). Conclusions: Hcy can induce calcium overload in NRICs by increasing oxidative stress, upregulating the sodium channel protein, and activating the late sodium current and phosphorylating CaMKⅡδ.

microRNA-145-3p inhibits non-small cell lung cancer cell migration and invasion by targeting PDK1 via the mTOR signaling pathway.

The mammalian target of rapamycin (mTOR) pathway is dysregulated in more than 50% of all human malignancies and is a major target in cancer treatment. In this study, we explored the underlying mechanism involving microRNA-145-3p (miR-145-3p) in the development and progression of non-small cell lung cancer (NSCLC) by targeting PDK1 via the mTOR signaling pathway. NSCLC tissues and adjacent normal tissues were obtained from 83 NSCLC patients. miR-145-3p, PDK1, and mTOR levels were determined by quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry. Human NSCLC cell lines A549 and H1299 were transfected with miR-145-3p and siPDK1 to confirm the effect of miR-145-3p and PDK1 on NSCLC cells in vitro. Cell growth was evaluated by a CCK8 assay. Cell motility and chemotaxis analysis were determined by the scratch test and chemotaxis assay, respectively. The protein levels of PDK1 and mTOR were measured using the western blotting. Results showed lower level of miR-145-3p and higher levels of PDK1 and mTOR in NSCLC tissues compared to the adjacent normal tissues. In vitro results showed that cell growth, cell motility, and chemotaxis were all inhibited in cells transfected with miR-145-3p and those transfected with siPDK. Additionally, dual luciferase reporter gene assay helped confirmed that PDK1 is a target of miR-145. Finally, levels of PDK1, mTOR, and phosphorylated-mTOR were lower in cells transfected with miR-145-3p as well as those with siPDK1. These findings indicate that miR-145-3p may inhibit cell growth, motility, and chemotaxis in NSCLC by targeting PDK1 through suppressing the mTOR pathway.

Tacrolimus protects vascular endothelial cells from injuries caused by Ox-LDL by regulating endoplasmic reticulum stress.

OBJECTIVE: To study the protective effect of tacrolimus on endothelial cells under the action of oxidized low-density lipoprotein (Ox-LDL), and to explore the mechanism of tacrolimus in protecting endothelial cells. MATERIALS AND METHODS: Human umbilical vein endothelial cells (HUVEC) were used and they were divided into the blank group, tacrolimus group, ox-LDL injury group, and tacrolimus + ox-LDL (tacrolimus treatment) group, for treatment. The cell viability was detected via methyl thiazolyl tetrazolium (MTT) assay; the cytotoxic response was detected via lactate dehydrogenase (LDH) release assay, and the proportion of apoptotic cells was detected via flow cytometry. Moreover, the releases of superoxide dismutase (SOD), reactive oxygen species (ROS), were detected, and the expression levels of apoptosis-related molecules, and the endoplasmic reticulum stress (ERS)-related molecules were detected via polymerase chain reaction (PCR) and Western blotting. RESULTS: Ox-LDL could significantly reduce the viability of endothelial cells, increase the cytotoxicity and induce the apoptosis, while tacrolimus could effectively reduce such a toxic effect in a dose-dependent manner (p<0.05). Also, tacrolimus could significantly reduce the oxidative stress response of cells caused by ox-LDL and the expressions of ERS-related proteins, and protect the cells from oxidative stress injury (p<0.05). CONCLUSIONS: Tacrolimus regulates ERS of endothelial cells to reduce ox-LDL-induced injuries.

Association between KIR gene polymorphisms and type 1 diabetes mellitus (T1DM) susceptibility: A PRISMA-compliant meta-analysis.

BACKGROUND: Type 1 diabetes mellitus (T1DM) is a T-cell mediated autoimmune disease with a complex genetic and immunological background. Evidence suggests that killer cell immunoglobulin-like receptor (KIR) genes are associated with T1DM, but the results are inconsistent. Here, we conducted a meta-analysis to comprehensively evaluate the effect of KIR genes on the risk of T1DM. METHODS: The PubMed, Web of Science, the Chinese Biomedical Database, and Chinese National Knowledge Infrastructure databases were systematically searched to select studies on the association between KIR polymorphisms and T1DM. The quality of each study was scoring in term of the Newcastle-Ottawa Scale. Pooled odds ratios (ORs) and 95% confidence intervals (CIs) were used to assess the strength of this association. Subgroup analysis stratified by ethnicity was also conducted. Funnel plot and Egger test were conducted to assess the publication bias. RESULTS: A total of 13 independent case-control studies comprising 2076 T1DM cases and 1967 controls were included in this meta-analysis. We found a negative association between the KIR2DL1 polymorphism and susceptibility to T1DM in the overall population (OR = 0.71, 95%CI = 0.51-0.98, P = .038), but not in ethnic-specific analysis. Additionally, a negative association between the KIR2DS1 polymorphism and susceptibility to T1DM was found in the Asians (OR = 0.76, 95%CI = 0.63-0.92, P = .004), but not in the Caucasians. However, the associations could not withstand Bonferroni correction. Conversely, no association between the other KIRs genes (KIR2DL2, KIR2DL3, KIR2DL4, KIR2DL5, KIR2DS2, KIR2DS3, KIR2DS4, KIR2DS5, KIR3DL1, KIR3DL2, KIR3DL3, and KIR3DS1) and T1DM susceptibility was found in overall and subgroup ethnicity. No publication bias was detected in all comparisons. CONCLUSIONS: In summary, this meta-analysis suggested that the KIR2DL1 and 2DS1 polymorphism might be a potential protective factor for T1DM in the specific ethnicity. Further subtle design studies with more sample size are still needed for a definitive conclusion.