[Progress and challenge in intelligent syndromic surveillance for infectious diseases].

Intelligent syndromic surveillance is an important part of multi-point triggering and multi-channel surveillance system of intelligent early warning of infectious diseases in China, and an inevitable development process of traditional syndromic surveillance as the constant emergence of new technologies. Intelligent syndromic surveillance collects not only the medical data of patients seeking medical care in hospitals but also massive non-medical information. However, along with its rapid development, challenges in intelligent syndromic surveillance have emerged, such as information explosion, cost-effective balance, information sharing, data security and privacy. This paper summarizes the concept and development of intelligent syndromic surveillance to provide references for the method and technique development of intelligent early warning of infectious diseases and new thought for the prevention and control of infectious diseases in China and in the world.

[Renal insufficiency and venous thromboembolism].

Venous thromboembolism (VTE) is of high incidence and prevalence worldwide. Renal insufficiency has high disease burden with insidious development and is accompanied with disorder of coagulation system. A higher prevalence of VTE has been observed among patients with renal insufficiency whereas VTE patient with renal insufficiency had higher rates of adverse outcomes. Recent evidence indicated that renal insufficiency was an important risk factor for both short and long-term prognosis for VTE. Renal function also affects the choice of anticoagulation therapy and dosage adjustment of drugs. We conducted a comprehensive review of the pathogenesis, mechanism, prognosis and treatment strategy for VTE patients who comorbid renal insufficiency by searching the latest and most advanced national and international articles, to provide integrated information for the prevention and treatment for VTE patients.

[Differential diagnosis of pure ground glass nodules in lung with spectral CT imaging].

Objective: To evaluate the differential diagnostic performance of quantitative parameters derived from the spectral CT imagingin pure ground-glass nodules. Methods: A total of 44 patients with pure ground glass nodules underwent chest energy spectrum CT and with known subsequently pathological findings in the Imaging Department of the Second Affiliated Hospital of Soochow University from August 2017 to September 2019 were retrospectively analyzed. Among them, there are 18 males and 26 females, aged from 26 to 79 (51±12) years. They were divided into as the inflammatory group (n=12), pre-invasive adenocarcinoma group (n=17) and invasive adenocarcinoma group (n=15). The aforementioned three groups were further reclassified as non-invasive adenocarcinoma group (inflammatory lesion+pre-invasive lesion) and invasive adenocarcinoma group in order to evaluating the values of water concentration (WC) for the determination of adenocarcinoma infiltration status. The values of WC derived from the arterial and venous phase of the lesion, iodine concentration (IC), standardized iodine concentration (NIC) were measured respectively.The slope of the energy spectral curve (K40-70KeV) derived from the arterial and venous phase of the lesion was also calculated. One-way ANOVA analysis was performed to compare the differences of the three groups and the multiple comparison method was used for further comparing. Intraclass correlation efficient (ICC) was used to assess the consistency of the three times of measurements. The area under curve(AUC) of Receiver Operating Characteristic (ROC) was conducted to evaluate the diagnostic performance of water based values. Results: The values of WC in the arterial and venous phases were significantly different. As in the inflammatory group, the pre-invasive lesion group and the invasive adenocarcinoma group, the values of WC was (291.95±58.66) mg/cm3, (297.61±63.96) mg/cm3and (374.52±60.62) mg/cm3 of the arterial phase, and (277.07±33.78) mg/cm3, (291.74±50.49) mg/cm3 and (373.33±75.12) mg/cm3 of the venous phase, respectively(all P<0.05). Further comparison demonstrated that no significant difference was observed for the values of WC derived from the arterial phases and venous phases between the inflammatory lesion group and the pre-invasive lesion group (all P>0.05).There were an significant differences between the invasive adenocarcinoma group, the inflammatory lesion group and the pre-invasive lesion group (all P<0.05). The values of WC derived from the venous phase achieved the largest AUC (0.770) for differentiating invasive adenocarcinoma from non-invasive adenocarcinoma (inflammatory lesions+pre-invasive lesions) in the pure ground glass nodules. The sensitivity and specificity were 66.67% and 93.10%, respectively, when using 349.31 mg/cm³ as the optimal threshold. The slope of the spectral curve and iodine-related parameters (IC, NIC) derived from arterial or venous phases among the three groups were not significantly different (all P>0.05). Conclusion: The values of WC derived from the spectral CT can better distinguish inflammatory, pre-invasive lesions and invasive adenocarcinoma, which is helpful for the qualitative analysis for pure ground glass nodules.

[Establishment of a nomogram model for predicting hematoma expansion in intracerebral hemorrhage and its multidimensional evaluation].

Objective: To establish a nomogram model for hematoma expansion (HE) prediction after intracerebral hemorrhage (ICH) and evaluate its performance in a multidimensionally way. Methods: A total of 348 ICH patients who were firstly diagnosed and hospitalized in the Second Affiliated Hospital of Soochow University from January 2017 to December 2019 were collected retrospectively. There were 236 males and 112 females, and their age ranged from 18 to 94 (62.0±14.6) years. All patients were divided into HE group (n=121) or non-HE group (n=227) according to the presence or absence of HE. The clinical and imaging features were compared between the two groups. Multivariate logistic regression analysis was performed for determining the independent predicting factors for HE prediction and a Nomogram model was established by using these factors. Receiver operating characteristic (ROC) curve, calibration curve, and decision curve analysis (DCA) were used to evaluate the prediction effectiveness, accuracy and clinical practicability of the model, respectively. Bootstrap method was used for internal validation. Results: There were significant differences in onset time, swirl sign, history of anticoagulants administrations, systolic blood pressure when admission, Glasgow coma scale (GCS) scores and RBC distribution width between the two groups[(1.77(1.0, 2.5) h vs 2(1, 3) h, 72 cases (59.5%) vs 94 cases (41.4%), 17 cases (14.0%) vs 15 cases (6.6%), (170.69±29.19) mmHg(1 mmHg=0.133 kPa) vs (163.84±26.07) mmHg, 11(8, 14) scores vs 14(10, 15) scores, 44.3% (41.2%, 46.8%) vs 42.4% (40.1%, 45.3%);respectively, all P<0.05]. Multivariate logistic regression analysis demonstrated that onset time (OR=0.809, 95%CI: 0.682-1.961, P=0.015), swirl sign (OR=0.562, 95%CI:0.349-0.905, P=0.018), history of anticoagulants administrations (OR=0.394, 95%CI: 0.180-1.861, P=0.020), and GCS (OR=0.881, 95%CI: 0.815-1.952, P=0.001) were the predicting factors for HE. The area under the curve (AUC) of the Nomogram model was 0.735(95%CI: 0.687-0.805), which demonstrated that the model has an ideal prediction effectiveness. The calibration curve showed that the prediction probability of HE of the model fits well with the actual probability, and with high calibration. DCA showed relatively wide range of optional threshold probability of the model (ranging from 14% to 72%), the clinical practicability of this model was high. The internal validation results showed a C-index of 0.703, indicated a good discrimination power. Conclusion: The established Nomogram model can predict the HE of ICH with good prediction effectiveness, discrimination power and with good clinical practicability, which can be capable of providing an intuitive and visual guidance tool for timely identifying ICH patients who may have HE.

[The value of 3.0T magnetic resonance -diffusion kurtosis imaging in the differential diagnosis of rectal mucinous adenocarcinoma and common adenocarcinoma].

Objective: To investigate the value of 3.0T MRI diffusion kurtosis imaging (DKI) quantitative histogram parameters in the differential diagnosis of rectal mucinous adenocarcinoma (MC) and common adenocarcinoma (AC). Methods: One hundred and ten patients from Department of Radiology, the Second Affiliated Hospital of Soochow University between September 2015 and September 2019 with complete magnetic resonance imaging (MRI) and DKI results confirmed by surgery and pathology were retrospectively analyzed, including 16 patients in MC group and 94 patients in AC group. Two physicians outlined the region of interest (ROI) on the DKI image with b=1 000 s/mm(2), and obtained quantitative DKI parameters, including the diffusion coefficient (D value) and kurtosis coefficient (K value) corrected for non-Gaussian distribution. The apparent diffusion coefficient (ADC) values of quantitative parameters of diffusion-weighted imaging (DWI) were obtained through image registration, and histogram analysis was performed to obtain the mean value, 25th percentile, 50th percentile, 75th percentile, skewness and kurtosis of the above parameters, respectively. The difference between the quantitative histogram parameter analysis results of the rectal MC group and the AC group was evaluated, and the main indicators and multivariate comprehensive analysis indicators was screened, and the effectiveness of quantitative histogram parameters related to histopathological classification in the differential diagnosis of rectal MC and AC was evaluated. Results: There was no significant differences in gender, age, lesion location, T stage or N stage between MC group and AC group (all P>0.05). The multivariate binary logistic stepwise regression screening showed that D50th percentile and K25th percentile are statistically significant indicators (B values were 2 966.166 and -4.550, respectively; Wals values were 9.000 and 15.720, respectively; and P values were 0.003 and <0.001, respectively). The combined area under the curve of the two indictors was 0.85, but there was no statistically significant difference in pairwise comparison using DeLong method (P>0.05). The results of histogram analysis of quantitative parameters measured by the two physicians were consistent, and the inter-group correlation coefficient ranged from 0.880 to 0.981. Conclusions: The quantitative parameter histogram analysis of the DKI double-index model is helpful for the differentiation of rectal MC and AC, in which the D50th percentile and K25th percentile have differential diagnosis significance, and are superior to the ADC value of the single-index model.

[Diagnostic value of gadobenate dimeglumine enhanced MRI in hepatic fibrosis of rats].

Objective: To evaluate the value of Gd-BOPTA enhanced MRI in the staging of liver fibrosis. Method: Fifty male SD rats (6-week-old, 180-220 g) were divided into the modeling group (n=42) and the control group (n=8). The model of liver fibrosis in the modeling group was established by carbon tetrachloride (animal license No. SYXK (Su) 2017-0043). From week 2 to week 10, rats in the modeling group (n=4) and control group (n=1) were randomly selected to scan 1 h(RER1), 2 h(RER2) and 3 h(RER3) after injection of Gd-BOPTA, respectively, to measure the relative enhancement rate (RER) of liver parenchyma. The shape of intrahepatic bile duct and the degree of enhancement at each time point were observed. Results: Forty-two rats (34 rats in the modeling group and 8 rats in the control group) completed the experiment. RER1, RER2 and RER3 of the control group were 1.44±0.37, 1.22±0.37 and 0.84±0.28 respectively. RER1, RER2 and RER3 of the modeling group were respectively: S1 (n=6): 1.49±0.48, 1.29±0.39, 0.91±0.38;S2 (n=9): 1.48±0.44, 1.34±0.37, 1.04±0.40;S3 (n=11): 1.49±0.43, 1.37±0.39, 1.21±0.30; S4 (n=8): 1.49±0.44, 1.40±0.37, 1.24±0.40. There was no significant difference in RER1 and RER2 values between the control group and the liver fibrosis group (F=0.022, P=0.999; F=0.301, P=0.875). There were significant differences between the control group and RER3 values of hepatic fibrosis stage S3 and S4 (t=2.249, P=0.031; t=2.274, P=0.029), there was no significant difference between the remaining groups (all P>0.05).In the control group, the intrahepatic bile duct was obviously strengthened within 1 hour after enhancement, and walked naturally. The intrahepatic bile duct was slightly enhanced 1h after the enhancement of S3-S4 stage of hepatic fibrosis, and the intrahepatic bile duct was significantly enhanced 2-3 hours later, with distorted alignment. Conclusion: Delayed 3 hours liver parenchymal RER and intrahepatic bile duct distortion delay enhancement after Gd-BOPTA enhancement contribute to the S3-S4 diagnosis of liver fibrosis.

[Applied research of "quadri-low" combined with automatic tube current modulation and iterative model reconstruction technology in head and neck CT angiography].

Objective: To investigate the feasibility of low tube voltage, low contrast medium concentration, injection rate and volume (quadri-low) combined with automatic tube current modulation (ATCM) and iterative model reconstruction (IMR) technology in head and neck CT angiography (CTA). Methods: A total of 70 patients whose body mass index (BMI)<25 kg/m(2) underwent head and neck CTA and digital subtraction angiography (DSA) from January to July 2017 were enrolled in this prospective study. According to random number table, patients were divided into two groups: group A (n=35) was scanned according to the protocol of 120 kV, 150 mAs, 50 ml and 5 ml/s iopromide (370 mg/ml) and filtered back projection (FBP) reconstruction; group B (n=35) was scanned with 80 kV, ATCM with mean tube current of 100 mAs, 30 ml and 3 ml/s iohexol (300 mg/ml) and IMR; the other parameters kept consistent between the two groups. The maximum transverse neck diameter at the level of the hyoid bone, artery CT value and image noise were measured, signal to noise ratio (SNR), contrast to noise ratio (CNR) and figure of merit (FOM) were calculated, and the image quality was evaluated subjectively and compared with those reconstructed by DSA. Scan length, volume CT dose index (CTDIvol) and dose length product (DLP) were recorded, and the effective dose (ED) was calculated. The chi-square and independent-sample t tests were used to compare the inter-group differences in these aforementioned data. Resutls: No significant difference was found in general information between the two groups. No significant difference existed in artery CT value, image noise, SNR and CNR between the two groups (t=-1.170-1.365, all P>0.05); however, the FOM of group B (74±40) was significantly higher than that in group A (12±4) (Z=-7.195, P=0.000). The image quality of the two groups met the requirement of clinical diagnosis[(4.1±0.7) vs (4.2±0.8) points, Z=-0.592, P>0.05], no significant difference was found in subjective evaluation and diagnostic efficacy. The CTDIvol, DLP and ED in group B were all significantly lower than those in group A (Z=-7.728, -7.202, -7.206, all P<0.05). The iodine load and iodine delivery rate (IDR) of group B was lower than that of group A (18.5 g vs 9.0 g, 1.85 mg/s vs 0.90 mg/s), and they were reduced for 51.4% in group B. Conclusions: For patients of BMI <25 kg/m(2,) low tube voltage, low contrast medium concentration, injection rate and volume combined with ATCM and IMR technology can significantly decrease radiation dose, iodine load and IDR while maintain the image quality in head and neck CTA examination.

[The quantitative evaluation of early radiation-induced changes in the salivary glands using MRI].

Objective: To quantitatively evaluate the early radiation injury of salivary glands in patients with nasopharyngeal carcinoma (NPC) after intensity-modulated radiotherapy (IMRT). Methods: Twenty patients with NPC between 2014 and 2015 from the Second Affiliated Hospital of Soochow University were retrospectively analyzed.All patients underwent an MRI scan before and after IMRT.The volumes, T(1)WI, T(2)WI signal intensity(SIs) and apparent diffusion coefficient (ADCs) of the parotid and submandibular glands were measured.The relative signal intensity (RSIs) of each salivary gland was calculated with cerebrospinal fluid as control.The quantitative parameters of salivary glands were compared before and after radiotherapy. Results: The volumes (cm(3)) and T(1)WI RSIs of the parotid and submandibular glands (14.88±6.00, 5.21±1.76, 2.98±1.05, 1.88±0.42, respectively) were significantly lower than those before radiotherapy (22.26±8.26, 7.76±2.45, 3.58±1.02, 2.27±0.50, respectively) (t=9.921, 4.013, 10.126, 4.202, respectively, P=0.000 for all). The T(2)WI RSIs and ADCs (×10(-3) mm(2)/s) of the parotid and submandibular glands (0.50 ± 0.08, 0.41±0.04, 1.31±0.19, 1.50±0.13, respectively) were significantly higher than those before radiotherapy (0.45±0.07, 0.33±0.05, 1.02±0.21, 1.23±0.13, respectively) (t=-4.846, -9.276, -9.957, -10.679, respectively, P=0.000 for all). The volumes of parotid and submandibular glands were correlated with ADCs (r=-0.512, P=0.000; r=-0.358, P=0.001; respectively). The volumes and ADCs of submandibular glands were correlated with T(1)WI RSIs and T(2)WI RSIs(P<0.05). Conclusion: MRI can quantitatively evaluate the early changes of salivary glands after radiotherapy of nasopharyngeal carcinoma as a noninvasive method, and has high clinical application potential.

Revealing extraordinary tensile plasticity in layered Ti-Al metal composite.

Layered Ti-Al metal composite (LMC) fabricated by hot-pressing and hot-rolling process displays higher ductility than that of both components. In this paper, a combination of digital image correlation (DIC) and X-ray tomography revealed that strain delocalization and constrained crack distribution are the origin of extraordinary tensile ductility. Strain delocalization was derived from the transfer of strain partitioning between Ti and Al layer, which relieved effectively the strain localization of LMC. Furthermore, the extensive cracks of LMC were restricted in the interface due to constraint effect. Layered architecture constrained the distribution of cracks and significantly relieved the strain localization. Meanwhile, the transfer of strain partitioning and constrained crack distribution were believed to inhibit the strain localization of Ti and change the deformation mechanisms of Ti. Our finding enriches current understanding about simultaneously improving the strength and ductility by structural design.

A novel approach to accelerate the reaction between Ti and Al.

Pure Ti foils and SiCp/Al composite foils were employed to investigate the parabolic growth kinetics of TiAl3 at 660°C. Compared with pure Al foils, the introduction of SiC particles significantly refined TiAl3 grain size by the solid solution of silicon. Corresponding refinement mechanisms were concluded from the perspective of the nucleation of TiAl3. Micromechanics analysis shows that the fine TiAl3 grains own a small viscous resistance, and subsequently an improvement in the reaction rate could be achieved. This meaningful law also applies extensively to Ni/Al and Fe/Al systems.

Formation of intermetallic compound layer in multi-laminated Ni-(TiB2/Al) composite sheets during annealing treatment.

Solid-state reactive diffusion between Ni and Al was investigated during annealing at 650°C by employing multi-laminated Ni-(TiB(2)/Al) composite sheets. In multi-laminated Ni-(TiB(2)/Al) composite sheets annealed up to 5min NiAl(3) was the only phase observed in the diffusion zone, and Ni(2)Al(3) appeared after longer annealing time. Most grains of Ni(2)Al(3) showed equiaxed morphology rather than columnar microstructures like NiAl(3), due to the low concentration gradients of Al and Ni at the Ni/NiAl(3) interface. The preferential formation of this intermetallic compound NiAl(3) in multi-laminated Ni-(TiB(2)/Al) composite sheets was predicted using an effective heat of formation model. The present work indicated that both Ni and Al interdiffused, and the formation of NiAl(3) was a reaction-diffusion process.

Protein phosphatase 2A interacts with and directly dephosphorylates RelA.

Nuclear factor-kappa B (NF-kappa B)/Rel transcription factors are key regulators of a variety of genes involved in inflammatory responses, growth, differentiation, apoptosis, and development. There are increasing lines of evidence that NF-kappa B/Rel activity is controlled to a great extent by its phosphorylation state. In this study, we demonstrated that RelA physically associated with protein phosphatase 2A (PP2A) subunit A (PR65). Both the N- and C-terminal regions of RelA were responsible for the PP2A binding. RelA co-immunoprecipitated with PP2A in melanocytes in the absence of stimulation, indicating that RelA forms a signaling complex with PP2A in the cells. RelA was dephosphorylated by a purified PP2A core enzyme, a heterodimer formed by the catalytic subunit of PP2A (PP2Ac) and PR65, in a concentration-dependent manner. Okadaic acid, an inhibitor of PP2A at lower concentration, increased the basal phosphorylation of RelA in melanocytes and blocked the dephosphorylation of RelA after interleukin-1 stimulation. Interestingly, PP2A immunoprecipitated from melanocytes was able to dephosphorylate RelA, whereas PP2A immunoprecipitated from melanoma cell lines exhibited decreased capacity to dephosphorylate RelA in vitro. Moreover, in melanoma cells in which I kappa B kinase activity was inhibited by sulindac to a similar level as in melanocytes, the phosphorylation state of RelA and the relative NF-kappa B activity were still higher than those in normal melanocytes. These data suggest that the constitutive activation of RelA in melanoma cells (Yang, J., and Richmond, A. (2001) Cancer Res. 61, 4901-4909) could be due, at least in part, to the deficiency of PP2A, which exhibits decreased dephosphorylation of NF-kappa B/RelA.

Constitutive activity of the human beta(1)-adrenergic receptor in beta(1)-receptor transgenic mice.

We tested the hypothesis that the human beta(1)-adrenergic receptor displays constitutive activity and that beta-adrenergic antagonists differ in their ability to modulate this constitutive activity. Transfection of the cDNAs of the human beta(1)- and beta(2)-adrenergic receptors into COS-7 cells caused increases in basal cAMP that were proportional to the receptor levels, thus demonstrating constitutive activity for both subtypes. At comparable receptor levels, the increase in basal cAMP was about 5-fold higher for the beta(2)- than for the beta(1)-subtype. As a model for enhanced beta-adrenergic signaling at the whole-organ level, we used transgenic mice with heart-specific overexpression of the human beta(1)-adrenergic receptor. In this model, the beta(1)-adrenergic receptor displayed constitutive activity as evidenced by a higher spontaneous beating rate of isolated right atria from beta(1)-transgenic versus wild-type mice. This difference was abolished by the addition of CGP20712A, demonstrating inverse agonist properties of this compound. We then tested whether various beta-adrenergic antagonists currently in clinical use for the treatment of heart failure differ in their ability to modulate constitutive activity of the cardiac beta(1)-adrenergic receptor. The beta(1)-selective antagonists metoprolol and bisoprolol showed significant inverse agonist activity at the beta(1)-adrenergic receptor. Carvedilol behaved as a neutral antagonist and xamoterol displayed marked partial agonist activity. We conclude that the human beta(1)-adrenergic receptor displays constitutive activity that is considerably lower than that of the beta(2)-subtype. beta-Adrenergic antagonists currently in clinical use differ in their ability to exert inverse agonist activity at the human beta(1)-adrenergic receptor, which may contribute to their therapeutic effects.

Phosphorylation-independent association of CXCR2 with the protein phosphatase 2A core enzyme.

Protein phosphatase 2A (PP2A) is postulated to be involved in the dephosphorylation of G protein-coupled receptors. In the present study, we demonstrate that the carboxyl terminus of CXCR2 physically interacts with the PP2A core enzyme, a dimer formed by PP2Ac and PR65, but not with the PP2Ac monomer, suggesting direct interaction of the receptor with PR65. The integrity of a sequence motif in the C terminus of CXCR2, KFRHGL, which is conserved in all CC and CXC chemokine receptors, is required for the receptor binding to the PP2A core enzyme. CXCR2 co-immunoprecipitates with the PP2A core enzyme in HEK293 cells and in human neutrophils. Overexpression of dominant negative dynamin 1 (dynamin 1 K44A) in CXCR2-expressing cells blocks the receptor association with the PP2A core enzyme, and an internalization-deficient mutant form of CXCR2 (I323A,L324A) also exhibits impaired association with the PP2A core enzyme, suggesting that the receptor internalization is required for the receptor binding to PP2A. A phosphorylation-deficient mutant of CXCR2 (331T), which has previously been shown to undergo internalization in HEK293 cells, binds to an almost equal amount of the PP2A core enzyme in comparison with the wild-type CXCR2, suggesting that the interaction of the receptor with PP2A is phosphorylation-independent. The dephosphorylation of CXCR2 is reversed by treatment of the cells with okadaic acid. Moreover, pretreatment of the cells with okadaic acid increases basal phosphorylation of CXCR2 and attenuates CXCR2-mediated calcium mobilization and chemotaxis. Taken together, these data indicate that PP2A is involved in the dephosphorylation of CXCR2. We postulate that this interaction results from direct binding of the regulatory subunit A (PR65) of PP2A to the carboxyl terminus of CXCR2 after receptor sequestration and internalization.

Identification of a motif in the carboxyl terminus of CXCR2 that is involved in adaptin 2 binding and receptor internalization.

Agonist treatment of cells expressing the chemokine receptor, CXCR2, induces receptor phosphorylation and internalization through a dynamin-dependent mechanism. In the present study, we demonstrate that a carboxyl terminus-truncated mutant of CXCR2 (331T), which no longer undergoes agonist-induced phosphorylation, continues to undergo ligand-induced internalization in HEK293 cells. This mutant receptor exhibits reduced association with beta-arrestin 1 but continues to exhibit association with adaptin 2 alpha and beta subunits. Replacing Leu320-321 and/or Ile323-Leu324 with Ala (LL320,321AA, IL323,324AA, and LLIL320,321,323,324AAAA) in wild-type CXCR2 or 331T causes little change in ligand binding and signaling through Ca(2+) mobilization but greatly impairs the agonist-induced receptor sequestration and ligand-mediated chemotaxis. The LL320,321AA, IL323,324AA, and LLIL320,321,323,324AAAA mutants of CXCR2 exhibit normal binding to beta-arrestin 1 but exhibit decreased binding to adaptin 2alpha and beta. These data demonstrate a role for the LLKIL motif in the carboxyl terminus of CXCR2 in receptor internalization and cell chemotaxis and imply a role for adaptin 2 in the endocytosis of CXCR2.

Inhibition of calcium/calmodulin-dependent protein kinase II in rat hippocampus attenuates morphine tolerance and dependence.

Learning and memory have been suggested to be important in the development of opiate addiction. Based on the recent findings that calcium/calmodulin-dependent protein kinase II (CaMKII) is essential in learning and memory processes, and morphine treatment increases CaMKII activity in hippocampus, the present study was undertaken to examine whether inhibition of hippocampal CaMKII prevents morphine tolerance and dependence. Here, we report that inhibition of CaMKII by intrahippocampal dentate gyrus administration of the specific inhibitors KN-62 and KN-93 to rats significantly attenuated the tolerance to the analgesic effect of morphine and the abstinence syndrome precipitated by opiate antagonist naloxone. In contrast, both KN-04 and KN-92, the inactive structural analogs of KN-62 and KN-93, failed to attenuate morphine tolerance and dependence, indicating that the observed effects of KN-62 and KN-93 are mediated through inhibition of CaMKII. Furthermore, administration of CaMKII antisense oligonucleotide into rat hippocampal dentate gyrus, which decreased the expression of CaMKII specifically, also attenuated morphine tolerance and dependence, while the corresponding sense oligonucleotide of CaMKII did not exhibit such inhibitory effect. Moreover, the KN-62 treatment abolished the rewarding properties of morphine as measured by the conditioned place preference. These results suggest that hippocampal CaMKII is critically involved in the development of morphine tolerance and dependence, and inhibition of this kinase may have some therapeutic benefit in the treatment of opiate tolerance and dependence.

Suppression of phospholipase C blocks Gi-mediated inhibition of adenylyl cyclase activity.

The potential effect of inhibition of phospholipase C on the response of Gi-coupled receptors was investigated in neuroblastoma x glioma hybrid (NG108-15) cells. The phospholipase C specific inhibitor 1-[6-((17beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl]-1H -pyrrole-2,5-dione (U73122), which did not affect basal and forskolin-stimulated adenylyl cyclase activities, time- and dose-dependently blocked delta-opioid receptor-mediated inhibition of adenylyl cyclase activity, the EC50 (0.5 microM) of which was consistent with that for inhibition of bradykinin-dependent phospholipase C activation (EC50 = 1 microM). U73122 treatment also blocked functional responses of m4 muscarinic receptor and alpha2-adrenoceptor in NG108-15 cells and three opioid receptors (mu, delta and opioid receptor-like receptor (ORL1)) in human neuroblastoma SK-N-SH cells. 1-[6-((17Beta-3-Methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl]-2, 5-pyrrolidinedione (U73343), an inactive analog of U73122, did not show any effect, which suggests that the blockade by U73122 of Gi-coupled receptor-mediated signaling is probably mediated through inhibition of phospholipase C, although a possible direct modification of G proteins can not be excluded. Furthermore, treatment with U73122 but not U73343 blocked the GTP-induced inhibition of adenylyl cyclase, indicating blockade at the level of Gi proteins.

N-Methyl-D-aspartate attenuates opioid receptor-mediated G protein activation and this process involves protein kinase C.

The effects of N-methyl-D-aspartate (NMDA) on opioid receptor-mediated G protein activation were explored in neuroblastoma X glioma hybrid (NG108-15) cells. Treatment of the cells with NMDA resulted in a remarkable attenuation of [35S]guanosine-5'-O-(3-thio)triphosphate binding stimulated by [D-Pen2,D-Pen5]-enkephalin (DPDPE), a delta-opioid receptor agonist. The effects of NMDA were dose and time dependent with an IC50 value of 5 nM and could be blocked by NMDA receptor antagonists. After NMDA treatment, the DPDPE dose-response curve shifted to the right (EC50 value increased approximately 7-fold, from 6 to 40 nM), and the maximal response induced by DPDPE was reduced by approximately 60%. The effects of NMDA were reversible, and the DPDPE response could recover within 60 min. The functional responses of delta-, mu-, and kappa-opioid receptors in primarily cultured neurons also were attenuated significantly by NMDA treatment. The inhibitory effects of NMDA on opioid receptor-mediated G protein activation could be blocked by coadministration of the protein kinase C (PKC) inhibitors or by elimination of the extracellular Ca2+. Correspondingly, NMDA treatment of NG108 cells significantly elevated cellular PKC activity and stimulated Gialpha2 phosphorylation. Transient transfection into NG108-15 cells of the wild-type Gialpha2 and a mutated Gialpha2 (Ser144Ala) resulted in a 2-fold increase in DPDPE-stimulated G protein activation. The DPDPE responses were greatly inhibited by NMDA treatment in the wild-type Gialpha2-transfected cells but much less affected in the mutant Gialpha2-transfected cells. In summary, NMDA attenuates opioid receptor/G protein coupling, and this process requires activation of PKC.

Inhibition of phospholipase C blocks opioid receptor-mediated activation of Gi proteins.

Activation of phospholipase C (PLC) in response to stimulation of delta-opioid, m4 muscarinic and alpha2 adrenergic receptors was observed in NG108-15 cells. Treatment with PLC specific inhibitors, U73122 and ET-18-OCH3, blocked delta-opioid receptor-mediated activation of G proteins with no effect on opioid binding to the receptors. U73122 treatment also suppressed functional responses of m4 muscarinic and alpha2 adrenergic receptors in NG108-15 cells. Furthermore, the G protein activation mediated by mu- and delta-opioid receptors and opioid receptor-like receptor (ORL1) were abolished by U73122 in SK-N-SH neuroblastoma cells. Inhibition of adenylyl cyclase induced by high concentrations of GTP was blocked by U73122, suggesting that blockade is at the level of G proteins. Our results thus indicated that inhibition of PLC leads to blockade of Gi protein activation mediated by opioid receptors or other Gi-coupled receptors.