[Effects and clinical significance of NLRP3 inflammasome activated by IL-17A in CRSwNP].

Objective: To investigate the effects and clinical significance of NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome activated by interleukin (IL)-17A in chronic rhinosinusitis with nasal polyps (CRSwNP). Methods: Patients underwent nasal endoscopic surgery in the Third Affiliated Hospital of Sun Yat-sen University from January 2020 to December 2021 were collected, including 28 CRSwNP (including 19 males and 9 females, aged 19 to 67 years), 22 chronic rhinosinusitis without nasal polyps (CRSsNP) and 22 controls. qRT-PCR was used to detect the expressions of IL-17A, NLRP3, IL-1ß and IL-18 in the three groups, and their correlations were analyzed. The positions of IL-17A, NLRP3 and IL-18 in nasal polys were analyzed by immunofluorescence. Western Blotting and ELISA were employed to detect the expression of NLRP3, IL-1ß and IL-18 in the human nasal epithelial cells after using IL-17A stimulation or IL-17A receptor inhibitor. Immunofluorescence was used to observe the NLRP3, IL-1ß, and IL-18 protein expression after IL-17A stimulating human nasal epithelial cells, and after the use of IL-17A receptor inhibitor and NLRP3 inhibitor MCC950. The correlations between NLRP3, IL-1ß, IL-18 and CT scores, nasal endoscopic scores, visual analogue scale (VAS) scores, and sino-nasal outcome test (SNOT) 22 scores of CRSwNP patients were analyzed. SPSS 20.0 software was used for statistical analysis. Results: The expressions of IL-17A, NLRP3, IL-1ß and IL-18 in the tissues of CRSwNP patients were significantly higher than those in CRSsNP group(P=0.018,P<0.001,P=0.005, P=0.016) and the control group(all P<0.001). IL-17A was positively correlated with the expression of NLRP3, IL-1ß, and IL-18(r ralue was 0.643,0.650,0.629,respectively, all P<0.05). IL-17A, NLRP3, and IL-18 were co-localized in the epithelial propria of polyp tissue. IL-17A stimulated the expressions of NLRP3, IL-1ß, and IL-18 in human nasal epithelial cells. After the use of IL-17A receptor inhibitor, the expressions of NLRP3, IL-1ß, and IL-18 were significantly down-regulated. After the use of NLRP3 inhibitor MCC950, IL-17A was significantly down-regulated to promote the expression of NLRP3, IL-1ß, and IL-18. The expressions of NLRP3, IL-1ß and IL-18 were positively correlated with CT, nasal endoscopy, VAS, and SNOT22 scores in patients with CRSwNP. Conclusions: IL-17A promotes the release of IL-1ß and IL-18 by activating the NLRP3 inflammasome and aggravates the severity of the disease in CRSwNP.

[The expression profile and potential regulatory mechanism of ACE2 in chronic rhinosinusitis with nasal polyps].

Objective: To preliminarily analyze the expression of angiotensin-converting enzyme 2 (ACE2) and to investigate its potential regulatory mechanism in chronic rhinosinusitis with nasal polyps (CRSwNP). Methods: Patients underwent nasal endoscopic surgery in the Third Affiliated Hospital of Sun Yat-sen University from February 2020 to May 2021 were selected, including 17 males and 6 females, aging from 23 to 66 years old. Expression of ACE2 was evaluated via immunohistochemical staining in controls with non-chronic rhinosinusitis, non-eosinophilic CRSwNP (non-ECRSwNP), and eosinophilic CRSwNP (ECRSwNP) tissue, respectively. Correlations between ACE2 and the indicated Th1/Th2-related cytokines (IFN-γ, IL-4, IL-5, IL-13, IL-25, IL-33, TSLP and periostin) were analyzed based on GSE72713 dataset. Protein-protein interaction (PPI) network was constructed via string database, immune infiltration of GSE72713 dataset was evaluated using cibersort algorithm. ACE2 was comprehensively analyzed by microRNA regulatory network, gene set enrichment analysis (GSEA) and pharmacological analysis. Statistical analysis was performed using GraphPad 7.0 and SPSS 20.0 software. Results: ACE2 was up-regulated in non-ECRSwNP compared with ECRSwNP. Microarray analysis showed that ACE2 was positively correlated with IFN-γ while inversely correlated with IL-5, IL-13 and periostin significantly. Analysis of immune infiltration suggested that ACE2 expression correlated positively with the number of M1 macrophage while negatively with M2 macrophage. GSEA demonstrated that interferon-related signaling pathways were up-regulated in non-ECRSwNP, and miRNA-200B/miRNA-200C/miRNA-429 pathways targeting ACE2 were enriched in ECRSwNP. Results of pharmacological analysis indicated that ampicillin was able to promote the expression of ACE2 whereas acetaminophen could down regulated the expression of ACE2. Conclusion: Expression pattern of ACE2 is varied in non-ECRSwNP and ECRSwNP, which may be related to the different infiltration of indicated cytokines and different regulatory pathways of miRNA.

[Expression of CD8+Treg cells in chronic rhinosinusitis and its correlation with eosinophilic infiltration].

Objective: To detect the percentages of CD8+Treg cells in the nasal mucosa and peripheral blood of chronic rhinosinusitis (CRS) and to explore their correlation with eosinophilic infiltration. Methods: Thirty-three chronic rhinosinusitis with polyp (CRSwNP), 26 chronic rhinosinusitis without polyp (CRSsNP) and 27 control patients who were collected with the nose mucosal tissue and peripheral blood in the Third Affiliated Hospital of Sun Yat-sen University from March 2017 to October 2018 were selected, including 59 males and 27 females, aging from 18 to 72 years. Hematoxylin and eosin (HE) staining was used to observe the number of eosinophils in the nasal tissues and to classify the CRS into eosinophilic CRS (ECRS) and non-eosinophilic CRS (Non-ECRS). Flow cytometry was used to detect the percentages of CD4+ and CD8+T cells in lymphocytes of nasal mucosa and peripheral blood. The percentages of CD8+Foxp3+Treg cells, CD8+Foxp3-IL-10+Treg cells, CD8+IFN-γ+T cells (Tc1), CD8+IL-4+T cells (Tc2) and CD8+IL-17A+T cells (Tc17) in lymphocytes of nasal mucosa and peripheral blood were also tested. Besides, the percentages of Foxp3+TGF-ß+Treg cells and Foxp3+IL-10+Treg cells in CD8+T cells were determined. All data were represented by M (IQR). GraphPad 7.0 and SPSS 16.0 were used for illustration and statistical analysis. Results: The percentage of CD8+T cells (37.75%(17.35%)) was higher than that of CD4+T cells (4.72%(4.29%)) in nasal mucosa (Z=-5.70, P<0.001), while lower (23.60%(9.33%)) than that of CD4+T cells (44.05% (10.93%)) in peripheral blood (t=9.72, P<0.001). CRSwNP patients possessed the highest Tc2 (1.82% (1.22%)) and Tc17 (1.93% (2.32%)) percentages than CRSsNP (Tc2: 0.84% (0.79%); Tc17: 0.54% (1.04%)) and control (Tc2: 1.09% (0.92%); Tc17: 0.47% (0.51%), both P<0.05) patients. While, CRSwNP patients possessed the lowest CD8+Foxp3+Treg cells percentage (0.10% (0.32%)) than CRSsNP (0.43% (1.45%)) and control (0.48% (0.83%), Z value was -2.24, -2.22, respectively, P value was 0.025, 0.027, respectively). The percentages of Foxp3+TGF-ß+Treg cells and Foxp3+IL-10+Treg cells of CD8+T cells in nasal mucosa in CRSwNP were also lower than controls (Z value was 1.46, 0.49, respectively, both P=0.001). Moreover, the percentage of CD8+Foxp3-IL-10+Treg cells of CD8+T cells was decreased in nasal mucosa of CRSwNP patients (0.14% (0.28%)) when compared with that of CRSsNP (0.89% (0.81%), Z=0.61, P=0.03). ECRS patients had the lower percentages of CD8+Foxp3+Treg cells (0.07% (0.44%)) and CD8+Foxp3-IL-10+Treg cells (0.13% (0.21%)) than Non-ECRS patients (CD8+Foxp3+Treg cells: 0.53% (0.75%); CD8+Foxp3-IL-10+Treg cells: 0.29% (0.76%), t value was 2.14, 2.78, respectively, both P<0.05). The percentage of CD8+Foxp3+Treg cells and the ratio of CD8+Foxp3-IL-10+T per CD8+T cells were negatively correlated with the percentage of eosinophils in CRS patients(R2 value was 0.56, 0.78, respectively, both P<0.001). There was no significant difference in the distribution of CD8+Fxop3+Treg cells and CD8+Fxop3-IL-10+Treg cells in peripheral blood among different groups. Conclusion: The percentages of CD8+Treg cells decrease in CRSwNP patients, especially in ECRS patients, which are opposite to that of Tc2 and Tc17, and negatively correlate with the eosinophils percentage. This indicates that the decrease in the ratio of CD8+Treg cell may be associated with the immune-imbalance and eosinophilic infiltration in nasal mucosa of CRS patients.

[miR-18a enhances the radiosensitivity of nasopharyngeal carcinoma cells through inducing autophagy].

Objective: To explore the impacts of miR-18a overexpression or depression on the radiosensitivities of nasopharyngeal carcinoma cell line CNE1 and CNE2 and underlying mechanisms. Methods: CNE1 and CNE2 were transfected with miR-18a mimics, inhibitor and the corresponding control vectors. qRT-PCR and western blot were used to determine the ataxia telangiectasia mutated (ATM) expressions in CNE1 and CNE2. CNE1 and CNE2 with stably expressing miR-18a and miR-18a siRNA were constructed. Methyl thiazolyl tetrazolium (MTT) assay was used to detect the impacts of the miR-18a overexpression or depression combined with irradiation on the cell growth. Flow cytometry was used to detect the cell apoptosis and cell cycle. Colony formation assay was used to evaluate the raodiosensitivities of cells. Acridine orange (AO) staining and western blot were used respectively to test the autophagy and the expressions of related proteins. Independent samples t test was used to compare the mean value between groups by using SPSS 16.0. Results: ATM mRNA was decreased significantly in CNE1 and CNE2 cells transfected with 100 or 200 nmol/L miR-18a mimics for 48 hours (CNE1: RQ=0.174±0.139 and 0.003±0.001, t=9.939 and 19 470.783;CNE2: RQ=0.024±0.008 and 0.019±0.012, t=270.230 and 137.746, respectively, all P<0.001). ATM proteins were also decreased after transfected with 100 or 200 nmol/L miR-18a mimics for 72 hours. While in the cells transfected with 100 and 200 nmol/L miR-18a inhibitor for 48 hours, the expressions of ATM mRNA were upregulated significantly (CNE1: RQ=9.419±2.495 and 2.500±1.063, t=-4.427 and -41.241; CNE2: RQ=7.210±0.171 and 115.875±15.805, t=-62.789 and -12.589, all P<0.05), and the expressions of ATM proteins increased after transfected for 72 hours. The growth of cells with miR-18a overexpression plus 4 Gy irradiation were obviously inhibited compared to that of cells with the 4Gy irradiation alone; while the growth of miR-18a-inhibited cells increased compared to that of cells with 4 Gy irradiation alone (all P<0.05). CNE1 transfected with 100 nmol/L miR-18a mimics plus 4 Gy irradiation showed the higher apoptosis rate than the cells with 4 Gy irradiation alone ((22.9±2.1)% vs. (16.3±1.0)%, t=-4.870, P<0.01). Compared to the cells with 4 Gy irradiation alone, miR-18a-overexpressed cells plus 4 Gy irradiation decreased their percentages in G1 phases ((20.2±3.0)% vs. (29.8±4.4)%, t=3.119) and G2/M phases ((21.5±0.9)% vs. (33.4±3.1)%, t=6.410, P<0.05 for both), and increased their percentages in S phases ((56.7±4.9)% vs. (36.8±6.4)%, t=-4.246, P<0.05), and these cells possessed less colony number after exposure to different doses of irradiation, more autophagy-lysosome number, and more expressions of LC3 proteins (all P<0.05). There were no significant differences in the expressions of p62 expressions between different groups of cells. Conclusion: Overexpression of miR-18a can enhance the radiosensitivities of NPC cells by targeting ATM to abrogate G1/S, G2/M arrest and to induce autophagy and apoptosis.

The combination of soluble tumor necrosis factor receptor type 1 and fibroblast growth factor 21 exhibits better prediction of renal outcomes in patients with type 2 diabetes mellitus.

PURPOSE: Numerous biomarkers of diabetic kidney disease (DKD) are associated with renal prognosis but head-to-head comparisons are lacking. This study aimed to examine the association of soluble tumor necrosis factor receptor type 1 (sTNFR1), fibroblast growth factor 21 (FGF-21), endocan, N-terminal pro-brain natriuretic peptide (NT-pro-BNP), and renal outcomes of patients with or without clinical signs of DKD. METHODS: A total of 312 patients were enrolled in a prospective observational study that excluded individuals with estimated glomerular filtration rates (eGFR) < 30 mL/min/1.73 m2. Composite renal outcomes included either a > 30% decline in eGFR and worsening albuminuria or both from consecutive tests of blood/urine during a 3.5-year follow-up period. RESULTS: Higher sTNFR1 and FGF-21, rather than endocan and NT-pro-BNP, levels were associated with renal outcomes but the significance was lost after adjusting for confounders. However, sTNFR1 levels ≥ 9.79 pg/dL or FGF-21 levels ≥ 1.40 pg/dL were associated with renal outcomes after adjusting for the confounders (hazard ration [HR] 2.76, 95% confidence interval [CI] 1.36-5.60, p = 0.005 for sTNFR1 level; HR 1.95, 95% CI 1.03-3.69, p = 0.03 for FGF-21 level). The combination of both levels exhibited even better association with renal outcomes than did either one alone (adjusted HR 4.45, 95% CI 1.86-10.65, p = 0.001). The results were consistent among patients with preserved renal function and normoalbuminuria. CONCLUSION: Both sTNFR1 and FGF-21 levels were associated with renal outcomes of in patients with type 2 diabetes, and the combination of the abovementioned markers exhibits better predictability.

[The expression of Notch pathway in nasal polyps and its correlation with the decreased Treg percentage and eosinophil infiltration].

Objective: To investigate the correlation between Notch pathway expression in nasal polyps and Treg percentage and Eos infiltration. Methods: Patients with chronic sinusitis and simple nasal septum deviation who received nasal endoscopic surgery in the Third Affiliated Hospital of Sun Yat-Sen University between November 2012 and August 2018 were selected and enrolled in CRS group and control group respectively. Nasal mucosa tissues were collected from 30 CRSsNP patients (14 males and 16 females aged from 18 to 63), 58 CRSwNP patients (38 males and 20 females aged from 18 to 65) and 29 patients (19 males and 10 females aged from 20 to 57), who underwent nasal endoscopic surgery for correction of simple nasal septum deviation. Hematoxylin-eosin(HE) staining was used to observe the infiltration of eosinophilic granulocytes in the tissues and to classify chronic sinusitis with polyps (CRSwNP) into eosinophilic chronic rhinosinusitis with nasal polyps (Eos-CRSwNP)and non-eosinophilic chronic rhinosinusitis with nasal polyps (Eos-CRSwNP). Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of Notch pathway receptors (Notch-l, 2, 3, 4) and their ligands (Jagded-l, Jagded-2, Delta-l, Delta-3and Delta-4) in the nasal mucosa of each group, as well as the expression of Th2 cytokines (IL-4, IL-5, IL-13), eosinophilic cationic protein (ECP)and the key transcription factor Foxp3 in Treg cells. Finally, flow cytometry was used to detect CD4+CD25+Foxp3+ Treg cells in nasal mucosa of each group. Results: Compared with controls, the expression of Th2 cytokines (IL-4, IL-5, IL-13) in CRSsNP and non-Eos-CRSwNP patients was the highest in Eos-CRSwNP (F=16.930,9.197,9.116, all P<0.05). Foxp3 had the lowest expression in Eos-CRSwNP patients and was lower than non-Eos-CRSwNP patients (F=2.780,P<0.05), and was negatively correlated with ECP (r=-0.326,P<0.05). Compared with controls, Eos-CRSwNP patients in CRSsNP patients and non-Eos-CRSwNP patients exhibited a significantly lower frequency of CD4+CD25+Foxp3+Treg cells (F=13.140, all P<0.01). The expression of Notch-l and Jagged-l in Eos-CRSwNP was significantly higher than that of the controls, CRSsNP patients and non-Eos-CRSwNP patients (F=5.953/F=6.380, P<0.05). In the nasal polyp group, the expression of Notch-l and Jagged-l showed significantly negative correlation with Foxp3 (r=-0.611/-0.346, all P<0.05), and positive correlation with Th2 cytokines (IL-4, IL-5, IL-13) and ECP, respectively (r=0.781/0.459,0.621/0.601,0.605/0.490,0.464/0.668, all P<0.05). There was no significant difference in the expression of receptor and ligand of the other Notch pathway among the groups. Conclusion: Abnormal activation of Notch-l/Jagged-l pathway may be involved in decreasing Treg ratio in Eos-CRSwNP, thereby promoting Th2 inflammatory response and Eosinophil infiltration.

[Expression and cellular provenance of interleukin 17A in non-eosinophilic chronic rhinosinusitis with nasal polyps].

Objective: To investigate the expression and cellular provenance of interleukin 17A (IL-17A) in non-eosinophilic chronic rhinosinusitis with nasal polyps (nECRSwNP), and to analyze the possible reasons for its different expression. Methods: Samples were collected from 14 patients with eosinophilic chronic rhinosinusitis with nasal polyps (ECRSwNP) and 28 patients with nECRSwNP, who underwent functional endoscopic sinus surgery in the Third Affiliated Hospital of Sun Yat-sen University from January 2017 to May 2018, including 33 males and 9 females, with the age ranging from 18 to 65 years old. Enzyme linked immune sorbent assay (ELISA) and flow cytometry were used to investigate the expression and cellular origins of IL-17A in the nasal tissue of ECRSwNP and nECRSwNP groups. Then the difference of quantity and differentiation ability of the major cells producing IL-17A between ECRSwNP and nECRSwNP groups were analyzed by flow cytometry. Finally, the expressions of IL-6, transforming growth factor-ß(TGF-ß), and IL-23, which were considered as the important factors in promoting Th17/Tc17 differentiation in CRSwNP and their correlation with IL-17A, were analyzed by ELISA. Statistical analysis was performed using IBM SPSS 20. Results: The IL-17A protein levels and IL-17A(+)lymphocyte percentages were higher in nECRSwNP group compared with that of the ECRSwNP group (158.56 (167.76) pg/ml (M(QR)) vs. 9.42 (11.33) pg/ml, 10.21%±1.54% (x±s) vs. 3.93%±0.80%, Z=2.95, t=3.62, all P<0.01). Tc17 cells (CD8(+)T cells producing IL-17A) and Th17 cells (CD4(+)T cells producing IL-17A) were major IL-17A producers in both ECRSwNP and nECRSwNP group. Further analysis revealed that there was no significant difference in quantity of CD8(+)and CD4(+)T cells between ECRSwNP and nECRSwNP group, but the differentiation ability about CD8(+)and CD4(+)T cells differentiating into Tc17 and Th17 in nECRSwNP group was stronger than that in ECRSwNP. The high expressions of IL-6 and TGF-ß, which were considered as the important factors in promoting Th17/Tc17 differentiation were also found in nECRSwNP group compared with ECRSwNP (56.07 (234.25) pg/ml vs. 8.27 (12.51) pg/ml, (5.44±0.34) pg/ml vs. (4.17±0.22) pg/ml, Z=2.426, t=2.29, all P<0.05). But the difference in expression of IL-23 was not significant difference between the two groups. Moreover, the expression of IL-17A showed significantly positive correlation with IL-6 (r=0.615, P=0.009). No positive correlation between IL-17A and TGF-ß or IL-23 was observed. Conclusions: The expression of IL-17A in nasal mucosa of nECRSwNP patients is significantly higher than that of ECRSwNP, which is due to the increase of expression and differentiation of Tc17/Th17 cells. IL-17A shows positive correlation with IL-6 in CRSwNP, which is the important factor in promoting Th17/Tc17 differentiation.

Case Report of an 18 Year Old Girl with Solid Pseudo-papillary Tumor of Pancreas.

Solid Pseudo-papillary Tumor (SPT) of the pancreas is a rare tumor which typically affects young women without any significant clinical symptoms. Solid Pseudopapillary Tumor usually shows an indolent behavior and only rare cases recur and/or metastasize after complete resection. Here is a case report of 18 years old girl who presented to our centre with complaints of severe epigastric pain and underwent pancreatic parenchyma saving surgery for a large pancreatic head mass. In conclusion, Solid Pseudo-papillary Tumor being a large tumor possess a low malignant potential in which R0 resection has excellent prognosis.

The effects of nasal irrigation with various solutions after endoscopic sinus surgery: systematic review and meta-analysis.

BACKGROUND: Nasal irrigation is commonly performed in patients with chronic rhinosinusitis after functional endoscopic sinus surgery. This study systematically assessed the clinical efficacy of nasal irrigation from the medical literature. METHODS: The PubMed, Embase and Cochrane Central Register of Controlled Trials databases were searched using a comprehensive strategy, limited to English-language articles, published from October 1971 to March 2017, and comprising human subjects. RESULTS: A total of 824 trials were identified, 5 of which, involving 331 participants, were included in this systematic review. After selection, only three trials were eligible for inclusion in a meta-analysis. Nasal irrigation using normal saline and various solutions was found to be effective in reducing symptom scores and endoscopic scores for chronic rhinosinusitis patients after functional endoscopic sinus surgery. Comparison of outcome measures, such as eosinophil count reduction, revealed that various solutions are more effective than normal saline alone; however, no statistical significance was found in terms of reduced symptom or endoscopic scores. CONCLUSION: Based on the current limited evidence, nasal irrigation is an effective therapy for chronic rhinosinusitis patients after functional endoscopic sinus surgery. However, when comparing various solutions with normal saline, no significant difference was found in symptom scores or endoscopic scores.

[Mutant Rad50 enhances killing effects of radiation on nasopharyngeal carcinoma cell line CNE1].

Objective: To investigate the killing effects of radiation and mutant Rad50 transfection on human nasopharyngeal carcinoma cell line CNE1. Methods: The experimental groups included: control group, Ad-Rad50-GFP group, Ad-EGFP group, irradiation group, Ad-Rad50-GFP combined with irradiation group, and Ad-EGFP combined with irradiation group. CNE1 cells were transfected with recombinant adenoviral vector Ad-Rad50-GFP carrying mutant Rad50 gene. The expressions of Mre11, Rad50, Nbs1, and relevant constituents composing MRN complex were detected by Western Blot. Neutral comet assay was used to detect the effect of mutant Rad50 on restoration process of DNA damage. Cell growth curve was used to evaluate the growth inhibition of CNE1 by mutant Rad50 and radiation. Results: Expressions of Mre11, Rad50, and Nbs1 in cells of Ad-Rad50-GFP group were less significantly than those in control group when irradiation was completed (0.48 vs 0.62, 0.42 vs 0.5, and 0.53 vs 0.69, respectively, P<0.05) and 24 hours after irradiation (0.41 vs 0.69, 0.46 vs 0.58, and 0.34 vs 0.78, respectively, P<0.05). The mean tail moment (MTM) in Ad-Rad50-GFP plus irradiation group was higher than that in irradiation group when irradiation was completed (16.06 vs 14.8, P<0.05), 24 hours after irradiation (58.23 vs 15.89, P<0.05) and 48 hours after irradiation: (45.12 vs 11.42, P<0.05). Seven days after irradiation, the cells in Ad-Rad50-GFP plus irradiation group was less than those in control group or irradiation group (both P<0.05). Conclusion: Mutant Rad50 enhances killing effects of radiation on nasopharyngeal carcinoma cell line CNE1.

[Psychopathological characteristics in patients with deviation of nasal septum: a preliminary analysis].

Objective: To investigate the psychopathological characteristics in patients with deviation of nasal septum. Methods: Between May 2015 and December 2015, fourty-four patients with deviated nasal septum and 37 patients with vocal cord polyp as control were included in this study. Psychological characteristics were evaluated by a series of questionnaire instruments including symptom checklist-90 (SCL-90), self-rating depression scale (SDS) and self-rating anxiety scale (SAS). Visual analogue scale (VAS) and rhinomanometry through front nostril were used to evaluate nasal symptom. The correlation between psychological characteristics and nasal symptom was evaluated. SPSS 20.0 software was used to analyze the data. Results: The SCL-90 score in nasal septal deviation group was 130.4±48.3. The total score and total average score of SCL-90 had no significant difference between nasal septal deviation group and the Chinese standard or control group(t value was 0.469, 0.112, 1.575, 1.564, respectively, all P>0.05). The scores of somatization, depression and anxiety factors in nasal septal deviation group were higher than control group (t value was 2.380, 2.133, 1.969, respectively, all P<0.05). The proportion of positive patients in these three factors between nasal septal deviation group and control group had significant differences (χ2 value was 11.585, 9.610, 5.429, respectively, all P<0.05). The scores of SDS and SAS in nasal septal deviation group were 46.0±10.6 and 43.0±10.2, which were higher than that in the Chinese standard and control group (t value was 5.342, 6.236, 1.476, 3.013, respectively, all P<0.05). There were 9 patients companying with depression or anxiety (20.5%, 20.5%, respectively) and 5 patients companying with depression and anxiety in nasal septal deviation group (11.4%). There were positive correlation not only between the scores of SDS and the depression factor of SCL-90 but also between the scores of SAS and the anxiety factor of SCL-90 (Z=0.415, P=0.005, Z=0.445, P=0.002, respectively). The scores of SDS and SAS had positive correlation (Z=0.392, P=0.008). The VAS score of nasal obstruction was 6.0±3.2. The rhinomanometry in inspiratory and expiratory phase were (0.202±0.140) kPa·S/cm3 and (0.230±0.161) kPa·S/cm3. Besides the positive correlation between the rhinomanometry in inspiratory phase and SDS (Z=0.332, P=0.045), the psychological scores, including SCL-90 score, depression, anxiety factors score, SAS and SDS, had no correlation with VAS scores and rhinomanometry (r value was -0.030, -0.052, -0.026, 0.107, 0.185, 0.066, 0.160, 0.203, respectively, all P>0.05). Conclusions: High prevalence of depression and anxiety is found in patients with deviation of nasal septum. The SCL-90 score is consistent with SDS and SAS. Besides the positive correlation between the rhinomanometry in inspiratory phase and SDS, the psychological scores (SCL-90 score, depression, anxiety factors score, SAS and SDS) have no correlation with VAS score and rhinomanometry.

T lymphocyte mediated lysis of mitomycin C treated Tenon's capsule fibroblasts.

AIMS: To evaluate the effect of T cell co-culture on mitomycin C treated and untreated Tenon's capsule fibroblasts. METHODS: IL-2 dependent allogeneic T cells were incubated over a monolayer of mitomycin C treated or control fibroblasts. Fibroblast numbers were evaluated by direct counts using phase contrast microscopy. To determine whether T cell mediated lysis was a consequence of MHC mismatch, co-culture experiments were repeated with autologous T cells. The effect of Fas receptor blockade was established by co-incubation with a Fas blocking (M3) antibody. RESULTS: T cell co-culture resulted in a dramatic reduction in fibroblast survival compared to mitomycin C treatment alone (p = 0.032). T cell killing required fibroblast/lymphocyte cell to cell contact and was observed in both allogeneic and autologous co-culture experiments. Fas blocking antibodies did not significantly inhibit T cell killing (p = 0.39). CONCLUSION: T cells augment mitomycin C treated fibroblast death in vitro. Similar mechanisms may contribute to the cytotoxic effect of mitomycin C in vivo and account for the largely hypocellular drainage blebs that are observed clinically.

NOS inhibitors decrease hypoxia-induced ATP reductions in respiring cerebrocortical slices.

BACKGROUND: Excess neuronal nitric oxide (NO) production might cause adenosine triphosphate loss and cellular damage in hypoxic brain parenchyma. 31P nuclear magnetic resonance spectroscopy was used to study hypoxic intracellular responses in perfused respiring cerebrocortical slices, in which NO scavenging by hemoglobin is absent, during NO synthase blockade and NO augmentation. METHODS: Adenosine triphosphate concentrations were monitored at 4.7 Tesla in respiring slices before, during, and after 60 min of hypoxia (oxygen tension < 5 mmHg). Slices were not treated or were pretreated with 27 microM L-nitroarginine methyl ester (L-NAME), 27 microM 7-nitroindozole (7-NI), or 27 microM L-nitroarginine. Nitrotyrosine:tyrosine ratios of slice extracts were measured using high-performance liquid chromatography. Cresyl violet-stained sections (2 microm) from random slices were examined histologically. RESULTS: After 60 min of hypoxia, adenosine triphosphate decreased to < or = 3, < or = 3, 65 +/- 6, and 25 +/- 4% of control in slices that were untreated or treated with L-nitroarginine, L-NAME, and 7-NI, respectively. After 120 min of hyperoxic recovery, adenosine triphosphate levels returned to control values in slices pretreated with L-NAME and 7-NI, but to only 30% of control in untreated or L-nitroarginine-treated slices. Nitric oxide donors administered during posthypoxic recovery partially antagonized the adenosine triphosphate recovery found with L-NAME and 7-NI. Nitric oxide synthase activity in slice homogenates, assayed via conversion of L-arginine to citrulline, was < or = 2% of control after all inhibitory treatments. The nitrotyrosine:tyrosine ratio increased by 52% in slices treated with 7-NI and by 200-300% in all other groups. Pretreatment with L-NAME and 7-NI reduced histologic evidence of cell swelling. CONCLUSION: Neuronal NO is associated with rapid adenosine triphosphate reductions and peroxynitrite formation in acutely hypoxic cerebrocortical slices.

Fructose-1,6-bisphosphate preserves adenosine triphosphate but not intracellular pH during hypoxia in respiring neonatal rat brain slices.

BACKGROUND: Fructose-1,6-bisphosphate (FBP) sometimes provides substantial cerebral protection during hypoxia or ischemia. 31P/1H nuclear magnetic resonance spectroscopy of cerebrocortical slices was used to study the effects of FBP on hypoxia-induced metabolic changes. In addition, 13C-labeled glucose was administered and 13C nuclear magnetic resonance spectroscopy was used to search for FBP-induced modulations in glycolysis and the pentose-phosphate pathway. METHODS: In each experiment, 80 slices (350 microm) obtained from ten 7-day-old Sprague-Dawley rat litter mates were placed together in a 20-mm nuclear magnetic resonance tube, perfused, and subjected to 30 min of hypoxia (PO2 < 3 mmHg). Nine experiments were performed, with n = 3 in each of three groups: (1) no treatment with FBP; (2) 60 min of prehypoxia treatment with FBP (2 mM); and (3) 60 min of posthypoxia treatment with FBP (2 mM). 31P/1H Interleaved nuclear magnetic resonance spectra at 4.7 T provided average adenosine triphosphate, intracellular pH, and lactate. Cresyl violet stains of random slices taken at predetermined time points were studied histologically. Some experiments had [2-13C]glucose in the perfusate. Slices from these studies were frozen for perchloric acid extraction of intracellular metabolites and studied with high-resolution 13C nuclear magnetic resonance spectroscopy at 11.75 T. RESULTS: With no pretreatment with FBP, hypoxia caused an approximately 50% loss of adenosine triphosphate, an approximately 700% increase in lactate, and a decrease in intracellular pH to approximately 6.4. Pretreatment with FBP resulted in no detectable loss of adenosine triphosphate, no increase in lactate, and minimal morphologic changes but did not alter decreases in intracellular pH. 13C Nuclear magnetic resonance spectra of extracted metabolites showed that pretreatment caused accumulation of [1-13C]fructose-6-phosphate, an early pentose-phosphate pathway metabolite. Posthypoxic treatment with FBP had no effects compared with no treatment. CONCLUSIONS: During severe hypoxia, pretreatment with FBP completely preserves adenosine triphosphate and almost completely preserves cell morphology but does not alter hypoxia-induced decreases in intracellular pH. Pretreatment also substantially augments the flux of glucose into the pentose-phosphate pathway.

Effect of brain, body, and magnet bore temperatures on energy metabolism during global cerebral ischemia and reperfusion monitored by magnetic resonance spectroscopy in rats.

To record brain temperature for comparison with rectal and temporalis muscle temperatures in preliminary studies before MR spectroscopy experiments, a thermistor was inserted into the basal ganglia in eight anesthetized, ventilated, and physiologically monitored rats. The rats were placed in an MR spectrometer and subjected to 60 min of global cerebral ischemia and 2 h of reperfusion without radiofrequency (RF) pulsing. Body temperature was maintained at 37.5-38.0 degrees C (normothermia) or 36.5-37.0 degrees C (mild hypothermia). Brain temperature during ischemia, which dropped to 31.9 +/- 0.3 (hypothermia) and 33.6 +/- 0.5 degrees C (normothermia), correlated with temporalis muscle temperature (r2 = 0.92) but not with body or magnet bore temperature measurements. Ischemia reduced brain temperature approximately 1.7 degrees C in rats subjected to mild hypothermia (1 degree reduction of body temperature). Parallel MR spectroscopy experiments showed no significant difference in energy metabolites between normothermic and hypothermic rats during ischemia. However, the metabolic recovery was more extensive 20-60 min after the onset of reperfusion in hypothermic rats, although not thereafter (P < 0.05). Mild hypothermia speeds metabolic recovery temporarily during reperfusion but does not retard energy failure during global ischemia in rats.

Adult rat brain-slice preparation for nuclear magnetic resonance spectroscopy studies of hypoxia.

BACKGROUND: When perfused neonatal brain slices are studied ex vivo with nuclear magnetic resonance (NMR) spectroscopy, it is possible to use 31P detection to monitor levels of intracellular adenosine triphosphate (ATP), cytosolic pH, and other high-energy phosphates and 1H detection to monitor lactate and glutamate. Adult brain slices of high metabolic integrity are more difficult to obtain for such studies, because the adult cranium is thicker, and postdecapitation revival time is shorter. A common clinical anesthesia phenomenon--loss of temperature regulation during anesthesia, with surface cooling and deep hypothermia, was used to obtain high-quality adult rat cerebrocortical slices for NMR studies. METHODS: Spontaneously breathing adult rats (350 g), anesthetized with isoflurane in a chamber, were packed in ice and cooled until rectal temperatures decreased to approximately 30 degrees C. An intraaortic injection of heparinized saline at 4 degrees C further cooled the brain to approximately 18 degrees C. Slices were obtained and then recovered at 37 degrees C in oxygenated medium. Interleaved 31P/1H NMR spectra were acquired continually before, during, and after 20 min of no-flow hypoxia (PO2 approximately 0 mmHg). Histologic (Nissl stain) measurements were made from random slices removed at different times in the protocol. Three types of pretreatment were compared in no-flow hypoxia studies. The treatments were: (1) hyperoxia; (2) hypercapnia (50% CO2); and (3) hypoxia, which was accomplished by washing the slices with perfusate equilibrated with 100% N2 and maintaining a 100% N2 gas flow in the air space above the perfusate. RESULTS: During hyperoxia, 31P NMR metabolite ratios were identical to those seen in vivo in adult brains, except that, in vitro, the Pi peak was slightly larger than in vivo. A lactate peak was seen in in vitro 1H spectra of slices after metabolic recovery from decapitation, although lactate is barely detectable in vivo in healthy brains. The in vitro lactate peak was attributed to a small population of metabolically impaired cells in an injury layer at the cut edge. NMR spectral resolution from the solenoidal coil exceeded that obtained in vivo in surface coil experiments. Phosphocreatine and ATP became undetectable during oxygen deprivation, which also caused a three- to sixfold increase in the ratio of lactate to N-acetyl-aspartate. Within experimental error, all metabolite concentrations except pHi recovered to control values within 2 h after oxygen restoration. Nissl-stained sections suggested that pretreatment with hypercapnia protected neurons from cell swelling during the brief period of no-flow oxygen deprivation. CONCLUSIONS: Perfused, respiring adult brain slices having intact metabolic function can be obtained for NMR spectroscopy studies. Such studies have higher spectral resolution than can be obtained in vivo. During such NMR experiments, one can deliver drugs or molecular probes to brain cells and obtain brain tissue specimens for histologic and immunochemical measures of injury. Important ex vivo NMR spectroscopy studies that are difficult or impossible to perform in vivo are feasible in this model.

Modulation of glutamate-induced intracellular energy failure in neonatal cerebral cortical slices by kynurenic acid, dizocilpine, and NBQX.

The severity and rapidity of acute, glutamate-induced energy failure were compared in live cerebral cortical slices. In each experiment 80 live cerebral cortical slices (350 microns thick) were obtained from neonatal Sprague-Dawley rats, suspended and perfused in a nuclear magnetic resonance (NMR) tube, and studied at 4.7 T with interleaved 31P/1H NMR spectroscopy. NMR spectra, obtained continually, were determined as 5-min averages. Slices were perfused for 60 min with artificial cerebrospinal fluid (ACSF) containing either glutamate alone or glutamate mixed with one of three glutamate-receptor antagonists: kynurenate, dizocilpine (MK-801), and 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(F)quinoxaline (NBQX). Dose-dependent decreases in high-energy phosphates were studied during glutamate exposure (0.5 to 10 mM), with and without antagonist protection. Energy recovery after glutamate exposures was measured during a 60-min washout with glutamate-free, antagonist-free ACSF. Reversible and irreversible energy failures were characterized by changes in intracellular pH, and by changes in relative concentrations of ATP, phosphocreatine (PCr), and inorganic phosphate. No changes were observed in intracellular levels of N-acetylaspartate and lactate. Some special studies were also done using R-(-)-2-amino-5-phosphonovaleric acid (100 microM) and tetrodotoxin (1 mM) to examine glutamate receptor specificity in this tissue model. Dizocilpine (150 microM) best ameliorated the energy failure caused by 2.0 mM glutamate. With dizocilpine the maximum ATP decrease was only 6 +/- 5%, instead of 35 +/- 7%.(ABSTRACT TRUNCATED AT 250 WORDS)

Relationship between extracellular neurotransmitter amino acids and energy metabolism during cerebral ischemia in rats monitored by microdialysis and in vivo magnetic resonance spectroscopy.

The time-course of changes in extracellular glutamate and energy metabolism during 30 or 60 min of complete cerebral ischemia and 60-90 min of reperfusion was investigated by microdialysis and magnetic resonance spectroscopy in parallel groups of rats. During the first 10 min of ischemia, adenosine triphosphate (ATP) was completely depleted, and lactate increased 10-fold; after 30 min, intracellular pH had decreased to 6.33 +/- 0.11. ATP and lactate did not change further between 30 and 60 min of ischemia. Glutamate increased 30-fold between 10 and 30 min of ischemia and continued to increase in the 60-min ischemia group. After 30 min of reperfusion, glutamate had returned to pre-ischemic levels in both groups. The cellular energy state recovered within 50-60 min after 30 min of ischemia but never returned to more than 60% of baseline values after 60 min of ischemia. The continued increase in extracellular glutamate after total depletion of ATP suggests that glutamate release during ischemia is not entirely energy dependent. Ca(2+)-independent glutamate release and failure of energy-dependent glutamate re-uptake mechanisms may result in continued increase in extracellular glutamate. The rapid normalization of extracellular glutamate after 30 and 60 min of ischemia despite differences in the recovery of energy metabolism suggests that the glutamate levels were reduced by an energy-independent mechanism, such as diffusion into the restored circulation.

Citrate alterations in primary and metastatic human prostatic adenocarcinomas: 1H magnetic resonance spectroscopy and biochemical study.

The objectives of this study were to quantitatively verify the low levels of citrate previously observed in primary human prostatic adenocarcinomas and to determine whether citrate was further reduced in metastatic prostatic cancer. This was accomplished by comparison of citrate concentrations of DU 145 xenografts (a poorly differentiated human prostatic adenocarcinoma cell line grown in nude mice) with concentrations in primary human adenocarcinomas. Following in vivo 1H NMR studies of DU 145 xenografts, citrate concentrations of DU 145 xenografts and surgically removed primary prostatic adenocarcinoma tissue were determined by quantitative high resolution 1H NMR and enzymatic assay. The most significant findings of this study were that citrate concentrations in primary human adenocarcinomas (3.74 +/- 0.19 mumol/g wet weight) were significantly lower than those observed for normal and benign hyperplastic (BPH) prostatic tissues. Furthermore there was a further ten-fold reduction of citrate associated with DU 145 xenografts (0.31 +/- 0.028 mumole/g wet weight) compared with primary prostatic cancer. DU 145 xenografts also exhibited higher levels of uridine diphosphosugars and choline containing metabolites relative to primary prostatic adenocarcinomas. These findings support the hypothesis that citrate is low in primary prostatic cancer and further reduced in metastatic disease.

Effect of glucose and confluency on phosphorus metabolites of perfused human prostatic adenocarcinoma cells as determined by 31P MRS.

A series of perfused cell 31P MRS studies were conducted using a well established human prostate adenocarcinoma cell line (DU 145) at different phases of growth, and exposed to varying glucose concentrations during growth. The spectral characteristics of perfused DU 145 cells were compared with the same cells grown in nude mice (xenografts). Perfused DU 145 cells had lower levels of inorganic phosphate and phosphocreative relative to in vivo nude mice xenografts. 31P MR spectra obtained from perfused cells at different phases of growth and exposed to varying glucose concentrations during grown suggest that increases in diphosphodiester levels are associated with high glucose concentrations and confluency. Perfused DU 145 cells grown in 5.5 mM glucose and harvested at log phase of growth best reflected the phosphorus MR spectra of the same cell line grown in nude mice.