Idiopathic pulmonary fibrosis (IPF) is a chronic interstitial pneumonia characterized by progressive accumulation of fibroblastic foci and destruction of the alveolar structure. Due to an incomplete understanding of the mechanism of the occurrence and progression of IPF, currently no effective means have been available for its early screening or treatment. With a poor overall prognosis, the patients with IPF have a median survival of only 2-4 years. In recent years, several studies have confirmed that dozens of molecules are involved in the development of IPF and can be used as potential biomarkers. These biomarkers play important roles in early diagnosis (such as SP-D, MMP-7, and osteopontin), prognostic evaluation (such as telomerase length, KL-6, mtDNA, HSP-70, LOXL2, CXCL13, miRNA, ICAM-1, and CCL18), and guiding treatment of IPF (such as TOLLIP rs3750920 genotype, SAMS score, and SP-D), and also provide potential therapeutic targets (such as TERT, TERR, RTEC, and PARN).
Amino Acid Oxidoreductases ; metabolism ; Biomarkers ; analysis ; Disease Progression ; Humans ; Idiopathic Pulmonary Fibrosis ; diagnosis ; therapy ; Intracellular Signaling Peptides and Proteins ; metabolism ; Prognosis

OBJECTIVETo determine the expressions of endothelial nitric oxide synthase traffic inducer (NOSTRIN) and endothelial nitric oxide synthase (eNOS) in the testis tissue of azoospermia patients, and investigate their correlation with the pathogenesis of azoospermia.

METHODSWe detected the expressions of NOSTRIN and NOSTRIN mRNA by immunohistochemistry and RT-PCR respectively, determined the activity of eNOS by spectrophotometry, and measured the stable metabolic end product NO, NO2- / NO3-, by nitrate reductase assay in the testis tissues of 17 patients with idiopathic azoospermia (the azoospermia group) and 10 normal men (the normal group).

RESULTSNOSTRIN and NOSTRIN mRNA were expressed in the spermatogonia, sertoli cells, stromal cells and vascular endothelial cells, more lowly in the azoospermia than in the normal group (0.312 +/- 0.076 versus 0.793 +/- 0.082, P < 0.01). The activity of eNOS was significantly increased in the idiopathic azoospermia patients ([33.727 +/- 3.58] U/mg) compared with the normal men ([17.69 +/- 3.84] U/mg) (P < 0.01). The level of NO2- / NO3- was significantly higher in the azoospermia than in the normal group ([48.56 +/- 8.49] micromol/L versus [25.37 +/- 9.61] micromol/L, P < 0.01). The expression of NOSTRIN showed a significant negative correlation with the activity of eNOS (r = -0.57, P < 0.01) as well as with the level of NO2- / NO3- (r = -0.61, P < 0.01) in the testis tissue of the idiopathic azoospermia patients.

CONCLUSIONThe expression of NOSTRIN is decreased, while the activity of eNOS and the level of NO2- / NO3- increased in the testis tissue of azoospermia patients, which may be associated with the pathogenesis of azoospermia.

Adult ; Azoospermia ; metabolism ; Humans ; Intracellular Signaling Peptides and Proteins ; metabolism ; Male ; Nitrates ; analysis ; Nitric Oxide Synthase Type III ; metabolism ; Nitrites ; analysis ; Spermatogenesis ; Testis ; metabolism

OBJECTIVETo investigate the influence and significance of gastric bypass surgery on hepatic gluconeogenesis in type 2 diabetic Goto Kakizaki(GK) rats.

METHODSForty GK rats were randomly divided into Roux-en-Y gastric bypass group(group A) and sham operation group(group B). Differences in glucose tolerance experiment(OGTT) at preoperative and postoperative 1, 2 and 4 weeks were compared and weight was recorded. Glycated hemoglobin levels were measured preoperatively and 4 weeks postoperatively. The animals were sacrificed 4 weeks after surgery and liver tissues were harvested to detect the relative expression of mRNA and protein of glucose 6 phosphatase(G-6-P) and phosphoenol pyruvate kinase(PEPCK) with RT-PCR and Western blot.

RESULTSFasting blood glucose levels were 6.5, 4.9, and 4.7 mmol/L in group A, and were 10.3, 10.4, and 12.5 mmol/L in group B, and the differences between two groups were statistically significant(P<0.05). The blood glucose level at 2 h after stomach lavage were 8.3, 6.4 and 5.5 mmol/L in group A, and were 21.4, 23.8 and 24.7 mmol/L in group B at postoperative 1, 2, 4 weeks, and the differences between two groups were statistically significant(P<0.05). The glycosylated hemoglobin at postoperative 4 weeks was(6.8±1.0)%, significantly lower than that in group B[(7.9±0.8)%, P<0.05]. Hepatic G-6-P and PEPCK mRNA relative expression at postoperative 4 weeks was reduced by 21.0% and 25.9% respectively as compared to group B, and the protein expression reduced as well. Immunohistochemistry showed that hepatic glycogen sedimentary in group A increased significantly.

CONCLUSIONThe relative mRNA and protein level of key enzymes of hepatic gluconeogenesis are significantly decreased after Roux-en-Y gastric bypass surgery and hepatic gluconeogenesis is reduced, which may be a potential mechanism of the decrease of blood glucose.

Animals ; Blood Glucose ; analysis ; Diabetes Mellitus, Experimental ; metabolism ; surgery ; Diabetes Mellitus, Type 2 ; metabolism ; surgery ; Gastric Bypass ; Gluconeogenesis ; Glucose-6-Phosphatase ; metabolism ; Glycated Hemoglobin A ; metabolism ; Intracellular Signaling Peptides and Proteins ; metabolism ; Liver ; enzymology ; Male ; Phosphoenolpyruvate Carboxykinase (GTP) ; metabolism ; Rats

OBJECTIVETo investigate the expression of doublecortin-like kinase 1(DCLK1)in gastric cancer and its prognostic significance.

METHODSThe expression of DCLK1 was examined by immunohistochemical staining of paraffin-embedded tumor specimens from 122 patients who underwent curative gastrectomy for gastric cancer at Peking Union Medical College Hospital between July 2002 and December 2006. Survival curves were described by the Kaplan-Meier method and compared by the Log-rank test. Univariate and multivariate analysis was performed with the Cox proportional hazard model.

RESULTSThe expression of DCLK1 in tumor cells was significantly upregulated in 51 of 122 patients. High expression of DCLK1 in tumor cells was strongly correlated with pN stage(P=0.029)and lymphovascular invasion(P=0.029). Kaplan-Meier analysis revealed that patients with DCLK1 high expression had a significantly lower 5-year overall survival(OS)rate than that of patients with DCLK1 low expression(39. 0% vs. 65. 8%, P=0.001), as well as a significantly lower 5-year disease-free survival(DFS)rate(37. 0% vs. 64. 5%, P=0.001). Univariate and multivariate analyses showed that DCLK1 expression(both P=0.036)was an independent factor for predicting OS and DFS rate.

CONCLUSIONSHigh expression of DCLK1 in gastric cancer cells is associated with pN stage and lymphovascular invasion. It may be a predictor for poor survival in patients undergoing surgery for gastric cancer.

Disease-Free Survival ; Gastrectomy ; Humans ; Intracellular Signaling Peptides and Proteins ; metabolism ; Kaplan-Meier Estimate ; Multivariate Analysis ; Prognosis ; Proportional Hazards Models ; Protein-Serine-Threonine Kinases ; metabolism ; Stomach Neoplasms ; diagnosis ; epidemiology ; metabolism

OBJECTIVETo investigate the effects of gastric bypass surgery(GBP) on hepatic phosphoenolpyruvate carboxykinase(PEPCK) mRNA expression in type 2 diabetic Goto-Kakizaki rats.

METHODSMale GK rats were randomized into three groups: gastric bypass surgery(n=10), sham operation with diet restriction(n=10), and sham operation alone(n=10). Liver specimens of GK rats were collected during the intraoperative period for self-control study and 8 weeks after surgery. Fasting blood glucose, food intake, and body weight were recorded before surgery and 1, 2, 4, 8 weeks after surgery. The expression of PEPCK mRNA was measured by real-time PCR.

RESULTSThe fasting plasma glucose level decreased from(17.6±2.1) mmol/L before surgery to(7.5±0.9) mmol/L 8 weeks after surgery in GBP group. The level of PEPCK mRNA decreased from 1.08±0.38 before surgery to 0.41±0.10 8 weeks after surgery, significantly lower than that in sham operation alone group(1.04±0.12)(P<0.01). The level of PEPCK mRNA in diet restriction group increased from 1.15±0.16 before surgery to 2.54±0.82 8 weeks after surgery(P<0.01). The expression of PEPCK mRNA in diet restriction was significantly higher than that in CBP group(P<0.01).

CONCLUSIONSGBP can significantly improve hyperglycemia in type 2 diabetic GK rat models, which may be associated with the decrease of hepatic PEPCK mRNA level.

Animals ; Blood Glucose ; analysis ; Diabetes Mellitus, Experimental ; enzymology ; surgery ; Diabetes Mellitus, Type 2 ; enzymology ; surgery ; Gastric Bypass ; Intracellular Signaling Peptides and Proteins ; genetics ; metabolism ; Liver ; enzymology ; Male ; Phosphoenolpyruvate Carboxykinase (GTP) ; genetics ; metabolism ; RNA, Messenger ; genetics ; Rats

Myristoylated alanine-rich C kinase substrate (MARCKS) is a widely distributed protein kinase C (PKC) substrate and has been implicated in actin cytoskeletal rearrangement in response to extracellular stimuli. Although MARCKS was extensively examined in various cell culture systems, the physiological function of MARCKS in the central nervous system has not been clearly understood. We investigated alterations of cellular distribution and phosphorylation of MARCKS in the hippocampus following kainic acid (KA)-induced seizures. KA (25 mg/kg, i.p.) was administered to eight to nine week-old C57BL/6 mice. Behavioral seizure activity was observed for 2 h after the onset of seizures and was terminated with diazepam (8 mg/kg, i.p.). The animals were sacrificed and analyzed at various points in time after the initiation of seizure activity. Using double-labeling immunofluorescence analysis, we demonstrated that the expression and phosphorylation of MARCKS was dramatically upregulated specifically in microglial cells after KA-induced seizures, but not in other types of glial cells. PKC alpha, beta I, beta II and delta, from various PKC isoforms examined, also were markedly upregulated, specifically in microglial cells. Moreover, immunoreactivities of phosphorylated MARCKS were co-localized in the activated microglia with those of the above isoforms of PKC. Taken together, our in vivo data suggest that MARCKS is closely linked to microglial activation processes, which are important in pathological conditions, such as neuroinflammation and neurodegeneration.
Up-Regulation/drug effects ; Time Factors ; Seizures/chemically induced/*metabolism ; Protein Kinase C-delta/analysis ; Protein Kinase C-alpha/analysis ; Protein Kinase C/*analysis ; Protein Biosynthesis/drug effects ; Phosphorylation/drug effects ; Microscopy, Confocal ; Microglia/cytology/drug effects/*metabolism ; Mice, Inbred C57BL ; Mice ; Membrane Proteins/*analysis/metabolism ; Kainic Acid/*toxicity ; Isoenzymes/analysis ; Intracellular Signaling Peptides and Proteins/*analysis/metabolism ; Immunohistochemistry ; Animals

OBJECTIVETo investigate the relationship between expressions of ING1 gene and genes of human telomerase reverse transcriptase (hTERT) and telomerase-associated protein 1 (hTP1) in human gliomas.

METHODSThe expressions of ING1 mRNA and p33(ING1) protein, hTERT mRNA and protein, and hTP1 mRNA and protein in seventy human glioma specimens with different malignant grades were studied using in situ hybridization and immunohistochemistry.

RESULTSAll of the 70 gliomas collected expressed hTP1 mRNA and protein and among them, 62 (88.6%) and 58 (82.9%) out of 70 expressed hTERT mRNA and protein respectively. The quantities of the four kinds of positive cells were correlated positively with one another (r = 0.758 - 0.882, P < 0.000 5), and all of them were significantly fewer in gliomas of WHO grade I - II than in grade III gliomas and the most in grade IV gliomas (P < 0.05 approximately 0.01). 66 (94.3%) and 62 (88.6%) out of 70 gliomas expressed ING1 mRNA and p33(ING1) protein respectively. The quantities of their positive cells were also correlated positively with each other (r = 0.831, P < 0.000 5), but the positive cells were more in gliomas of WHO grade I - II than in grade III gliomas and the fewest in grade IV gliomas (P < 0.01). The quantities of positive cells of ING1 mRNA and p33(ING1) protein were correlated negatively with those of hTERT mRNA and protein as well as hTP1 mRNA and protein respectively (r = -0.211 to -0.384, P < 0.05 approximately 0.001).

CONCLUSIONSThe results suggest that all of the parameters concerned are valuable in evaluating the biological behavior of gliomas. In glioma cells, overexpressions of hTERT and hTP1 genes might be significant in inhibiting the expression of ING1 gene. The abnormal expressions of the three genes play possibly the important roles in the development and malignant progression of gliomas.

Adolescent ; Adult ; Aged ; Carrier Proteins ; analysis ; genetics ; Cell Cycle Proteins ; DNA-Binding Proteins ; Female ; Genes, Tumor Suppressor ; Glioma ; genetics ; metabolism ; Humans ; Immunohistochemistry ; In Situ Hybridization ; Inhibitor of Growth Protein 1 ; Intracellular Signaling Peptides and Proteins ; Male ; Membrane Transport Proteins ; Middle Aged ; Nuclear Proteins ; Proteins ; genetics ; RNA, Messenger ; analysis ; Telomerase ; analysis ; genetics ; Tumor Suppressor Proteins

OBJECTIVETo construct the NLS(ING1)-GFP vector, transfer it into MRC-5 cells and establish a cell model expressing NLS (ING1)-GFP fusion protein.

METHODSFirstly, cDNA fragment of nuclear locating sequence (NLS) of inhibitor of growth-1 gene (ING1) was gained by RT-PCR and inserted into multi-clone site of pEGFP-C1 to construct the NLS (ING1)-GFP expression vector. Then the vector was used to transfect the MRC-5 cells to observe the subcellular signal localization of green fluorescence protein (GFP).

RESULTSWe successfully constructed the expressing vector of NLS (ING1)-GFP fusion protein. After transferring the fusion expressing vector into MRC-5 cells, we observed that green fluorescence signal located in the cell nucleus. However, the green fluorescence signal located in the cytoplasm in MRC-5 cells transfected with pEGFP-C1 control only expressing GFP.

CONCLUSIONIn living cells, physiologically p33 ING1b locates absolutely in nucleus. The p33(ING1b) NLS plays a decisive role in the transporting process of subcellular localization.

Base Sequence ; Cell Line ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; genetics ; metabolism ; Humans ; Inhibitor of Growth Protein 1 ; Intracellular Signaling Peptides and Proteins ; genetics ; metabolism ; Microscopy, Fluorescence ; Molecular Sequence Data ; Nuclear Proteins ; genetics ; metabolism ; Recombinant Fusion Proteins ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Analysis, DNA ; Transfection ; Tumor Suppressor Proteins ; genetics ; metabolism

The distinction between benign and malignant thyroid tumors is critical for the management of patients with thyroid nodules. We applied immunohistochemical staining for galectin-3, HBME-1, cytokeratin 19 (CK19), high molecular weight cytokeratin (HMWCK), cyclin D1 and p27(kip1) in 295 thyroid lesions to determine their diagnostic accuracy. The expression of all markers was significantly associated with differentiated thyroid carcinoma (DTC).The sensitivity for the diagnosis of DTC was 94.7% with galectin-3, 91.3% with HBME-1, and 90.3% with CK19. The specificities of these markers were 95.5%, 69.7%, and 83.1%, respectively. Combining these markers, co-expression of galectin-3 and CK19 or galectin-3 and HBME-1 was seen in 93.2% of carcinomas but in none of the benign nodules. Comparing follicular variant of papillary carcinoma (FVPC) with follicular carcinoma (FC), the expression of galectin-3, CK19, and HMWCK was significantly higher in FVPC. When comparing FC with FA, the expression of galectin-3 and HBME-1 was significantly higher in FC. These results suggest that 1) galectin-3 is a useful marker in the distinction between benign and malignant thyroid tumors, 2) the combined use of HBME-1 and CK19 can increase the diagnostic accuracy, and 3) the use of CK19 and HMWCK can aid in the differential diagnosis between PC and FC.
Adenocarcinoma, Follicular/diagnosis/metabolism ; Carcinoma, Papillary, Follicular/diagnosis/metabolism ; Cyclin D1/analysis ; Cyclin-Dependent Kinase Inhibitor p27/analysis ; Diagnosis, Differential ; Galectin 3/analysis ; Humans ; Immunohistochemistry ; Intracellular Signaling Peptides and Proteins/analysis ; Keratin-19/analysis ; Keratins/analysis ; Sensitivity and Specificity ; Thyroid Gland/chemistry/*pathology ; Thyroid Nodule/*diagnosis/metabolism ; Tumor Markers, Biological/*analysis

OBJECTIVETo investigate whether HIV-1 protease inhibitor nelfinavir alters the insulin stimulated phosphorylation of insulin signaling parameters in rat insulinoma INS-1 cells.

METHODSINS-1 cells were incubated with nelfinavir for 48 h and stimulated with 100 nmol/L insulin for 2 min. Immunoprecipitation and Western blot analysis of the insulin stimulated insulin receptor substrate (IRS)-1,-2 and Akt-Thr(308) phosphorylation were performed on cell lysates. Cytotoxic effects of nelfinavir were measured by cell count with trypan blue and MTT reduction test.

RESULTNelfinavir decreased insulin stimulated phosphorylation of IRS-2 and Akt-Thr(308) in a dose-dependent manner; for 10 micromol/L of nelfinavir, the decrease was 52% and 55%, respectively.

CONCLUSIONTreatment with nelfinavir might impair IRS-2-mediated signaling in pancreatic beta cells.

Animals ; Cell Line, Tumor ; Cell Survival ; drug effects ; HIV Protease Inhibitors ; pharmacology ; Insulin Receptor Substrate Proteins ; Insulinoma ; metabolism ; Intracellular Signaling Peptides and Proteins ; Islets of Langerhans ; drug effects ; physiology ; Nelfinavir ; pharmacology ; Pancreatic Neoplasms ; metabolism ; Phosphoproteins ; analysis ; physiology ; Protein-Serine-Threonine Kinases ; analysis ; Proto-Oncogene Proteins ; analysis ; Proto-Oncogene Proteins c-akt ; Rats ; Signal Transduction ; drug effects