3-nitrotyrosine and Atrophy

Calcineurin inhibitors (CNIs) are effective immunosuppressive agents for the successful treatment of childhood steroid-resistant nephrotic syndrome (SRNS). Because these patients require long-term treatment, the identification of early markers of CNI-induced nephrotoxicity (CNIN) is imperative. The monitoring of CNI trough levels, serum creatinine, and glomerular filtration rate is not an accurate marker of CNIN. The present study has been undertaken to identify early markers of CNIN in SRNS patients. Twenty-four pediatric SRNS patients were included with paired renal biopsies, before initiation (time zero biopsy) and at least 1 year after CNI therapy (protocol renal biopsy) with standard dosage. Semiquantitative morphologic grading of the histologic features was done for assessing CNIN. Immunohistochemical markers for oxidative stress (nitrotyrosine [NT]), fibrogenic cytokine (transforming growth factor β1 [TGF-β1]), and endothelial injury (endothelial nitric oxide synthase [eNOS]) were evaluated. In addition, ultrastructural study was done to assess mitochondrial injury in endothelial and tubular epithelial cells. The protocol renal biopsies in comparison with time zero biopsies showed significant increase in glomerulosclerosis, juxtaglomerular apparatus hyperplasia, tubular atrophy, interstitial fibrosis, arteriolar hyalinosis, and smooth muscle vacuolization (P < .05 - P < .001). Significantly higher immunoexpression of eNOS (91.6%), NT (71%), and TGF-β1 (87.5%) was noted in posttreatment biopsies. Mean mitochondrial injury grade among post-CNI cases in endothelial cells and proximal tubular cells was 2.28 and 1.4, whereas in pre-CNI, it was 0.28 and 0.27, respectively. We propose that immunohistochemical overexpression of NT, eNOS, and TGF-β1 is an early marker of CNIN. Endothelial and proximal tubular mitochondrial injury may play an important role in the pathogenesis of CNIN.

Topics: Adolescent; Atrophy; Biomarkers; Biopsy; Calcineurin Inhibitors; Child; Child, Preschool; Early Diagnosis; Endothelial Cells; Female; Fibrosis; Humans; Hyperplasia; Immunohistochemistry; Immunosuppressive Agents; Infant; Kidney; Kidney Diseases; Kidney Tubules, Proximal; Male; Mitochondria; Nephrotic Syndrome; Nitric Oxide Synthase Type III; Oxidative Stress; Predictive Value of Tests; Time Factors; Transforming Growth Factor beta1; Treatment Outcome; Tyrosine; Up-Regulation

Since epidemiologic studies suggest that tobacco smoke toxins, e.g. the nicotine-derived nitrosamine ketone (NNK) tobacco-specific nitrosamine, can be a co-factor in alcohol-related brain disease (ARBD), we examined the independent and additive effects of alcohol and NNK exposures on spatial learning/memory, and brain insulin/IGF signaling, neuronal function and oxidative stress.. Adolescent Long Evans rats were fed liquid diets containing 0 or 26% caloric ethanol for 8 weeks. During weeks 3-8, rats were treated with i.p. NNK (2 mg/kg, 3×/week) or saline. In weeks 7-8, ethanol groups were binge-administered ethanol (2 g/kg; 3×/week). In week 8, at 12 weeks of age, rats were subjected to Morris Water Maze tests. Temporal lobes were used to assess molecular indices of insulin/IGF resistance, oxidative stress and neuronal function.. Ethanol and NNK impaired spatial learning, and NNK ± ethanol impaired memory. Linear trend analysis demonstrated worsening performance from control to ethanol, to NNK, and then ethanol + NNK. Ethanol ± NNK, caused brain atrophy, inhibited insulin signaling through the insulin receptor and Akt, activated GSK-3β, increased protein carbonyl and 3-nitrotyrosine, and reduced acetylcholinesterase. NNK increased NTyr. Ethanol + NNK had synergistic stimulatory effects on 8-iso-PGF-2α, inhibitory effects on p-p70S6K, tau and p-tau and trend effects on insulin-like growth factor type 1 (IGF-1) receptor expression and phosphorylation.. Ethanol, NNK and combined ethanol + NNK exposures that begin in adolescence impair spatial learning and memory in young adults. The ethanol and/or NNK exposures differentially impair insulin/IGF signaling through neuronal growth, survival and plasticity pathways, increase cellular injury and oxidative stress and reduce expression of critical proteins needed for neuronal function.

Topics: Acetylcholinesterase; Animals; Atrophy; Dinoprost; Drug Synergism; Ethanol; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Insulin; Insulin Resistance; Male; Maze Learning; Nicotine; Nitrosamines; Oxidative Stress; Phosphorylation; Protein Carbonylation; Rats; Receptor, IGF Type 1; Ribosomal Protein S6 Kinases, 70-kDa; Signal Transduction; Somatomedins; Spatial Learning; tau Proteins; Temporal Lobe; Tyrosine

Induction of inducible nitric oxide synthase (iNOS) may be involved in carcinogenesis of the stomach, because nitric oxide (NO) derived from iNOS can exert DNA damage and post-transcriptional modification of target proteins. In the present study, we investigated the correlation between endoscopic findings and iNOS mRNA expression/NO-modified proteins in the gastric mucosa.. Fifty patients were prospectively selected from subjects who underwent upper gastrointestinal chromoendoscopy screening for abdominal complaints. The Helicobacter pylori (H. pylori) status of patients was determined by anti-H. pylori IgG antibody levels. We classified the mucosal area of the fundus as F0, fine small granules; F1, edematous large granules without a sulcus between granules; F2, reduced-size granules with a sulcus between granules; and F3, irregular-sized granules with extended sulcus between granules. Gastritis was graded using the visual analog scale of the Updated Sydney System. The expression of interleukin (IL)-8 and iNOS mRNA was assayed in gastric biopsy specimens by reverse transcription-polymerase chain reaction. NO-modified proteins were analyzed by Western blotting using novel monoclonal antibodies against nitrotyrosine.. A total of 91.7% (11/12) of the F0 group was H. pylori-negative, whereas 94.7% (36/38) of the F1-3 groups was H. pylori-positive. Spearman's analysis showed good correlation between the endoscopic grading and the score of chronic inflammation (r=0.764) and glandular atrophy (r=0.751). The expression of IL-8 mRNA was significantly increased in F1, F2, and F3 cases compared with the F0 group, with no significant differences among them. iNOS mRNA was significantly increased in the F3 group compared with the other groups, with increased nitration of tyrosine residues of proteins.. The proposed classification by chromoendoscopy is useful for screening patients for atrophic and iNOS-expressing gastric mucosa with NO-modified proteins in H. pylori-associated atrophic gastric mucosa.

Topics: Antibodies, Bacterial; Atrophy; Biomarkers; Gastric Mucosa; Gastritis, Atrophic; Gastroscopy; Helicobacter Infections; Helicobacter pylori; Humans; Interleukin-8; Nitric Oxide; Nitric Oxide Synthase Type II; Prospective Studies; Proteins; RNA, Messenger; Severity of Illness Index; Tyrosine