Vitamin D Status During Puberty in French Healthy Male Adolescents

The vitamin D status was determined on one to four occasions either after summer (September–October) or after winter (March–April) in 175 male adolescents (13–17 years), resulting in 394 measurements of serum 25-hydroxyvitamin D (25(OH)D) and intact parathyroid hormone (iPTH). The subjects lived in a rural area to the north of Paris (49° N). After summer the 25(OH)D concentration was 58.5 ± 18.0 nmol/l (mean ± SD), while after winter it had fallen to 20.6 ±6.0 nmol/l (p= 0.0001). Meanwhile the iPTH concentration was 2.76 ± 0.97 pmol/l (mean ± SD) after summer and increased to 4.20 ± 1.21 pmol/l after winter (p= 0.0001). All the results were pooled and a nonlinear population model with random parameters was used to describe the relationship between serum iPTH and 25(OH)D. When the concentration of 25(OH)D was higher than 83 nmol/l, an iPTH mean ‘plateau’ level at 2.48 pmol/l was reached. When 25(OH)D concentrations fell below 83 nmol/l, the increase in iPTH concentration accelerates, and when the mean 25(OH)D concentration was equal to or lower than 10 nmol/l the mean iPTH level (4.97 pmol/l) was twice as high as the ‘plateau’ value. Received: 26 November 1998 / Accepted: 15 February 1999     

Global Skeletal Uptake of 99mTc-Methylene Diphosphonate (GSU) in Patients Affected by Endocrine Diseases: Comparison with Biochemical Markers of Bone Turnover

This study aimed to clinically validate the global skeletal uptake (GSU) of ^99mTc-methylene diphosphonate (^99mTc-MDP), and to compare it with a marker of bone formation (i.e. serum osteocalcin or OC) and an index of bone resorption (i.e. urinary deoxypyridinoline or U-DPD) in different endocrine disorders affecting the skeleton. We studied 29 female patients with thyrotoxicosis (TT), 27 with primary hyperparathyroidism (PHPT), 16 with acromegaly (AC), 15 with Cushing’s syndrome (CS), and altogether 110 healthy women matched for age, BMI and menstrual status. In all subjects total body digital scan images (TBDS) were acquired at 5 min and at 4 h after the administration of ^99mTc-MDP; the whole body retention (WBR) of the tracer was measured by counting two identical sets of rectangular ROIs, and GSU was subsequently calculated by drawing an irregular ROI on 4 h TBDS images. Serum OC was assessed by IRMA and urinary DPD by fluorometric detection after reverse phase high pressure chromatography. In TT patients GSU (40.0 ± 5.1 vs 36.5 ± 4.8%), OC (19.1 ± 11.8 vs 7.1 ± 2.9 mg/l) and U-DPD (62.4 ± 42.7 vs 19.5 ± 5.3 pmol/pmol) were significantly (p<0.01) higher than in controls. PHPT patients showed GSU (47.2 ± 6.6 vs 37.8 ± 5.3%), OC (38.6 ± 40.9 vs 8.2 ± 2.5 mg/l), and U-DPD (55.0 ± 51.3 vs 21.9 ± 6.1 pmol/pmol) values significantly (p<0.001) higher than controls. In CS patients, GSU (39.6 ± 6.4 vs 32.7 ± 3.5%; p<0.01) and U-DPD (22.8 ± 8.4 vs 16.5 ± 2.7 pmol/pmol; p<0.05) were higher, whereas OC (3.6 ± 2.4 vs 5.2 ± 1.9 mg/l; p<0,05) was lower than in controls. In AC patients, GSU (34.9 ± 5.3 vs 35.2 ± 3.4%) did not differ significantly from controls, whereas OC (16.8 ± 8.8 vs 6.9 ± 2.9 mg/l; p<0.001) and U-DPD (30.9 ± 13.6 vs 21.0 ± 5.7 pmol/pmol; p<0.01) were higher. Stepwise multivariate linear regression analysis was performed with disease activity, creatinine clearance, age, and years since menopause as predictor variables and GSU or OC or U-DPD as dependent variables. The significant partial regression coefficients (r) were: in TT, free triiodothyronine (fT3) with GSU (r = 0.37; p<0.005), Ln OC (r = 0.30; p = NS), Ln U-DPD (r = 0.76; p<0.0001), respectively; in PHPT, PTH with GSU (r = 0.74; p<0.001), Ln OC (r = 0.50; p<0.05), Ln U-DPD (r = 0.64; p<0.001); in CS Ln urinary free cortisol with OC (r = −0.68; p<0.001) and U-DPD (r = 0.66; p<0.05). Our data suggest that GSU could represent a valuable clinical tool for evaluating bone turnover rate in PHPT, CS, TT but not in AC. The behavior of GSU and OC and U-DPD is non-uniform in disorders characterized by a marked uncoupling between bone formation and resorption. Received: 26 December 2001 / Accepted: 31 May 2002     

N-Acetylcysteine and Desferoxamine Reduce Pulmonary Oxidative Stress Caused by Hemorrhagic Shock in a Porcine Model

Aim of the study: To investigate the pulmonary oxidative stress and possible protective effect of N-Acetylcysteine (NAC) and Desferoxamine (DFX)in a porcine model subjected to hemorrhagic shock. Materials and Methods: Twenty-one pigs were randomly allocated to Group-A (sham, n = 5), Group-B (fluid resuscitation, n = 8) and Group-C (fluid, NAC and DFX resuscitation, n = 8). Groups B and C were subjected to a 40-min shock period induced by liver trauma, followed by a 60-min resuscitation period. During shock, the mean arterial pressure (MAP) was maintained at 30–40 mmHg. Resuscitation consisted of crystalloids (35 mL/kg) and colloids (18 mL/kg) targeting to MAP normalization (baseline values ± 10%). In addition, Group-C received pretreatment with NAC 200 mg/kg plus DFX 2 g as intravenous infusions. Thiobarbituric Acid Reactive Substances (TBARS), protein carbonyls and glutathione peroxidase (GPx) activity were determined in lung tissue homogenates. Also, histological examination of pulmonary tissue specimens was performed. Results: TBARS were higher in Group-B than in Group-A or Group-C: 2.90 ± 0.47, 0.57 ± 0.10, 1.78 ± 0.47 pmol/μg protein, respectively (p < 0.05). Protein carbonyls content was higher in Group-B than in Group-A or Group-C: 3.22 ± 0.68, 0.89 ± 0.30, 1.95 ± 0.54 nmol/mg protein, respectively (p > 0.05). GPx activity did not differ significantly between the three groups (p > 0.05). Lung histology was improved in Group-C versus Group-B, with less alveolar collapse, interstitial edema and inflammation. Conclusion: NAC plus DFX prevented the increase of pulmonary oxidative stress markers and protein damage after resuscitated hemorrhagic shock and had beneficial effect on lung histology. NAC/DFX combination may be used in the multimodal treatment of hemorrhagic shock, since it may significantly prevent free radical injury in the lung.     

Glutathione depletion and increased apoptosis rate in human cystinotic proximal tubular cells

We have determined levels of glutathione (GSH), ATP, mitochondrial complex activity and apoptosis rate in proximal tubular cells (PTCs) exfoliated from urine in cystinotic (n=9) and control (n=9) children. Intracellular GSH was significantly depleted in cystinotic PTCs compared with controls (6.8 nmol GSH/mg protein vs 11.8 nmol GSH/mg protein; P<0.001), but there were no significant differences in mitochondrial complex activities or ATP levels under basal conditions. Cystinotic PTCs showed significantly increased apoptosis rate. After PTCs had been stressed by hypoxia, there was further depletion of GSH in cystinotic and control PTCs (2.4 nmol GSH/mg protein vs 7.2 nmol GSH/mg protein; P<0.001). Hypoxic stress led to increased complex I and complex IV activities in control but not in cystinotic PTCs. ATP levels were significantly reduced in cystinotic PTCs after hypoxic stress (12.2 nmol/mg protein vs 26.9 nmol/mg protein; P<0.001). GSH depletion occurs in this in vitro model of cystinotic PTCs, is exaggerated by hypoxic stress and may contribute to reduced ATP and failure to increase complex I/IV activities. Apoptotic rate is also increased, and these mechanisms may contribute to cellular dysfunction in cultured, human cystinotic PTCs.     

Wintertime Vitamin D Deficiency in Male Adolescents: Effect on Parathyroid Function and Response to Vitamin D3 Supplements

The first part of this study consisted of an 18 month follow-up of the vitamin D status and parathyroid function in a group of 54 French male adolescents, aged from 13 to 16 years old and all pupils of a jockey training school. During the 18 month period four samplings were made, one every 6 months. The first was during September of the first year, the second and third during March and October of the second year, and the last in March of the third year. Therefore we had two main periods: summer and winter. The summer 25-hydroxyvitamin D (25(OH)D) concentrations were higher (71.6 ± 19.9 and 52.4 ± 16.5 nmol/l) than the winter ones (20.4 ± 6.9 and 21.4 ± 6.1 nmol/l). Conversely, the winter intact parathyroid hormone (iPTH) serum levels (4.18 ± 1.18 and 4.11 ± 1.35 pmol/l) were higher than the summer ones (2.44 ± 0.82 and 2.71 ± 0.71 pmol/l). At the two winter time points the 25(OH)D concentrations were lower than 25 nmol/l (10 ng/ml) in 72% (2nd year) and 68% (3rd year) of the adolescents. In the second part of the study we tried a vitamin D₃ supplementation procedure designed to maintain the 25(OH)D and iPTH postsummer serum levels throughout the winter. Pairs of male adolescents matched for height, weight and Tanner pubertal stage were randomly assigned to either vitamin D₃ supplementation (2.5 mg, i.e., 100 000 IU) administered orally at three specific periods (end of September, November and January) or no vitamin D₃ treatment (control subjects). Blood was collected just before the first intake of vitamin D₃ and 2 months after the last intake (March). The control subjects had blood drawn at the same time points. In the vitamin D₃-treated subjects, the concentrations of 25 (OH)D (55.3 ± 11.5 nmol/l) and of iPTH (3.09 ± 1.16 pmol/l) in March and September (53.8 ± 12.3 nmol/l and 2.75 ± 1.26 pmol/l) were not significantly different. In the control subjects, March 25(OH)D levels (21.0 ± nmol/l were low, with values below 25 nmol/l in 78% of subjects, and iPTH concentrations (3.97 ± 1.08 pmol/l) were significantly (p<0.001) higher than in September (2.91 ± 0.81 pmol/l). The constant vitamin D wintertime deficiency and wintertime rise in iPTH in adolescent French males throughout puberty has been demonstrated. In adolescents with low dairy calcium intakes, the vitamin D₃ treatment was sufficient to maintain 25(OH)D concentrations at their summer levels throughout winter and to prevent an excessive wintertime rise in iPTH levels. Received: 6 February 2001 / Accepted: 9 May 2001     

Bone quality, as measured by trabecular bone score in patients with adrenal incidentalomas with and without subclinical hypercortisolism

Patients with adrenal incidentalomas (AIs) and subclinical hypercortisolism (SH) have increased risk of fracture independent of bone mineral density (BMD) and possibly due to reduced bone quality. The trabecular bone score (TBS) has been proposed as a index of bone microarchitecture. The aim of the study was to investigate TBS in AI. In 102 AI patients, SH was diagnosed in the presence of at least two of the following: (1) urinary free cortisol >70 µg/24 h (193.1 nmol/L); (2) cortisol after 1‐mg dexamethasone suppression test (1‐mg DST) >3.0 µg/dL (82.8 nmol/L); or (3) adrenocorticotropic hormone (ACTH) <10 pg/mL (<2.2 pmol/L). In patients and in 70 matched controls, BMD was measured at lumbar spine (LS) and femur (neck [FN] and total [FT]) by dual X‐ray absorptiometry and TBS was assessed in the region of LS‐BMD; BMD and TBS data were reported as Z‐scores. In patients, vertebral deformities were assessed by radiograph. Patients with SH (n = 34) had lower LS‐BMD (?0.31 ± 1.17), FT‐BMD (?0.29 ± 0.91), and TBS (?3.18 ± 1.21) than patients without SH (n = 68, 0.31 ± 1.42, p = 0.03; 0.19 ± 0.97, p = 0.01; ?1.70 ± 1.54, p < 0.0001, respectively) and controls (0.42 ± 1.52, p = 0.02; 0.14 ± 0.76, p = 0.02; ?1.19 ± 0.99, p < 0.0001, respectively). TBS was inversely correlated with 1‐mg DST (β = ?0.26, t = ?2.79, p = 0.006) regardless of age, LS‐BMD, body mass index (BMI), and gender. The presence of fracture was associated with low TBS alone (odds ratio [OR], 4.8; 95% confidence interval [CI], 1.85–12.42, p = 0.001) and with the cluster low TBS plus low LS‐BMD (OR, 4.37; 95% CI, 1.71–11.4, p = 0.002), after adjustment for age, BMI, and gender. Low TBS plus low LS‐BMD showed a good specificity (79%) for predicting fractures, whereas normal TBS (ie, > ?1.5) plus normal LS‐BMD high specificity (88.1%) for excluding fractures. Finally, TBS predicted the occurrence of a new fracture in 40 patients followed for 24 months (OR, 11.2; 95%CI, 1.71–71.41, p = 0.012) regardless of LS‐BMD, BMI, and age. In SH, bone quality, as measured by TBS, is altered. TBS is useful in detecting AI patients at risk of fractures. © 2012 American Society for Bone and Mineral Research.     

Changes in the Biochemical Profiles of Mid-Cervically Located Adenosine A1 Receptors After Repeated Theophylline Administration in Adult Rats

Abstract

Background/Objective: Adenosine A1 receptors localized in the phrenic motoneurons (PMNs), where the axons of the descending bulbospinal respiratory make synaptic contacts, may be involved in theophylline- induced respiratory-related activity in rats. The objective of this study was to characterize the biochemical profiles of adenosine A., receptors in 2 groups of rats: (a) naive and (b) theophylline-treated (3-day oral administration).

Methods: Biochemical binding characteristics of adenosine A1 receptors in the C3 to C5 (PMN) of adult rats were assessed in naive (n = 6) and theophylline-treated animals (n = 6) using [3H]-DPCPX (10 pmol/L to 30 nmol/L), the specific adenosine A1 receptor antagonist in saturation-binding assays. Competition assays used theophylline as the competing ligand (20 mmol/L to 20 pmol/L), and protein concentration was determined with the Bradford assay using a range of standards (0.016-1.0 mg/mL).

Results: In saturation-binding assays in naive animals, the A1 receptor was characterized by a single binding site with Smax and Kd values of 256.00 ± 32.13 fmoi/mg protein and 2.89 ± 0.45 nmol/L, respectively. Analysis of the isotherm in theophylline-treated animals showed 1 site with Smax and Kd values of 219.00 ± 26.3 fmol/mg protein and 0.60 ± 0.21 nmol/L, respectively, and a second site characterized by Smax and Kd values of 492.6 ± 3.15 fmol/mg protein and 14.09 ± 2.06 n mol/L, respectively.

Conclusions: Theophylline administration revealed 2 binding sites on receptors (characterized by the specific adenosine A1 antagonist, [3H]-DPCPX) located in the vicinity of phrenic motoneurons (C3-C5). Alteration of the receptor profiles after theophylline may underlie the respiratory-related actions of the drug.     

Hepatocellular Glycogen in Alleviation of Liver Ischemia-Reperfusion Injury During Partial Hepatectomy

Background Temporary occlusion of liver blood supply for complex liver operation is common in liver surgery. However, hepatic vascular occlusion will undoubtedly impair liver function. This study was designed to elucidate the effect of hepatocellular glycogen in alleviation of liver ischemia-reperfusion injury during hepatic vascular occlusion for partial hepatectomy. Methods Fifty-seven patients were randomly divided into an experimental group (n = 29) and a control group (n = 28). In the experimental group, patients were given high-concentration glucose intravenously during 24 h before the operation. The hepatic lesion was resected after portal triad clamping in the two groups. Noncancer liver tissue was biopsied to measure hepatic tissue ATP content and change of malondialdehyde (MDA) and superoxide dismutase (SOD). Liver function of all patients was assessed by using an automatic biochemical analysis apparatus before the operation and the first and fifth days after operation. Results The mean hepatic vascular occlusion time in the experimental group was 19.21 ± 4.54 min and in the control group it was 21.04 ± 5.11 min. Hepatic tissue ATP content of the experimental group was significantly higher than that of the control group at the end of hepatic vascular occlusion (2.15 ± 0.39 μmol/g wet tissue vs. 1.33 ± 0.44, p < 0.01) and at the point of 1-h reperfusion (2.19 ± 0.29 μmol/g wet tissue vs. 1.57 ± 0.35, p < 0.01). There was significant difference in SOD activity between the two groups at the end of hepatic vascular occlusion (130.69 ± 30.49 NU/mg pr vs. 97.83 ± 26.23, p < 0.01) and at the point of 1-h reperfusion (139.55 ± 39.88 NU/mg pr vs. 114.74 ± 25.93, p < 0.01). Significant difference was shown in MDA content between the two groups at the end of hepatic vascular occlusion (3.02 ± 0.30 nmol/mg pr vs. 3.99 ± 0.49, p < 0.01) and at the point of 1-h reperfusion (3.81 ± 0.69 nmol/mg pr vs. 5.75 ± 1.17, p < 0.01). In addition, the liver function of the experimental group was significantly better than that of the control group the first and fifth days after the operation (p < 0.01). Conclusions Abundant intracellular glycogen may reduce liver ischemia-reperfusion injury caused by hepatic vascular occlusion. It is beneficial to give a large amount of glucose before a complex liver operation during which temporary occlusion of hepatic blood flow is necessary.     

Analysis of the oxidative catabolism of retinoic acid in rat Dunning R3327G prostate tumors

We studied the enzymatic characteristics of the oxidative catabolism of retinoic acid (RA) and its inhibition by liarozole-fumarate in homogenates of rat Dunning R3327G prostate tumors. Homogenates of rat liver were used as reference material. Both tumor and liver homogenates were able to catabolize retinoic acid. HPLC analysis revealed only very polar metabolites in tumors, while in the liver both metabolites with intermediate polarity and more polar metabolites were found. Kinetic analysis of retinoic acid catabolism showed a K_m of 1.7 ± 0.7 μM and a V_max of 4.2 ± 4.4 pmol polar RA metabolites/mg protein/hr for Dunning G tumor homogenates. In liver homogenates a K_m value of 4.3 ± 0.5 μM and a V_max value of 290 ± 120 pmol polar RA metabolites/mg protein/hr were obtained. Liarozole-fumarate inhibited retinoic acid catabolism in Dunning tumors and liver with IC₅₀ values of 0.26 ± 0.16 μM and 0.14 ± 0.05, respectively. The results suggest that rat Dunning R3327G tumors are able to metabolize retinoic acid in a manner similar to that found in rat liver but with a lower metabolizing capacity. © 1996 Wiley-Liss, Inc.     

The effect of prostate cancer and antianrogenic therapy on lipid peroxidation and antioxidant systems

In living organism, excessive free radicals oroxidative damage which occur as a result of deficient antioxidant defensive mechanisms by the effect of endogenous and exogenous factors, influences especially developmental steps of chemically induced cancers [1, 2]. In our study, plasma malondialdehyde level (MDA) as an indicator of lipid peroxidation, erythrocyte glutathione (GSH) level as an indicator of antioxidant state, glutathione reductase (GSH-Red), glutathione peroxidase (GSH-Px), glutathione-S-transferase (GST) as an antioxidant enzymes and plasma vitamin E level were detected in patients with prostate cancer (21 males; age, 69.4 ± 4.8 years) before and after three months of antiandrogenic therapy with goserelin acetate as luteinizing hormone releasing hormone (LHRH) analogue. Healthy people evaluated as a control group (20 males; age, 63.7 ± 3.9). Erythrocyte GSH levels, the activities of GSH-Red and GSH-Px and plasma vitamin E levels were found significantly low in patients with prostate cancer when compared with the healthy subjects (p < 0.01, p < 0.05, p ≤ 0.001 and p ≤ 0.001 respectively). Plasma MDA level and erythrocyte GST activity of patient group were significantly higher than the levels of control group (p ≤ 0.001 and p ≤ 0.001 respectively). After antiandrogenic therapy erythrocyte GSH level, GSH-Red, GSH-Px activity and plasma vitamin E level were found unchanged. Significant decrease in plasma MDA level and significant increase in erythrocyte GST activity were detected in patient group (p < 0.05 and p ≤ 0.01 respectively). The study has revealed the shift in the oxidant-antioxidant balance towards oxidative state in patients with metastatic prostate cancer. Our results showed that antiandrogenic therapy increased in GST activity, decreased in lipid peroxidation. This revised version was published online in August 2006 with corrections to the Cover Date.     

Regulation of amino acid arginine transport by lipopolysaccharide and nitric oxide in intestinal epithelial IEC-6 cells

As a precursor for nitric oxide (NO) synthesis and an immune-enhancing nutrient, amino acid L-arginine plays a critical role in maintaining intestine mucosal integrity and immune functions in sepsis. However, the relationship between intestinal arginine transport and NO synthesis in sepsis remains unclear. In the present study, we investigated the effects of lipopolysaccharide (LPS) and NO on the arginine transport in cultured rat intestinal epithelial IEC-6 cell. Near-confluent IEC-6 cells were incubated with LPS (0-50 μg/ml) in serum-free Dulbecco’s modified Eagles’s medium, in the presence and absence of the NO donor sodium nitroprusside (SNP, 0–500 μmol/L) and the inducible nitric oxide synthase (iNOS) inhibitor N-ω-nitro-L-arginine (NNA, 0–1000 μmol/L) for various periods of time (0-48 hours). Arginine transport activity, arginine transporter CAT1 mRNA and protein levels were measured with transport assay, Northern blot analysis, and Western blot analysis, respectfully. LPS increased arginine transport activity in a time- and dose-dependent fashion. Prolonged incubation of LPS (24 hours, 25 μg/ml) resulted in a 3-fold increase of arginine transport activity (control: 28 ±5; LPS: 92 ±20 pmol/mg/ min, P < 0.05), with the System y⁺ as the predominant arginine transport system, and a 2-fold increase of System y⁺ CAT1 mRNA and transporter protein levels (P < 0.05). LPS increased the arginine transport System y⁺ maximal velocity (V_max, control: 1484 ±180; LPS: 2800 ±230 pmol/mg/min, P<0.05) without affecting the transport affinity (K_m, control: 76 ±8; LPS: 84 ±14 μmol/L, p = NS). The LPSinduced arginine transport activity was blocked by sodium nitroprusside (SNP) (control: 25 ±6; LPS: 97 ±26*; SNP: 22 ±0.4⁺; LPS+SNP: 33 ±10.3⁺ pmole/mg/min, *P < 0.01 and ⁺p = NS, compared with control). In contrary, the LPS-induced arginine transport activity was further augmented by NNA (control: 18 ±3.2; LPS: 59 ±2.7*; NNA: 26.3 ±5.8; LPS + NNA: 127 ±18⁺ pmol/mg/min; *P < 0.01 compared with control and ⁺P < 0.01 compared with control or LPS). LPS-stimulates arginine transport activity in IEC-6 cells via a mechanism that involves increase of transport System y⁺ mRNA levels and transporter protein levels. The LPS-stimulated arginine transport activity is regulated by the availability of nitric oxide. Presented at the Forty-Sixth Annual Meeting of The Society for Surgery of the Alimentary Tract, Chicago, Illinois, May 14–18, 2005 (oral presentation). This work was supported in part by The Society for Surgery of Alimentary Tract Career Development Award (M.P.) and National Institute of Diabetes and Digestive and Kidney Disease Grant DK-62165 (M.P.).     

Effect of systemic insulin on protein kinetics in postoperative cancer patients

Background: Cancer cachexia is a significant cause of postoperative morbidity and mortality in patients with tumors of the upper gastrointestinal tract. Standard parenteral nutrition (TPN) has failed to alter this. The anabolic effect of insulin has been well documented, and its positive effect on protein economy in cancer patients has been recently demonstrated. This study examines the effect of high-dose insulin and parenteral nutrition on protein kinetics in postoperative cancer patients. Methods: Eleven patients underwent surgery for pancreatic, esophageal, or gastric carcinoma. Postoperatively, patients received standard TPN for 4 days (1 g/kg/day amino acids, 1,000 kcal/day dextrose, 100 g/day lipid), and hyperinsulinemic parenteral nutrition for 4 days (same as standard TPN plus 1.44 U/kg/day regular human insulin) in a crossover design. All patients received both treatments, and the order of treatment was determined randomly. Euglycemia was maintained during insulin infusion via a variable 30% dextrose infusion. Patients underwent protein metabolic studies after each treatment period and rates of whole body and skeletal muscle protein synthesis, breakdown, and net balance were determined by radioisotopic tracer methods using¹⁴C-leucine and³H-phenylalanine. Results: Compared with standard TPN (STD), hyperinsulinemic TPN (INS) resulted in a significant increase in skeletal muscle protein synthesis (INS: 52.04±10.22 versus STD: 26.06±6.71 nmol phe/100 g/min, p<0.05) and net balance of protein (INS: 7.75±4.61 versus STD: −15.10±6.44 nmol phe/100 g/min, p<0.01), but no difference in skeletal muscle protein breakdown (INS: 44.29±11.54 versus STD: 41.17±5.89 nmol phe/100 g/min). Whole-body net balance of protein also significantly increased with insulin-based TPN, compared with standard TPN (INS: 0.04±0.05 versus STD: −0.08±0.07 μmol leu/kg/min, p<0.05), but no difference in whole-body protein synthesis (INS: 2.52±0.15 versus STD: 2.49±0.15 μmol leu/kg/min) or whole-body protein breakdown (INS: 2.48±0.16 versus STD: 2.58±0.19 μmol leu/kg/min) was observed. Patients received significantly more calories during the hyperinsulinemic TPN period than during the standard TPN period. There was no difference in total, essential, or branched-chain amino acids, and no difference in serum free fatty acids, triglycerides, or cholesterol was observed between the two treatment periods. Conclusion: High-dose insulin in conjunction with hypercaloric parenteral nutrition causes improved skeletal muscle protein synthesis, skeletal muscle protein net balance, and whole-body protein net balance compared with standard TPN in postoperative cancer patients. Results of this study were presented at the 46th Annual Meeting of The Society of Surgical Oncology, Los Angeles, California, March 18–21, 1993. Dr. Pearlstone is recipient of the Resident's Clinical Prize.     

Effects of trimetazidine on the Akt/eNOS signaling pathway and oxidative stress in an in vivo rat model of renal ischemia-reperfusion

Renal ischemia reperfusion (I/R) injury, which occurs during renal surgery or transplantation, is the major cause of acute renal failure. Trimetazidine (TMZ), an anti-ischemic drug, protects kidney against the deleterious effects of I/R. However its protective mechanism remains unclear. The aim of this study is to examine the relevance of Akt, endothelial nitric oxide synthase (eNOS), and hypoxia inducible factor-1α (HIF-1α) on TMZ induced protection of kidneys against I/R injury. Wistar rats were subjected to 60?min of warm renal ischemia followed by 120?min of reperfusion, or to intraperitoneal injection of TMZ (3?mg/kg) 30?min before ischemia. In sham operated group renal pedicles were only dissected. Compared to I/R, TMZ treatment decreased lactate dehydrogenase (845?±?13 vs. 1028?±?30?U/L). In addition, creatinine clearance and sodium reabsorption rates reached 105?±?12 versus 31?±?11?μL/min/g kidney weight and 95?±?1 versus 68?±?5%, respectively. Besides, we noted a decrease in malondialdehyde concentration (0.33?±?0.01 vs. 0.59?±?0.03?nmol/mg of protein) and an increase in glutathione concentration (2.6?±?0.2 vs. 0.93?±?0.16?µg GSH/mg of protein), glutathione peroxidase (95?±?4 vs. 61?±?3?µg GSH/min/mg of protein), and superoxide dismutase (25?±?3 vs. 11?±?2?U/mg of protein) and catalase (91?±?12 vs. 38?±?9?μmol/min/mg of protein) activities. Parallely, we noted a significant increase in p-Akt, eNOS, nitrite and nitrate (18?±?2 vs. 8?±?0.1?pomL/mg of protein), HIF-1α (333?±?48 vs. 177?±?14?µg/mg of protein) and heme oxygenase-1 (HO-1) levels regarding I/R. TMZ treatment improves renal tolerance to warm I/R. Such protection implicates an activation of Akt/eNOS signaling pathway, HIF-1α stabilization and HO-1 activation.     

The effects of zinc supplementation on wound healing and metabolic status in patients with diabetic foot ulcer: A randomized,double‐blind,placebo‐controlled trial

This study was performed to determine the effects of zinc supplementation on wound healing and metabolic status in patients with diabetic foot ulcer. The current randomized, double‐blind, placebo‐controlled trial was conducted among 60 patients (aged 40–85 years old) with grade 3 diabetic foot ulcer. Participants were randomly divided into two groups (30 participants in each group) to take either 220 mg zinc sulfate supplements containing 50 mg elemental zinc or placebo daily for 12 weeks. After the 12‐week intervention, compared with the placebo, zinc supplementation was associated with significant reductions in ulcer length (?1.5 ± 0.7 vs. ?0.9 ± 1.2 cm, p = 0.02) and width (?1.4 ± 0.8 vs. ?0.8 ± 1.0 cm, p = 0.02). In addition, changes in fasting plasma glucose (?40.5 ± 71.0 vs. ?3.9 ± 48.5 mg/dl, p = 0.02), serum insulin concentration (?8.0 ± 15.4 vs. +1.1 ± 10.3 µIU/ml, p = 0.009), homeostasis model of assessment‐estimated insulin resistance (?3.9 ± 7.1 vs. +0.8 ± 5.9, p = 0.007), the quantitative insulin sensitivity check index (+0.01 ± 0.03 vs. ?0.002 ± 0.02, p = 0.04) and HbA1c (?0.5 ± 0.8 vs. ?0.1 ± 0.5%, p = 0.01) in the supplemented group were significantly different from the changes in these indicators in the placebo group. Additionally, significant increases in serum HDL‐cholesterol (+4.1 ± 4.3 vs. +1.1 ± 5.1 mg/dl, p = 0.01), plasma total antioxidant capacity (+91.7 ± 213.9 vs. ?111.9 ± 188.7 mmol/L, p < 0.01) and total glutathione (+68.1 ± 140.8 vs. ?35.0 ± 136.1 µmol/L, p = 0.006), and significant decreases in high sensitivity C‐reactive protein (?20.4 ± 24.6 vs. ?6.8 ± 21.3 µg/ml, p = 0.02) and plasma malondialdehyde concentrations (?0.6 ± 0.9 vs. ?0.2 ± 0.7 µmol/L, p = 0.03) were seen following supplementation with zinc compared with the placebo. Zinc supplementation for 12 weeks among diabetic foot ulcer patients had beneficial effects on parameters of ulcer size and metabolic profiles.     

Tumor Necrosis Factor-α, Interleukin-1β and Nitric Oxide: Induction of Liver Megamitochondria in Prehepatic Portal Hypertensive Rats

Abstact It has been shown that portal hypertension in the rat causes microvesicular hepatocytic fatty infiltration. Formation of megamitochondria (MG) is one of the most prominent alterations in steatosis. Because nitric oxide (NO), tumor necrosis factor-α (TNFα), and interleukin-1β (IL-1β) impair mitochondrial function, these mediators have been studied in prehepatic portal hypertensive rats to verify their coexistence with MG and therefore with steatosis. Male Wistar rats were divided into two groups: a control group (n = 7) and a group with partial portal vein hgation (n = 19) at 6 weeks of evolution. TNFα and IL-1β were quantified in liver by enzyme-linked immunosorbent assay, and NO was measured in the portal vein, suprahepatic inferior vena cava, and infrahepatic inferior vena cava by the Griess reaction. In portal hypertensive rats, the-serum concentration of NO of hepatic origin increases (132.10 ± 34.72 vs. 52.44 ± 11.32 nmol/ml; p < 0.001), as do TNF-α (2.02 ± 0.20 vs. 1.12 ± 0.43 μmol/mg protein) and IL-1β (18.95 ± 2.59 vs. 5.48 ± 1.70 μmol/mg protein) (p = 0.005) in the liver. The most frequent hepatic histologic findings are the presence of MG (p < 0.001), steatosis, and hyperplasia. An increase in hepatic release of NO, TNFα and IL-Iβ with MG formation is produced in rats with portal hypertension. Therefore these proinflammatory mediators and this morphologic mitochondrial alteration could both be involved in the etiopathogenesis of steatosis.     

The influence of laparotomy and laparoscopy on tumor growth in a rat model

Background: The effects of laparotomy and laparoscopy with different gases on subcutaneous and intraperitoneal tumor growth have not been evaluated yet. Methods: Tumor growth of colon adenocarcinoma DHD/K12/TRb was measured in rats after laparotomy, laparoscopy with CO₂ or air, and in control group. Cell kinetics were determined after incubation with carbon dioxide or air in vitro and tumor growth was measured subcutaneously and intraperitoneally after surgery in vivo. Results: In vitro, tumor cell growth increased significantly after incubation with air and CO₂. In vivo, intraperitoneal tumor weight was increased after laparotomy (1,203 ± 780 mg) and after laparoscopy with air (1,085 ± 891 mg) and with CO₂ (718 ± 690 mg) compared to control group (521 ± 221 mg) (p < 0.05). Subcutaneous tumor growth was promoted after laparotomy (71 ± 35 mg) and even more after laparoscopy with air (82 ± 45 mg) and CO₂ (99 ± 55 mg) compared to control group (36 ± 33 mg). Conclusions: Insufflation of air and CO₂ promote tumor growth in vitro. In vivo, intraperitoneal tumor growth seems to be promoted primarily by intraperitoneal air and subcutaneous tumor growth by CO₂. Received: 7 November 1996/Accepted: 3 December 1996     

Factors Predicting Nutritional Derangements in Patients with Obstructive Jaundice: Multivariate Analysis

Patients with obstructive jaundice (OJ) that requires surgery often have malnutrition associated with increased perioperative morbidity. This study investigated the factors influencing nutritional derangements in these patients. A series of 46 OJ patients were investigated prospectively (28 malignant tumors, 18 benign obstructions). A nutritional risk index of < 83.5 was used to define protein-calorie malnutrition. Liver function, cholecystokinin (CCK), tumor necrosis factor-α (TNFα), and endotoxin levels were determined. A multivariate analysis was performed, and an obstructive jaundice malnutrition index (OJMI) was obtained. Altogether, 22 (48%) OJ patients had malnutrition (33% with benign obstructions, 57% with malignant disease). Malnourished patients had higher serum bilirubin levels (258 ± 120 vs. 154 ± 62 mmol/L; p= 0.005), longer duration of jaundice (16 ± 9 vs. 9 ± 5 days; p= 0.03), and higher plasma levels of CCK (4.0 ± 1.3 vs. 1.7 ± 1.0 pmol/L; p= 0.005), alanine aminotransferase (ALT) (226 ± 209 vs. 187 ± 161 UI/L; p= 0.01), endotoxin (15 ± 10 vs. 6.5 ± 7.0 EU/L; p= 0.007), and TNFα (69 ± 82 vs. 23 ± 15 pg/ml; p= 0.008) than those without malnutrition. However, only serum bilirubin, CCK, ALT, and patient age were predictors for malnutrition by multivariate analysis. Malnutrition might be expected (95% confidence interval) in patients older than 68 years with increased bilirubin (> 290 mmol/L) and ALT (> 210 UI/L) levels that corresponded with an OJMI > 55. It was concluded that nutritional alterations in patients with obstructive jaundice were determined by the intensity of the biliary obstruction correlated with increased plasma CCK levels as well as with liver dysfunction and patient age.     

Elevation of glutathione and related enzyme activities in high-grade and metastatic extremity soft tissue sarcoma

Background: Glutathione is a free radical scavenger implicated in the chemoresistance of certain tumors. As treatment with chemotherapy has added little to improved survival in adult soft tissue sarcoma and little is known concerning the mechanisms of chemoresistance in sarcoma, we studied concentrations of glutathione (nmol/mg protein) and activities of-glutamylcysteine synthetase (GCS; nmol/mg protein/h) and-glutamyl transpeptidase (GGTP; U/mg protein) in extremity soft tissue sarcoma. Methods and Results: Tumor specimens (n=65) were frozen in liquid nitrogen at the time of resection. Fourteen low-grade tumors, 40 high-grade tumors, and 11 pulmonary metastases were analyzed. Glutathione concentrations and GGTP activity were significantly lower in low-grade (3.97±0.7 nmol/mg protein and 1.07±0.2 U/mg protein) than in high-grade (8.98±1.2 nmol/mg protein, p<0.001; 2.10±0.4 U/mg protein, p<0.002) tumors and pulmonary metastases (10.05±1.8 nmol/mg protein, p<0.008; 3.14±2.8 U/mg protein, p<0.04). While GCS activity was lower in low-grade (0.81±0.3 nmol/mg protein/h) than high-grade (1.49±0.5 nmol/mg protein/h) tumors and pulmonary metastases (1.03±0.2 nmol/mg protein/h), these differences were not significant. In those patients with a high-grade tumor presenting with a local recurrence, glutathione levels were higher in those patients who had received preoperative doxorubicin-based chemotherapy (9.25±1.7 nmol/mg protein; n=7) than in those who had no preoperative chemotherapy (4.71±3.1 nmol/mg protein; n=4, p=0.08). Conclusions: In extremity soft tissue sarcoma, glutathione concentration and GGTP activity are significantly elevated in patients with high-grade and metastatic sarcomas. In addition, there is a trend for increased glutathione levels in tumors previously exposed to doxorubicin-based chemotherapy. Glutathione may play a role in soft tissue sarcoma chemoresistance.This work was presented at the 49th Annual Cancer Symposium of the Society of Surgical Oncology, Atlanta, GA, U.S.A., March 21–24, 1996.     

Different concentrations of two small stress proteins, αB crystallin and HSP27 in human urological tumor tissues

Concentrations of two small stress proteins, αB crystallin and the 27-kDa heat shock protein (HSP27) were quantitated in tissues of the human normal genitourinary system and their tumors. Levels of HSP27 in renal cell carcinomas (mean ± SE: 1450 ± 262 ng/mg protein, n = 15) were significantly higher than in normal kidney (the cortex: 540 ± 99 ng/mg protein, n = 13; the medulla: 600 ± 106 ng/mg protein, n = 13) while those of αB crystallin tended to be increased without statistical significance. These findings were similar to those previously reported for renal cell tumors chemically induced in rats. Concentrations of αB crystallin in prostatic carcinoma tissues (410 ± 129 ng/mg protein, n = 10) were also significantly higher than in benign prostatic hyperplasia (54 ± 12 ng/mg protein, n = 14), whereas αB crystallin levels in testicular tumors including seminomas (2.1 ± 0.8 ng/mg protein, n = 11) and non-seminomas (5.2 ± 2.3 ng/mg protein, n = 9) were significantly lower than in normal testicular tissues (29.7 ± 6.2 ng/mg protein, n = 5). Both αB crystallin and HSP27 could be immunohistochemically localized in the normal kidney and renal cell carcinoma tissues. Received: 1 December 1997 / Accepted: 16 June 1998     

Prednisolone Co-Administered with Losartan Confers Renoprotection in Patients with IgA Nephropathy

Background.Treatment options for progressive IgA nephropathy are limited. Methods. We performed a small, randomized controlled trial to evaluate the effects of prednisolone (PSL, 30 mg/dL, gradually tapered to 5 mg/dL over two years) plus 50 mg/day of losartan (LST, an angiotensin II receptor blocker) or PSL alone on IgA nephropathy. We separated 38 patients (age, 33 ± 11 years; creatinine clearance, 103 ± 31 mL/min; proteinuria, 1.6 ± 0.5 g/day) into two groups that were treated with either PSL plus LST or PSL alone, and compared the proteinuria and creatinine clearance after two years. Baseline and histopathological data did not significantly differ between the two groups. Results. Two years of treatment in both groups significantly decreased proteinuria compared with baseline, and PSL plus LST (from 1.6 ± 0.6 to 0.3 ± 0.1 g/day, p < 0.05) was more effective than PSL alone (from 1.6 ± 0.3 to 0.5 ± 0.1 g/day, p < 0.05). Creatinine clearance in both groups was similar at the start of study but significantly differed at the end of the study (PSL plus LST, 104.3 ± 36.4 to 100.4 ± 38.9 mL/min; PSL alone, 103.4 ± 28.5 to 84.8 ± 34.3 mL/min, p < 0.05). Conclusions. Combined therapy with PSL plus LST appears to be more effective than PSL alone in reducing proteinuria and protecting renal function in patients with IgA nephropathy.