Study of the Effect of Hyperbaric Oxygen Therapy on the Viability of Dorsal Cutaneous Flaps in Tobacco-Exposed Rats.

BACKGROUND: Smoking causes a threefold increase in the risk of surgical complications in flaps. Hyperbaric oxygen therapy (HBOT) increases the viability of chronic wounds. However, there are few studies concerning the effects of HBOT on surgical flaps in patients who smoke. This study aimed to analyze the effect of HBOT on the viability of cutaneous flaps in tobacco-exposed rats. METHODS: Twenty Wistar rats were exposed to tobacco smoke for two months. Following this period, all animals underwent a dorsal cutaneous flap (3 × 10 cm) surgery and were divided into two groups: control (n = 10) and HBOT (n = 10). HBOT was performed in seven daily sessions (2 ATA, 90 min). After seven days, the animals were euthanized. The outcomes were total area, viable area, viable area/total area rate, analysis of dermal appendages and angiogenesis (hematoxylin-eosin), and gene expression analysis of iNOS and VEGF-a biomarkers. RESULTS: The HBOT group showed an increase in viable area compared with the control group (84% versus 47%, p = 0.009, respectively). The HBOT group also showed an increase in appendage units (1.69 ± 0.54 versus 1.87 ± 0.58, p = 0.04) and angiogenesis density (1.29 ± 0.45 versus 1.82 ± 0.64, p < 0.001) compared to the control group. There was a difference between the control and HBOT groups in iNOS levels (0.926 ± 1.4 versus 0.04 ± 0.1 p = 0.002, respectively). However, this study did not show a difference between the groups concerning the gene expression of VEGF-a. CONCLUSION: The use of hyperbaric oxygen therapy increased the viability of cutaneous flaps in tobacco-exposed rats and decreased iNOS mRNA levels; however, it did not change VEGF-a levels. Level of Evidence IV This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266.

Diabetes consequences in the fetus liver of the non-obese diabetic mice.

DM type 1 (T1D) incidence is increasing around 3% every year and represents risks for maternal and fetal health. The objective of this study was to explore the effects of diabetes on fetus liver cells in non-obese diabetic (NOD) mice. Hyperglycemic NOD (HNOD), normoglycemic NOD (NNOD) and BALB/c females were used for mating, and the fetus livers were collected at 19.5 gestation day (gd). HNOD group had reduced fetal weight (989.5±68.32 vs 1290±57.39 mg BALB/c, P<0.05) at 19.5 gd and higher glycemia (516.66±28.86 mg dl-1, P<0.001) at both 0.5 gd and 19.5 gd compared to other groups. The protein expression of albumin (ALB) was significantly reduced in HNOD group (0.9±0.2 vs 3.36±0.36 NNOD P<0.01, vs 14.1±0.49 BALB/c P<0.001). Reduced gene expression of ALB (1.34±0.12 vs 5.53±0.89 NNOD and 5.23±0.71 BALB/c, P<0.05), Hepatic Nuclear Factor-4 alpha (HNF-4α) (0.69±0.1 vs 3.66±0.36 NNOD, P<0.05) and miR-122 (0.27±0,10 vs 0.88±0.15 NNOD, P<0.05) was present in HNOD group. No difference for alpha-Fetoprotein (AFP) and gene expression was observed. In conclusion, our findings show the impacts of T1D on the expression of ALB, AFP, HNF-4α and miR-122 in fetus liver cells by using NNOD and HNOD mice.

Sorafenib prevents liver fibrosis in a non-alcoholic steatohepatitis (NASH) rodent model

Liver fibrosis occurring as an outcome of non-alcoholic steatohepatitis (NASH) can precede the development of cirrhosis. We investigated the effects of sorafenib in preventing liver fibrosis in a rodent model of NASH. Adult Sprague-Dawley rats were fed a choline-deficient high-fat diet and exposed to diethylnitrosamine for 6 weeks. The NASH group (n=10) received vehicle and the sorafenib group (n=10) received 2.5 mg·kg-1·day-1 by gavage. A control group (n=4) received only standard diet and vehicle. Following treatment, animals were sacrificed and liver tissue was collected for histologic examination, mRNA isolation, and analysis of mitochondrial function. Genes related to fibrosis (MMP9, TIMP1, TIMP2), oxidative stress (HSP60, HSP90, GST), and mitochondrial biogenesis (PGC1α) were evaluated by real-time quantitative polymerase chain reaction (RT-qPCR). Liver mitochondrial oxidation activity was measured by a polarographic method, and cytokines by enzyme-linked immunosorbent assay (ELISA). Sorafenib treatment restored mitochondrial function and reduced collagen deposition by nearly 63% compared to the NASH group. Sorafenib upregulated PGC1α and MMP9 and reduced TIMP1 and TIMP2 mRNA and IL-6 and IL-10 protein expression. There were no differences in HSP60, HSP90 and GST expression. Sorafenib modulated PGC1α expression, improved mitochondrial respiration and prevented collagen deposition. It may, therefore, be useful in the treatment of liver fibrosis in NASH.

Sorafenib prevents liver fibrosis in a non-alcoholic steatohepatitis (NASH) rodent model.

Liver fibrosis occurring as an outcome of non-alcoholic steatohepatitis (NASH) can precede the development of cirrhosis. We investigated the effects of sorafenib in preventing liver fibrosis in a rodent model of NASH. Adult Sprague-Dawley rats were fed a choline-deficient high-fat diet and exposed to diethylnitrosamine for 6 weeks. The NASH group (n=10) received vehicle and the sorafenib group (n=10) received 2.5 mg·kg(-1)·day(-1) by gavage. A control group (n=4) received only standard diet and vehicle. Following treatment, animals were sacrificed and liver tissue was collected for histologic examination, mRNA isolation, and analysis of mitochondrial function. Genes related to fibrosis (MMP9, TIMP1, TIMP2), oxidative stress (HSP60, HSP90, GST), and mitochondrial biogenesis (PGC1α) were evaluated by real-time quantitative polymerase chain reaction (RT-qPCR). Liver mitochondrial oxidation activity was measured by a polarographic method, and cytokines by enzyme-linked immunosorbent assay (ELISA). Sorafenib treatment restored mitochondrial function and reduced collagen deposition by nearly 63% compared to the NASH group. Sorafenib upregulated PGC1α and MMP9 and reduced TIMP1 and TIMP2 mRNA and IL-6 and IL-10 protein expression. There were no differences in HSP60, HSP90 and GST expression. Sorafenib modulated PGC1α expression, improved mitochondrial respiration and prevented collagen deposition. It may, therefore, be useful in the treatment of liver fibrosis in NASH.

Differential gene expression profiles of hepatocellular carcinomas associated or not with viral infection

Chronic hepatitis B (HBV) and C (HCV) virus infections are the most important factors associated with hepatocellular carcinoma (HCC), but tumor prognosis remains poor due to the lack of diagnostic biomarkers. In order to identify novel diagnostic markers and therapeutic targets, the gene expression profile associated with viral and non-viral HCC was assessed in 9 tumor samples by oligo-microarrays. The differentially expressed genes were examined using a z-score and KEGG pathway for the search of ontological biological processes. We selected a non-redundant set of 15 genes with the lowest P value for clustering samples into three groups using the non-supervised algorithm k-means. Fisher’s linear discriminant analysis was then applied in an exhaustive search of trios of genes that could be used to build classifiers for class distinction. Different transcriptional levels of genes were identified in HCC of different etiologies and from different HCC samples. When comparing HBV-HCC vs HCV-HCC, HBV-HCC/HCV-HCC vs non-viral (NV)-HCC, HBC-HCC vs NV-HCC, and HCV-HCC vs NV-HCC of the 58 non-redundant differentially expressed genes, only 6 genes (IKBKâ, CREBBP, WNT10B, PRDX6, ITGAV, and IFNAR1) were found to be associated with hepatic carcinogenesis. By combining trios, classifiers could be generated, which correctly classified 100 percent of the samples. This expression profiling may provide a useful tool for research into the pathophysiology of HCC. A detailed understanding of how these distinct genes are involved in molecular pathways is of fundamental importance to the development of effective HCC chemoprevention and treatment.

Differential gene expression profiles of hepatocellular carcinomas associated or not with viral infection.

Chronic hepatitis B (HBV) and C (HCV) virus infections are the most important factors associated with hepatocellular carcinoma (HCC), but tumor prognosis remains poor due to the lack of diagnostic biomarkers. In order to identify novel diagnostic markers and therapeutic targets, the gene expression profile associated with viral and non-viral HCC was assessed in 9 tumor samples by oligo-microarrays. The differentially expressed genes were examined using a z-score and KEGG pathway for the search of ontological biological processes. We selected a non-redundant set of 15 genes with the lowest P value for clustering samples into three groups using the non-supervised algorithm k-means. Fisher's linear discriminant analysis was then applied in an exhaustive search of trios of genes that could be used to build classifiers for class distinction. Different transcriptional levels of genes were identified in HCC of different etiologies and from different HCC samples. When comparing HBV-HCC vs HCV-HCC, HBV-HCC/HCV-HCC vs non-viral (NV)-HCC, HBC-HCC vs NV-HCC, and HCV-HCC vs NV-HCC of the 58 non-redundant differentially expressed genes, only 6 genes (IKBKbeta, CREBBP, WNT10B, PRDX6, ITGAV, and IFNAR1) were found to be associated with hepatic carcinogenesis. By combining trios, classifiers could be generated, which correctly classified 100% of the samples. This expression profiling may provide a useful tool for research into the pathophysiology of HCC. A detailed understanding of how these distinct genes are involved in molecular pathways is of fundamental importance to the development of effective HCC chemoprevention and treatment.

Detection of codon 12 mutation in the K-ras oncogene in pancreatic tumors.

Mutations at codons 12, 13, or 61 of the H-ras, K-ras, and N-ras have been detected in human neoplasias by a variety of techniques. Some of these techniques are very sensitive and can detect K-ras mutation in 90% of the cases of pancreatic adenocarcinomas. We analyzed 11 samples of pancreatic adenocarcinoma, three samples of pancreatic mucinous cystadenoma, and two samples without tumors in formalin-fixed paraffin embedded tissue sections. K-ras mutations at codon 12 were detected by a two-step PCR-enriched technique in all the samples of pancreatic adenocarcinoma, but not in cystadenoma or control samples. This technique may be useful for early detection of pancreatic cancer.

[Physiopathology of lung injury in acute pancreatitis]. / Fisiopatologia da lesão pulmonar na pancreatite aguda.

Clinically detectable signs of lung injury develop in up to 50-70 percent of patients with acute pancreatitis. Despite that, the physiopathology of the lung injury associated with acute pancreatitis is unclear so far. Pulmonary edema is the main respiratory complication in acute pancreatitis. Increased permeabilities of the pulmonary endothelial and alveolar epithelial barriers are the causes of the pulmonary edema. Several factors have been regarded as the cause to pulmonary edema: release of pancreatic-derived proteolytic enzymes, oxygen-free radicals, phospholipase A2, free fat acids, tumor necrosis factor, platelet activating factor, arachidonic acid metabolites and pulmonary embolization. Understanding lung injury physiopathology enables physicians to a better therapeutic approach of the patients with acute pancreatitis. The aim of this paper is to expose the theories that explain the pancreatic-derived lung injury.

[Protective effect of reduction of pancreatic enzyme content on the pulmonary disease induced by acute pancreatitis]. / Efeito protetor da redução do conteúdo enzimático do pâncreas na lesão pulmonar da pancreatite aguda.

Lung injury develop in up to 50-70 percent of patients with acute pancreatitis. Cerulein in physiological doses reduces the rate mortality of pancreatitis by decreasing the enzyme content of the pancreas. In order to assess the effect of acute reduction of pancreatic enzyme content on the pancreatitis pulmonary injury, pancreatitis was induced in Wistar rats by intraductal injection of 5 per cent sodium taurocholate: group I, with pancreatitis; group II, pancreatitis after decreasing pancreatic enzyme content; group III, control group. Dye Evans blue was used to evaluate the lung injury. The pancreatitis pulmonary injury was smaller in group II than group I (P < 0.05) but it was similar to control group (group III). In conclusion, it is speculated that the reduction of the pancreatic enzyme content reduces the pancreatitis pulmonary injury by decreasing the quantity of pancreatic enzymes reaching the systemic circulation.

[Lung injury in acute pancreatitis. Influence of pancreatic enzymes reduction]. / Lesão pulmonar na pancreatite aguda. Influência da redução do conteúdo enzimático do pâncreas.

UNLABELLED: A previous report has show that cerulein in physiological doses reduces the rate mortality of pancreatitis by decreasing the enzyme content of the pancreas. Clinically detectable signs of lung injury develop in up to 50-70 percent of patients with acute pancreatitis. The aim of the present study was to assess the effect of acute reduction of pancreatic enzyme content on the pancreatitis pulmonary injury. Experimental haemorrhagic pancreatitis was induced by intraductal injection of 5 per cent sodium taurocholate in two groups of Wistar rats: group I (pancreatitis) and group II (pancreatitis after decreasing pancreatic enzyme content). Dye Evans blue was used to evaluate the lung injury. The degree of histologically observed lesions were similar in both groups, but the pulmonary lesion was smaller in group II than group I (p < 0.05). IN CONCLUSION: 1) pancreatitis' pulmonary lesion may be related with pancreatic enzymes that reach the blood stream and 2) the reduction of the pancreatic enzyme content has a beneficial effect on acute pancreatitis and reduces its pulmonary injury.

[Effect of omeprazole administration on pancreatic content of enzymes in rats]. / Efeito da administração de omeprazol sobre o conteúdo pancreático de enzimas em ratos.

Gastric chloride acid plays an important role in pancreatic enzyme synthesis and secretion, mediated by cholecystokinin released in the duodenum. This study was designed to evaluate the influence of gastric acid suppression by omeprazole on pancreatic enzyme content. Eighteen male Wistar rats (180-220 g) were divided in two groups: I--control and II--omeprazole. Animals received by intraduodenal catheter 3 doses of 0.5 ml saline solution (NaCl 0.9%)--Group I or 5 mumol/Kg of omeprazole solution--Group II at 24 h intervals. All animals, after an overnight fasting period, were killed 3 h after the last dose. Serum amylase and pancreatic tissue content of protein, trypsinogen, elastase, lipase and phospholipase A2 were determinated. Omeprazole treated animals (group II) showed statistically significant lower levels of serum amylase and pancreatic trypsinogen content (P < 0.05). We believe that this effect is related with acid secretion suppression by omeprazole and that it may be mediated by cholecystokinin.

Reduction of pancreatic enzyme content and mortality in experimental acute pancreatitis in rats.

A previous report has shown that undernutrition reduces the mortality of acute experimental pancreatitis probably by decreasing pancreatic enzyme content. Cerulein in physiological doses reduces the enzyme content of the pancreas without any harmful effect on the organ. The aim of the present study was to asses the effect of acute reduction of pancreatic enzyme content on the outcome of acute pancreatitis. Two groups of male Wistar rats weighing 230-250 g were studied: group I, 12-h fasted animals, and group II, ad libitum-fed animals who received cerulein at the inframaximal dose (0.2 microgram kg-1 h-1) for 2 h. Cerulein administration resulted in the reduction of the pancreatic contents of chymotrypsinogen (71%), trypsinogen (55%), proelastase (60%), amylase (62%) and cathepsin B (45%) (P < 0.05). However, no significant reduction in pancreatic phospholipase content was observed. Acute pancreatitis was induced in group I after 12-h fasting and in group II at the end of cerulein infusion by retrograde injection o 0.5 ml of 2.5% Na+ taurocholate into the pancreatic duct. Ascites volume and the degree of histologically observed lesions were similar in both groups, but 72-h mortality was 56% in the control group (10/ 18) and 23% (5/22) in the cerulein group (P < 0.05). We speculate that the reduction of pancreatic enzyme content may exert its beneficial effect in acute pancreatitis by decreasing the quantity of pancreatic enzymes reaching the circulation and consequently their pathogenic effects.

Reduction of pancreatic enzyme content and mortality in experimental acute pancreatitis in rats

A previous report has shwon that undernutrition reduces the mortality of acute experimental pancreatitis probably by decreasing pancreatic enzyme content of the pancreas without any harmful effect on the organ. The aim of the present study was to assess the effect of acute reduction of pancreatic enzyme content on the outcome of acute pancreatitis. Two groups of males Wistar rats weighing 230-250 g were studied: group I, 12-h fasted animals, and group II, add libitum-fed animals who received cerulein at the inframaximal dose (0.2 µg kg-1h-1) for 2 h. Cerulein adminsitration resulted in the reduction of the pancreatic contents of chymotrypsinogen (71 percent), trypsinogen (55 percent), proelastase (60 percent), amylase (62 percent) and cathepsin B (45 percent) (P<0.05). However, no significant reduction in pancreatic phospholipase content was observed. Acute pancreatitis wass induced in group I after 12-h fasting and in group II at the end of cerulein infusion by retrograde injection of 0.5 ml of 2.5 percent Na+ taurocholate into the pancreatic duct. Ascites volume and the degree of histologically observed lesions were similar in both groups, but 72-h mortality was 56 percent in the control group (10/18) and 23 percent (5/22) in the cerulein group (P<0.05). We speculate that the reduction of pancreatic enzyme content may exert its beneficial effect in acute pancreatitis by decreasing the quantity of pancreatic enzymes reaching the circulation and consequently their pathogenic effects

[Simplified model of induction of experimental acute pancreatitis with a supra-maximal dose of cerulein]. / Modelo simplificado de indução de pancreatite aguda experimental com uso de dose supramáxima de ceruleína.

The study was performed to compare an usual method of induction of acute experimental pancreatitis with a simplified, easier and faster induction through a subcutaneous and intravenous injection of cerulein, with good reproducibility as compared to the literature. Four groups were studied. In the group I, continuous three hour intravenous injection of 15 micrograms/Kg of cerulein, was given. Group II was a control group with saline infusion. Group III received a subcutaneous injection of 20 micrograms/Kg and an intravenous injection of 20 micrograms/Kg of cerulein one hour later. Group IV was the control group with saline. The results of biochemical measurements, such as tecidual trypsinogen, chimotrypsinogen, proelastase, cathepsin and serum amylase, showed no difference between the two methods. Histologic study revealed edematous pancreatitis in group I and III, with moderate acinar necrose in group III. These results suggest that the proposed simplified method induces enough acute and edematous pancreatitis to allow studies in physiopathology and therapeutics.

[Protective effect of pancreatic enzyme depletion in the course of acute pancreatitis--experimental study in rats]. / Efeito protetor da depleção de enzimas pancreáticas na evolução da pancreatite aguda--estudo experimental em ratos.

This study was performed to assess the effects of the reduction of pancreatic enzymatic content in the evolution of acute pancreatitis (A.P.) in rats. We hypothesized that cerulein in physiologic dose reduces the enzymatic content of the pancreas and may decrease the mortality of A.P.. Four groups of rats have been studied. In the group I cerulein in a total dose of 8 ug/Kg was given, Group II was a control group, Group III received cerulein in inframaximal dose (0.4 micrograms/Kg) and Group IV was a control group with free access to diet and water. Acute pancreatitis was induced by retrograde injection of taurocholic acid into the pancreatic duct. The mortality was 60% in group I, 56% in group II, 23% in group III and 34% in group IV. These results suggest that by decreasing enzymatic content of the pancreas the severity of acute pancreatitis may be reduced.