Evidence for endogenous formation of tobacco-specific nitrosamines in rats treated with tobacco alkaloids and sodium nitrite

Carcinogenic tobacco-specific nitrosamines are present in tobacco products and are believed to play a significant role in human cancers associated with tobacco use. Additional amounts of tobacco-specific nitrosamines could be formed endogenously. We tested this hypothesis by treating rats with nicotine and sodium nitrite and analyzing their urine. Initially, we treated groups of rats with (S)-nicotine (60 micromol/kg) and NaNO2 (180 micromol/kg), (S)-nicotine alone, NaNO2 alone or 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK, 12 nmol/kg) by gavage twice daily for 4 days. We collected urine and analyzed for two metabolites of NNK; 4-(methylnitrosamino)-1-(3- pyridyl)-1-butanol and its glucuronide. We did not detect these metabolites in the urine of rats treated with nicotine alone or nicotine plus NaNO2, indicating that endogenous conversion of nicotine to NNK did not occur. However, the urine did contain N'- nitrosonornicotine (NNN), N'-nitrosoanabasine (NAB) and N'- nitrosoanatabine (NAT). Analysis of the (S)-nicotine used in this experiment demonstrated that it contained trace amounts of nornicotine, anabasine and anatabine. In a second experiment, we used an identical protocol to compare the endogenous nitrosation of this (S)-nicotine with that of synthetic (R,S)-nicotine, which did not contain detectable amounts of nornicotine, anabasine or anatabine. NNN (0.53 x 10(-3)% of nicotine dose), NAB (0.68%) and NAT (2.1%) were detected in the urine of the rats treated with the (S)-nicotine and NaNO2. NNN (0.47 x 10(- 3)% of dose), but not NAB or NAT, was present in the urine of the rats treated with synthetic (R,S)-nicotine and NaNO2. NNN probably formed via nitrosation of metabolically formed nornicotine. These results demonstrate for the first time that endogenous formation of tobacco- specific nitrosamines occurs in rats treated with tobacco alkaloids and NaNO2. The potential significance of the results with respect to nitrosamine formation in people who use tobacco products or nicotine replacement therapy is discussed.      

Expression of differential nitric oxide synthase isoforms in human normal gastric mucosa and gastric cancer tissue

The present study investigated the expression and distribution of three isoforms of nitric oxide synthase (NOS) in different anatomical regions of the human stomach and in gastric neoplastic tissues by immunohistochemistry using specific antibodies. Intracellular localization of individual isoenzymes of NOS was detected in normal gastric mucosa. Gastric cancer tissues had a marked reduction of all three NOS isoforms expression. The expression of the endothelial NOS, neuronal NOS and inducible NOS in the tumor tissue was significantly lower than in normal gastric mucosa (P = 0.01, P = 0.02, P < 0.01, respectively). In the tumor tissue the expression of inducible NOS was significantly lower than the expression of both constitutive forms of NOS (P < 0.01). There was a tendency to higher expression of both constitutive forms of NOS in earlier stages T2 of the tumor compared to advanced T4 tumor. In contrast, the expression of inducible NOS was higher than in the advanced T4 tumor than in the earlier stages T2 of the tumor. The mapping of the expression of endothelial NOS, neuronal NOS and inducible NOS in human stomach showed higher expression of NOS isoforms in the distal third than in the proximal third of the stomach (P = 0.03, P = 0.04, P = 0.01, respectively). We conclude that there is greater expression of NOS in the stomach corpus and in antrum than in the proximal third of the normal human stomach mirroring the anatomical predilection of common pathological changes in this part of the human stomach. Furthermore, there was loss of the expression of individual isoenzymes in gastric neoplasms.      

Absolute configuration of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol formed metabolically from 4-(methylnitrosamino)-1-(3-pyridyl)-1- butanone

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) is an important metabolite of the tobacco-specific nitrosamine 4-(methylnitrosamino)-1- (3-pyridyl)-1-butanone (NNK). Using the chiral derivatizing agent, (R)- (+)-alpha-methylbenzyl isocyanate [(R)-(+)-MBIC], previous work has shown that the enantiomeric ratio of metabolically formed NNAL and its glucuronide derivative may be species dependent. However, the absolute configuration of such NNAL has not been previously reported. Synthetically prepared racemic NNAL was converted to diastereomeric esters by reaction with (R)-(+)- and (S)-(-)-alpha-methoxy-alpha- (trifluoromethyl)phenylacetic acid (MTPA) chloride (Mosher's reagent) and the products were characterized by 1H-NMR. Based on chemical shift data, the absolute configuration of NNAL in each diastereomeric ester was assigned. Hydrolysis of (R)-NNAL-(R)-MTPA gave (R)-NNAL. This was converted to the corresponding carbamate by reaction with (R)-(+)-alpha- MBIC and the absolute configurations of the diastereomeric carbamates formed by reaction of (R)- and (S)-NNAL with (R)-(+)-MBIC were thereby assigned. Conversion of metabolically produced NNAL to the same carbamates allowed us to assign the NNAL formed from NNK by rat liver microsomes as (R)-NNAL. The major and minor NNAL-glucuronide diastereomers found in the urine of patas monkeys and humans exposed to NNK were similarly assigned; they were formed from (R)-NNAL and (S)- NNAL, respectively.      

MAGNOLOL REDUCES INFARCT SIZE AND SUPPRESSES VENTRICULAR ARRHYTHMIA IN RATS SUBJECTED TO CORONARY LIGATION

1. Magnolol is an active component of Magnolia officinalis. It is 1000-times more potent than α-tocopherol in inhibiting lipid peroxidation in rat heart mitochondira. In the present study, the in vivo antiarrhythmic and anti-ischaemic effects of magnolol in coronary ligated rats were investigated. 2. Male Sprague-Dawley rats were anaesthetized with urethane. Magnolol, at dosages of 10^?7, 10^?8 and 10^?9 g/kg, was adminstered intravenously 15 min before ligation of the coronary artery. 3. The incidence and duration of ventricular tachycardia and ventricular fibrillation during 30 min coronary ligation were significantly reduced by magnolol. Ventricular arrhythmias during 10 min reperfusion after the relief of coronary ligation were also reduced. 4. In rats subjected to 4h coronary ligation, 10^?7 and 10^?8 g/kg magnolol significantly reduced infarct size. 5. We conclude that magnolol may protect the myocardium against ischaemic injury and suppress ventricular arrhythmia during ischaemia and reperfusion.     

谷氨酸脱羧酶抑制剂L-苹果酸对小鼠学习记忆的影响

跳台法和Y迷宫法试验表明,小鼠po L-苹果酸600mg·kg^-1连续5d后对记忆的获得、巩固和再现均有明显的改善作用,并能促进空间辨别学习能力;L-苹果酸改善记忆的作用能被NMDA受体拮抗剂氯胺酮所拮抗。脑内游离氨基酸测定显示,L-苹果酸可明显降低小鼠脑内GABA水平,提高Glu/GABA比值。实验结果表明,脑内GABA水平下降,Glu/GABA比值升高对学习记忆有正性调节作用。     

Aberrant methylation of the Human Hedgehog interacting protein (HHIP) gene in pancreatic neoplasms

Hedgehog pathway overactivity has been implicated in the development of a variety of human cancers. The Human Hedgehog interacting protein (HHIP), a negative regulator of hedgehog signaling, has been shown to be underexpressed in pancreatic cancers. In this study we determined if the HHIP gene is a target for genetic and epigenetic alterations. While no mutations of HHIP were identified, we found complete methylation of the HHIP promoter CpG island in three pancreatic cancer cell lines, and partial hypermethylation in 13/17 (80%) pancreatic cancer cell lines, 35/75 (46%) primary pancreatic cancers and 14/18 (78%) pancreatic cancer xenografts, but no methylation in 13 normal pancreata.In pancreatic cancer cell lines, complete methylation was associated with absent or reduced HHIP expression by real-time RT-PCR. HHIP expression could be restored in methylated cell lines using epigenetic modifier drugs. Restoring the expression of HHIP in pancreatic cancer cells by 5-aza-2'-deoxycytidine led to a decrease in Gli reporter activity, consistent with downregulation of Hedgehog signaling. These results indicate in some pancreatic adenocarcinomas that HHIP is epigenetically inactivated by promoter methylation, and its silencing could contribute to the increased Hedgehog signaling observed in pancreatic neoplasms.     

Mine Blast Injuries - Our Experience

Background

The sudden increase in incidence and magnitude of mine blast injuries prompted us to highlight the problem and its management.

Methods

The cases of mine blast injuries occurring during mining and demining in a particular geographical area were analysed. Total 27 cases of mine blast injuries occurred during mining or demining operations in a period of 13 months.

Results

Various body regions were involved in the mine blast injuries but the main brunt was borne by feet and legs followed by multiple body regions due to splinters. 14 patients underwent below knee (BK) amputation while 4 patients required through knee (TK) amputations. The effect of blast was so severe that most of the cases required 2 to 5 times wound debridements. The initial aggressive debridement / open stump amputation saved the limb and life of all patients.

Conclusion

A mine blast causes extensive injuries and psychological trauma. Management is needed urgently, surgery is difficult, and amputation is often inevitable. Maximum lives and limbs can be saved with aggressive debridement, repeated inspections and dressings under anaesthesia and definitive closure at optimum time.Key Words: Amputation, Antipersonnel mine, Crush syndrome, Debridements, Mine blast injury, Secondary missiles, Shrapenels     

Scintigraphic diagnosis of tricuspid regurgitation

The authors describe a simple technique for diagnosis of tricuspid regurgitation. Red blood cells were labeled in vivo with 99mTc and 22 patients were studied with ECG-gated blood-pool imaging of the liver. A single region of interest was manually drawn around the liver and a time-activity curve obtained. The per cent change in liver counts during the cardiac cycle was found to be significantly higher in the 12 patients with tricuspid regurgitation (Group I) (mean, 4.04 +/- 1.6%; range, 1.3-21.4%) compared with the 10 controls (Group II) (mean, 0.35 +/- 0.16%; range, 0.013-1.3%) (p less than 0.05). Using a 1% change in liver counts as the criterion of a positive study, all 12 cases in Group I were diagnosed correctly, but there was one false positive in Group II; thus the sensitivity was 100% and the specificity 90%.