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Methamphetamine (MA) is an illicit stimulant that affects the central nervous system. Cytochrome P450 2D6 (CYP2D6) plays an important role in MA metabolism. Numerous allelic variants confer substantial variation in CYP2D6 activity among individuals. In the present study, we examined the frequencies of CYP2D6 alleles, including CYP2D6*1, *2, *4, *5, *10, *14A, *14B, *18, and *36, and multiplication, in 82 forensic autopsy cases of MA abusers and 567 autopsy cases in which MA was not detected (controls). Ultrarapid metabolizer (UM), extensive metabolizer (EM), intermediate metabolizer (IM), and poor metabolizer (PM) phenotypes were predicted from CYP2D6 genotypes. Of MA abusers, 64 subjects were predicted to be EM, 17 were IM, and 1 was UM. No MA abuser had the predicted PM phenotype. No significant differences in CYP2D6 phenotype frequencies were found between MA abusers and controls. MA and amphetamine (AMP) concentrations were measured in the right heart blood, left heart blood, peripheral external iliac blood, urine, pericardial fluid, and bone marrow of MA abusers. MA concentrations in urine and bone marrow were significantly higher in IM than in EM. AMP concentration was not associated with CYP2D6 phenotype in any body fluid. These results suggest that the MA concentration in body fluids is influenced by CYP2D6 phenotypes in the Japanese population., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Methamphetamine (MA) is an illicit psychostimulant that stimulates the release of catecholamines from sympathetic nerve terminals and is widely abused worldwide. Since catechol-O-methyltransferase (COMT) metabolizes catecholamines and mediates adrenergic, noradrenergic, and dopaminergic signaling responses, we investigated the effects of the COMT polymorphisms rs4633 and rs4680 on cerebrospinal fluid (CSF) catecholamine concentrations in autopsies of subjects who died of drug intoxication. 28 MA abusers and 22 fatal psychotropic drug intoxication cases were evaluated. No correlations were identified between rs4633 or rs4680 polymorphisms and CSF concentrations of adrenaline (Adr), noradrenaline (Nad), or dopamine (DA) in fatal psychotropic cases. However, among MA abusers, DA concentrations in the CSF were significantly higher in those with the T allele (CT and TT) of rs4633 than in CC genotype carriers (p=0.004). Moreover, among MA abusers, DA concentrations were significantly higher in those with the A allele (GA and AA) of rs4680 than in GG genotype carriers (p=0.017). In subsequent haplotype analyses of MA abusers, a strong correlation was identified between two COMT haplotypes and CSF DA concentrations (p=0.002). However, the CSF concentrations of Adr and Nad were not associated with COMT genotypes or haplotypes. The present results indicate that rs4633 and rs4680 polymorphisms influence CSF DA concentrations and MA toxicity in MA abusers., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Methamphetamine (MA) use sometimes causes rhabdomyolysis, which has been associated with mortality. We analyzed potential rhabdomyolysis-susceptibility genes from autopsy samples of 18 methamphetamine abusers. We examined mutations in the ryanodine receptor 1 (RYR 1), carnitine palmitoyltransferase II (CPT II), very long-chain acyl-CoA dehydrogenase (VLCAD), and cytochrome P450 (CYP) 2D6 genes. Different RYR1 mutations that caused amino acid substitutions ((612)Ala>Thr and (4295)Ala>Val) were identified in 2 cases. In the CPT II gene, there was a new mutation ((545)Glu>Ala) in 1 case and there were mutations that did not change activity in 17 cases. In the VLCAD gene, there were mutations that did not change activity in 6 cases. In the CYP2D6 gene, homozygosity for CYP2D6∗10, which is associated with significantly reduced metabolic activity, was found in 3 cases, while 2 cases carried a different previously unreported missense mutation ((344)Arg>Gln and (48)His>Tyr). RYR1 mutations and the new CPT II mutation identified in this study were not observed in a control group. Eighteen cases that were genetically analyzed were also investigated immunohistochemically to diagnose the possibility of rhabdomyolysis. However, there were no significant mutations that reduced enzyme activity in the suspected cases of rhabdomyolysis. These data suggested no obvious relationship between the genetic mutations observed in this study and rhabdomyolysis., (Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.)

To diagnose the cause of death in autopsy cases, systematic examinations, such as macroscopic, pathological, biochemical, and toxicological are important. In this case report, drug examinations also gave very useful information to diagnose the cause of death, fatal diabetic ketoacidosis (DKA). A female methamphetamine abuser in her forties was found dead lying on a hotel bed. Diagnosing her cause of death was difficult only from the macroscopic findings because there was no fatal and/or serious injury or disease. On toxicological examination, acetone was detected at a high concentration (682 microg/mL in blood, 887 microg/mL in urine) using gas chromatography (GC). Using gas chromatography-mass spectrometry (GC-MS), methamphetamine was detected in the blood, urine, hair, and visceral organs; however, these concentrations were low. At the same time, GC-MS examination revealed a high glucose peak. From the results of the biochemical examination of urine, acetoacetic acid was 1940 micromol/L, beta-hydroxybutyric acid was 14,720 micromol/L, and glucose was 4620 mg/dL. Histologically, Langerhans' islets in the pancreas were fibrotic and atrophic, and no insulin-immunoreactive cells were observed. The subsequent police investigation also revealed that she had contracted diabetes mellitus type 1; therefore, we concluded that her cause of death was DKA, due to a lack of insulin injection., (Copyright 2010 Elsevier Ireland Ltd. All rights reserved.)

Little is known about the role of glial cells in the striatum of chronic methamphetamine (METH) users. In this study, we immunohistochemically examined glial reactions in the striatum of chronic METH users who did not abstain from METH use and died of drug intoxication. Human glucose transporter 5 (hGLUT), a useful marker of microglia, and CR3.43, a major histocompatibility complex class II antigen specific for reactive microglia, were immunostained. Glial fibrillary acidic protein (GFAP) and S100 Beta were used for astrocyte immunohistochemistry. We analyzed 12 chronic METH users and 13 control subjects, and detected a 200-240% increase in the number of hGLUT5-positive cells in chronic METH users (p<0.01). However, we did not detect any proliferation of CR3.43-positive cells. The number of GFAP-positive astrocytes increased, but this increase was not significant (p>0.05). Moreover, S100B-positive cell density between the two groups was not significant (p>0.05). This study demonstrates the absence of reactive gliosis in the striatum of chronic METH users who did not abstain for prolonged periods from METH use. The results suggest that chronic METH use by itself did not activate glial cells in humans and reactive gliosis may not be involved in the mechanism underlying the loss of control in drug intake, which is a characteristic feature of drug addiction., ((c) 2009 Elsevier Ireland Ltd. All rights reserved.)

In this study, we investigated methamphetamine (METH)- induced peroxidative DNA damage in various regions of the rat brain. We injected METH to rats following 2 protocols. For the single administration experiment (group I), 50 mg/kg (i. p.) of METH was administered to observe the acute influence of METH. For the repeated administration experiment (group II), 10 mg/kg/day (i. p.) of METH was injected for 5 days. Immunohistochemically, peroxidative damage DNA, 8-hydroxy-2'- deoxyguanosine (8-OH-dG) was observed, and in situ apoptosis was also observed. In group I, immunoreactivity of 8-OH-dG was only enhanced in neurons of the nucleus accumben of METH-treated rats. On in situ apoptosis detection, positive findings were also enhanced in all examined parts compared to those in the control, though there were no significant increases in 8-OH-dG-immunopositive neurons except in the nucleus accumben. In group II, the nucleus accumben also showed enhanced 8-OH-dG immunopositivity compared to that in the control. There was no significant difference in apoptosis between the control and METH groups. Based on our observations, it is considered that METH induces oxidative DNA damage in the brain, especially in the nucleus accumben. However, those DNA damage might be caused differently between acute and chronic administration.

In this study, we observed renal damage and peroxidative injury as the acute or sub-acute effect of methamphetamine (MA) to determine whether MA intoxication can be diagnosed from immunohistochemical changes in the kidney. In addition, renal function was investigated in relation to the immunohistochemical changes. A single administration of MA (group I) (50mg/kg/ (i.p.)) and repeated administration (group II) (10mg/kg/day (i.p.) for 5 days) were designed as an acute model and a sub-acute or chronic model. Immunohistochemically, cell damage markers were observed. Then, renal function markers and minerals in blood were measured. Myoglobin and creatinine phosphokinase (CPK) in blood were also analyzed. In group I, ubiquitin immunoreactivity was enhanced only in the renal tubules. Creatinine increased, while K, Ca, and P decreased (P<0.01). CPK increased significantly (P<0.01). Therefore, it was suspected that MA might induce renal dysfunction with renal tubule damage. This damage might be related to leakage of CPK from muscle. In group II, 8-hydroxy-2'-deoxyguanosine (8-OH-dG) increased immunohistochemically and quantitatively (P<0.01). It was considered that oxidative DNA damage might be induced by repeated administration. It was considered that this study offers basic information for the evaluation of pathological changes in the kidney in MA-related autopsy cases.

It is known that methamphetamine (MA) causes rhabdomyolysis, myoglobinuria, and acute renal failure. We conducted an immunohistochemical study on the kidney of 22 forensic autopsy cases in which MA had been detected. Myoglobin was positive in 17 cases. The concentration of the blood MA in the myoglobin-positive cases (8.39+/-3.43 micromol/dl) was higher than -negative cases (0.198+/-0.076 micromol/dl). And, the 70 kDa heat shock protein (HSP70), 8-hydroxy-2'-deoxyguanosine (8-OH-dG), 4-hydroxy-2-nonenal (4-HNE), and Cu/Zn superoxide dismutase (SOD) were also stained positively in five, ten, 11, nine cases of examined, respectively. In addition, 80% of HSP70-positive cases were myoglobin-positive. Myoglobin was also observed in 60% of 8-OH-dG-positive, in 82% of 4-HNE-positive, and in 78% of SOD-positive cases, respectively. Therefore, myoglobin rather than MA itself might induce oxidative damage. From these results, it was considered that MA abuse had caused the skeletal muscle damage before death. In forensic autopsy cases of drug abusers, the antemortem situation is not often known. The present research suggested that in addition to the measurement of the concentration of MA, immunohistochemical staining of myoglobin, HSP70, 8-OH-dG, 4-HNE, and SOD offers important information for the diagnosis of MA poisoning.

Forensic autopsy cases detecting methamphetamine (MA) are usually diagnosed according to its toxicological concentration. It has been reported that the lethal blood concentration of MA is 4.48 microg/ml (3.0 micromol/dl). We autopsied two MA-detected cadavers, and immunohistochemical staining was performed on the skeletal muscle with an anti-myoglobin antibody, and on the kidney with an anti-the 70 kDa heat shock protein (HSP70) antibody. One case showed a high rectal temperature (40 degrees C). The toxicological examination revealed 0.75 microg/ml of MA in the blood, and 16.8 microg/ml in the urine. Myoglobin was negative and HSP70 was positive in the kidney immunohistochemically. From the toxicological and immunohistochemical findings, it was considered that the subject died of hyperthermia and acidosis caused by muscular hyperactivity. In another case, the autopsy revealed highly congested lungs, with dark-red bloody fluid and foam in the trachea and bronchus. MA (17.0 microg/ml) was detected in the blood. HSP70 was negative and myoglobin was positive immunohistochemically. It was thought that the subject died of acute MA intoxication based on the high MA concentration, although rhabdomyolysis was suspected. It is suggested that myoglobin and HSP70 immunostaining are useful to diagnose MA poisoning.

Forensic laboratories are often faced with cases in which methamphetamine hydrochloride-mixed blood is unable to be identified as human blood by immunochromatography against human hemoglobin A0. The application of mRNA expression analysis to samples that showed a false-negative with immunochromatography was investigated as an alternative approach that did not depend on the antigen-antibody reaction. Real-time PCR was used to examine the expression levels of blood markers such as glycophorin A, spectrin beta, and hemoglobin beta. Hemoglobin beta was the only marker that was specifically detected in blood, while glycophorin A was useful for determining human specificity. Hemoglobin beta showed good detection sensitivity and was detectable in 37-year-old blood stains. Hemoglobin beta was exclusively detectable in methamphetamine hydrochloride-mixed blood stains. Detergents and disinfectants did not significantly influence mRNA markers. The proposed mRNA expression analysis was suitable for human blood identification as an alternative method to immunochromatography. [ABSTRACT FROM AUTHOR]

Dopamine D2 receptor/ankyrin repeat and kinase domain containing 1 ( DRD2/ANKK1 ) gene polymorphisms have been associated with responses to psychotropic drugs and addiction. We analyzed two DRD2/ANKK1 polymorphisms, Taq1A and –141C Ins/Del, in 37 fatal methamphetamine (MA) intoxication cases and 235 control cases in which MA and psychotropic drugs were not detected. The association among polymorphism, cause of death, and cerebrospinal fluid (CSF) dopamine concentration was evaluated. The Taq1A polymorphism distribution in the fatal MA intoxication cases differed from in the controls ( P  = 0.030) with a significantly high A1/A1 + A1/A2 genotype frequency. No significant associations were observed between –141C Ins/Del polymorphisms and MA intoxication cases or between DRD2/ANKK1 polymorphisms and CSF dopamine concentrations. Our findings suggest that the DRD2/ANKK1 Taq1A polymorphism is associated with susceptibility to fatal MA intoxication. [ABSTRACT FROM AUTHOR]

Methamphetamine (MA) is an illicit psychostimulant that stimulates the release of catecholamines from sympathetic nerve terminals and is widely abused worldwide. Since catechol-O-methyltransferase (COMT) metabolizes catecholamines and mediates adrenergic, noradrenergic, and dopaminergic signaling responses, we investigated the effects of the COMT polymorphisms rs4633 and rs4680 on cerebrospinal fluid (CSF) catecholamine concentrations in autopsies of subjects who died of drug intoxication. 28 MA abusers and 22 fatal psychotropic drug intoxication cases were evaluated. No correlations were identified between rs4633 or rs4680 polymorphisms and CSF concentrations of adrenaline (Adr), noradrenaline (Nad), or dopamine (DA) in fatal psychotropic cases. However, among MA abusers, DA concentrations in the CSF were significantly higher in those with the T allele (CT and TT) of rs4633 than in CC genotype carriers (p=0.004). Moreover, among MA abusers, DA concentrations were significantly higher in those with the A allele (GA and AA) of rs4680 than in GG genotype carriers (p=0.017). In subsequent haplotype analyses of MA abusers, a strong correlation was identified between two COMT haplotypes and CSF DA concentrations (p=0.002). However, the CSF concentrations of Adr and Nad were not associated with COMT genotypes or haplotypes. The present results indicate that rs4633 and rs4680 polymorphisms influence CSF DA concentrations and MA toxicity in MA abusers. [ABSTRACT FROM AUTHOR]