Evolving etiologies, comorbidities, survival, and costs of care in adult encephalitis.

Encephalitis is a central nervous system disorder, often caused by infectious agents or aberrant immune responses. We investigated causes, comorbidities, costs, and outcomes of encephalitis in a population-based cohort. ICD-10 codes corresponding to encephalitis were used to identify health services records for all adults from 2004 to 2019. Data were cross-validated for identified diagnoses based on laboratory confirmation using univariate and multivariate statistical analyses. We identified persons with a diagnosis of encephalitis and abnormal cerebrospinal fluid (CSF) results (n = 581) in whom viral genome was detected (n = 315) in a population of 3.2 million adults from 2004 to 2019. Viral genome-positive CSF samples included HSV-1 (n = 133), VZV (n = 116), HSV-2 (n = 34), enterovirus (n = 4), EBV (n = 5), and CMV (n = 3) with the remaining viruses included JCV (n = 12) and HHV-6 (n = 1). The mean Charlson Comorbidity Index (2.0) and mortality rate (37.6%) were significantly higher in the CSF viral genome-negative encephalitis group although the mean costs of care were significantly higher for the CSF viral genome-positive group. Cumulative incidence rates showed increased CSF VZV detection in persons with encephalitis, which predominated in persons over 65 years with a higher mean Charlson index. We detected HSV-2 and VZV more frequently in CSF from encephalitis cases with greater material-social deprivation. The mean costs of care were significantly greater for HSV-1 encephalitis group. Encephalitis remains an important cause of neurological disability and death with a viral etiology in 54.2% of affected adults accompanied by substantial costs of care and mortality. Virus-associated encephalitis is evolving with increased VZV detection, especially in older persons.

Human pegivirus viremia in HCV/HIV co-infected patients: Direct acting antivirals exert anti-pegivirus effects.

BACKGROUND: Human pegivirus (HPgV) is a single-stranded RNA virus​ that is closely related to hepatitis C virus (HCV)​. HPgV has also been shown to infect patients with human immunodeficiency virus (HIV). The mechanisms and disease outcomes of HPgV infections are largely unknown, although it has been implicated in both cancer and neurological diseases. There are no established therapies for HPgV. OBJECTIVES: To estimate the prevalence of HPgV in a cohort of HCV/HIV co-infected patients undergoing treatment for HCV with direct acting antivirals (DAA) and investigate the effect of DAA therapy on HPgV infection. STUDY DESIGN: RNA was extracted from plasma samples collected at time points before, during, and after DAA. HPgV RNA abundance was quantified by droplet digital PCR assays targeting the NS5A and 5'UTR domains and confirmed by RT-qPCR. Clinical, demographic and treatment data were analysed. RESULTS: HPgV RNA was detected and quantified in 26 of 100 patients' plasma (26%) before starting DAA. Patients with detectable HPgV were more likely to be male, had higher peak HIV plasma levels, and a history of injection drug use. Patients receiving sofosbuvir/ledipasvir (n = 9) displayed significantly lower HPgV levels at time of DAA completion and had lower post-DAA HPgV rebound​ levels compared to patients receiving sofosbuvir/velpatasvir (n = 11) although both regimens significantly reduced viremia directly following DAA completion. Sustained suppression of HPgV was â€‹also observed among patients (n = 2) receiving pegylated-interferon. CONCLUSIONS: HPgV RNA ​was frequently detected in HCV/HIV co-infected patients and ​was​ supressed by DAA and pegylated interferon therapies with sofosbuvir-ledipasvir showing greatest antiviral activity. These findings suggest potential treatment strategies for HPgV infections​.

Accuracy of Ankle-Brachial Index, Toe-Brachial Index, and Risk Classification Score in Discriminating Peripheral Artery Disease in Patients With Chronic Kidney Disease.

The accuracy of ankle-brachial index (ABI) and toe-brachial index (TBI) in discriminating lower extremity peripheral artery disease (PAD) has not been evaluated in patients with chronic kidney disease (CKD). We measured ABI, TBI, and Doppler ultrasound in 100 predialysis patients with CKD without revascularization or amputation. Leg-specific ABI was calculated using higher systolic blood pressure (SBP) in posterior tibial or dorsalis pedis artery divided by higher brachial SBP; alternative ABI was calculated using lower SBP in posterior tibial or dorsalis pedis artery. PAD was defined as ≥50% stenosis detected by Doppler ultrasound. PAD risk classification score was calculated using cardiovascular disease risk factors. The area under the curve (AUC, 95% confidence interval [CI]) for discriminating ultrasound-diagnosed PAD was 0.78 (0.69 to 0.87) by ABI, 0.80 (0.71 to 0.89) by alternative ABI, and 0.74 (0.63 to 0.86) by TBI. Sensitivity and specificity were 25% and 97% for ABI ≤0.9, 41% and 95% for alternative ABI ≤0.9, and 45% and 93% for TBI ≤0.7, respectively. AUC (95% CI) of PAD risk classification score was 0.86 (0.78 to 0.94) with sensitivity and specificity of 95% and 60% for risk score ≥0.10, 76% and 76% for risk score ≥0.25, and 43% and 95% for risk score ≥0.55. Combining risk score with ABI, alternative ABI, and TBI increased AUC (95% CI) to 0.89 (0.82 to 0.96), 0.89 (0.80 to 0.98), and 0.87 (0.78 to 0.96), respectively. In conclusion, current ABI and TBI diagnostic criteria have high specificity but low sensitivity for classifying PAD in patients with CKD. PAD classification risk score based on cardiovascular disease risk factors improves the accuracy of PAD classification.

Progressive multifocal leukoencephalopathy and Creutzfeldt-Jakob disease: population-wide incidences, comorbidities, costs of care, and outcomes.

Neurological disorders associated with chronic infections are often progressive as well as challenging to diagnose and manage. Among 4.4 million persons from 2004 to 2019 receiving universal health, progressive multifocal leukoencephalopathy (PML, n = 58) and Creutzfeldt-Jakob disease (CJD, n = 93) cases were identified, revealing stable yearly incidence rates with divergent comorbidities: HIV/AIDS affected 37.8% of PML cases while cerebrovascular disease affected 26.9% of CJD cases. Most CJD cases died within 1 year (73%) although PML cases lived beyond 5 years (34.1%) despite higher initial costs of care. PML and CJD represent important neurological disorders with evolving risk variables and impact on health care.

Combination Treatment with Sodium Nitrite and Isoquercetin on Endothelial Dysfunction among Patients with CKD: A Randomized Phase 2 Pilot Trial.

BACKGROUND AND OBJECTIVES: Endothelial dysfunction is common among patients with CKD. We tested the efficacy and safety of combination treatment with sodium nitrite and isoquercetin on biomarkers of endothelial dysfunction in patients with CKD. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: This randomized, double-blind, placebo-controlled phase 2 pilot trial enrolled 70 patients with predialysis CKD. Thirty-five were randomly assigned to combination treatment with sodium nitrite (40 mg twice daily) and isoquercetin (225 mg once daily) for 12 weeks, and 35 were randomly assigned to placebo. The primary outcome was mean change in flow-mediated vasodilation over the 12-week intervention. Secondary and safety outcomes included biomarkers of endothelial dysfunction, inflammation, and oxidative stress as well as kidney function, methemoglobin, and adverse events. Intention-to-treat analysis was conducted. RESULTS: Baseline characteristics, including age, sex, race, cigarette smoking, history of hypertension and diabetes, use of renin-angiotensin system blockers, BP, fasting glucose, lipid profile, kidney function, urine albumin-creatinine ratio, and endothelial biomarkers, were comparable between groups. Over the 12-week intervention, flow-mediated vasodilation increased 1.1% (95% confidence interval, -0.1 to 2.3) in the treatment group and 0.3% (95% confidence interval, -0.9 to 1.5) in the placebo group, and net change was 0.8% (95% confidence interval, -0.9 to 2.5). In addition, changes in biomarkers of endothelial dysfunction (vascular adhesion molecule-1, intercellular adhesion molecule-1, E-selectin, vWf, endostatin, and asymmetric dimethylarginine), inflammation (TNF-α, IL-6, C-reactive protein, IL-1 receptor antagonist, and monocyte chemoattractant protein-1), and oxidative stress (oxidized LDL and nitrotyrosines) were not significantly different between the two groups. Furthermore, changes in eGFR, urine albumin-creatinine ratio, methemoglobin, and adverse events were not significantly different between groups. CONCLUSIONS: This randomized phase 2 pilot trial suggests that combination treatment with sodium nitrite and isoquercetin did not significantly improve flow-mediated vasodilation or other endothelial function biomarkers but also did not increase adverse events compared with placebo among patients with CKD. CLINICAL TRIAL REGISTRY NAME AND REGISTRATION NUMBER: Nitrite, Isoquercetin, and Endothelial Dysfunction (NICE), NCT02552888.

Impairment of Nrf2- and Nitrergic-Mediated Gastrointestinal Motility in an MPTP Mouse Model of Parkinson's Disease.

BACKGROUND: Gastrointestinal (GI) motility dysfunction is the most common non-motor symptom of Parkinson's disease (PD). Studies have indicated that GI motility functions are impaired before the onset of PD. AIMS: To investigate the underlying mechanism of PD-induced GI dysmotility in MPTP (1-methyl 4-phenyl 1,2,3,6-tetrahydropyridine)-induced animal model. METHODS: C57BL/6 mice were administered with or without a selective dopamine neurotoxin, MPTP, to induce parkinsonian symptoms. In addition to in vivo studies, in vitro experiments were also conducted in colon specimens using l-methyl-4-phenylpyridinium (MPP+), a metabolic product of MPTP. Gastric emptying, colon motility, nitrergic relaxation, and western blot experiments were performed as reported. RESULTS: MPTP-induced PD mice showed decreased expression of nuclear factor erythroid 2-related factor (Nrf2) and its target phase II genes in gastric and colon neuromuscular tissues. Decreased levels of tetrahydrobiopterin (BH4, a critical cofactor for nNOS dimerization) associated with uncoupling of nNOS in gastric and colon tissues exposed to MPTP. Impaired enteric nitrergic system led to delayed gastric emptying and slower colonic motility compared to the control mice. In vitro results in colon specimens confirm that activation of Nrf2 restored MPP+-induced suppression of alpha-synuclein, tyrosine hydroxylase (TH), Nrf2, and heme oxygenase-1. In vitro exposure to L-NAME [N(w)-nitro-L-arginine methyl ester], a NOS synthase inhibitor, reduced protein expression of TH in colon tissue homogenates. CONCLUSIONS: Loss of Nrf2/BH4/nNOS expression in PD impairs antioxidant gene expression, which deregulates NO synthesis, thereby contributing to the development of GI dysmotility and constipation. Nitric oxide appears to be important to maintain dopamine synthesis in the colon.

Heterophilic interference in specimens yielding false-reactive results on the Abbott 4th generation ARCHITECT HIV Ag/Ab Combo assay.

BACKGROUND: False-reactivity in HIV-negative specimens has been detected in HIV fourth-generation antigen/antibody or 'combo' assays which are able to detect both anti-HIV-1/HIV-2 antibodies and HIV-1 antigen. OBJECTIVES: We sought to characterize these specimens and determine the effect of heterophilic interference. STUDY DESIGN: Specimens previously testing as false-reactive on the Abbott ARCHITECT HIV Ag/Ab combo assay and re-tested on a different (Siemens ADVIA Centaur HIV Ag/Ab) assay. A subset of these specimens were also pre-treated with heterophilic blocking agents and re-tested on the Abbott assay. RESULTS: Here we report that 95% (252/264) of clinical specimens that were repeatedly reactive on the Abbott ARCHITECT HIV Ag/Ab combo assay (S/Co range, 0.94-678) were negative when re-tested on a different fourth generation HIV combo assay (Siemens ADVIA Centaur HIV Ag/Ab). All 264 samples were subsequently confirmed to be HIV negative. On a small subset (57) of specimens with available volume, pre-treatment with two different reagents (HBT; Heterophilic Blocking Tube, NABT; Non-Specific Blocking Tube) designed to block heterophilic antibody interference either eliminated (HBT) or reduced (NABT) the false reactivity when re-tested on the ARCHITECT HIV Ag/Ab combo assay. CONCLUSIONS: Our results suggest that the Abbott ARCHITECT HIV Ag/Ab combo assay can be prone to heterophilic antibody interference.

The multi-faceted dynamics of HIV-1 transmission in Northern Alberta: A combined analysis of virus genetic and public health data.

Molecular epidemiology has become a key tool for tracking infectious disease epidemics. Here, the spread of the most prevalent HIV-1 subtypes in Northern Alberta, Canada, was characterized with a Bayesian phylogenetic approach using 1146 HIV-1 pol sequences collected between 2007 and 2013 for routine clinical management purposes. Available patient metadata were qualitatively interpreted and correlated with onwards transmission using Fisher exact tests and logistic regression. Most infections were from subtypes A (n=36), B (n=815) and C (n=211). Africa is the dominant origin location for subtypes A and C while the subtype B epidemic was seeded from the USA and Middle America and, from the early 1990s onwards, mostly by interprovincial spread. Subtypes A (77.8%) and C (74.0%) were usually heterosexually transmitted and circulate predominantly among Blacks (61.1% and 85% respectively). Subtype B was mostly found among Caucasians (48.6%) and First Nations (36.8%), and its modes of transmission were stratified by ethnic origin. Compared to subtypes A (5.6%) and C (3.8-10.0%), a larger portion of subtype B patients were found within putative provincial transmission networks (20.3-29.5%), and this almost doubled when focusing on nationwide transmission clusters (37.9-57.5%). No clear association between cluster membership and particular patient characteristics was found. This study reveals complex and multi-faceted transmission dynamics of the HIV-1 epidemic in this otherwise low HIV prevalence population in Northern Alberta, Canada. These findings can aid public health planning.

Prevention and control of carbapenemase-producing organisms at a regional burns centre.

In many parts of the world, carbapenemase-producing organisms (CPOs) are endemic. The transfer of medical patients from such countries to the UK requires us to have control systems in place to avoid onward transmission. This report describes the experience of a regional burns centre challenged by its first four cases of CPO in two separate incidents. Key learning from our experience was the importance of CPOs being considered in empirical antibiotics for any patient from an endemic area. Using contact plates, we demonstrated high bacterial counts after cleaning and we describe a terminal cleaning strategy along with the importance of continuing staff engagement and education.

Pharmacokinetics of oxycodone after subcutaneous administration in a critically ill population compared with a healthy cohort.

This study aimed to characterise and compare the absorption pharmacokinetics of a single subcutaneous dose of oxycodone in critically ill patients and healthy subjects. Blood samples taken at intervals from two minutes to eight hours after a subcutaneous dose of oxycodone in patients (5 mg) and healthy volunteers (10 mg) were assayed using high performance liquid chromatography. Data were analysed using a non-compartmental approach and presented as mean (SD). Parameters were corrected for dose differences between the groups assuming linear kinetics. Ten patients (eight male, two female) and seven healthy male subjects were included. Maximum venous concentration and area under the concentration curve were approximately two-fold lower in the patient group for an equivalent dose, suggesting either reduced bioavailability or increased clearance: maximum venous concentration 0.14 ± 0.06 vs 0.05 ± 0.02 µg/ml (P <0.0001); area under the concentration curve 19.50 ± 9.15 vs 9.72 ± 2.71 µg/ml/minute (P <0.001) respectively. However, time to maximum venous concentration and mean residence time were not different, suggesting similar absorption rates: time to maximum venous concentration 22.10 ± 18.0 vs 20.50 ± 16.10 minutes (P=0.81); mean residence time 353 ± 191 vs 291 ± 80 minutes (P=0.26). Kinetic parameters were less variable in patients than in volunteers. The patients therefore had reduced exposure to subcutaneous oxycodone. This warrants further model-based analysis and experimentation. Dose regimens for subcutaneous oxycodone developed in healthy volunteers cannot be directly translated to critically ill patients.

Performing slow vital capacity in older people with and without cognitive impairment--is it useful?

BACKGROUND: most patients with moderate cognitive impairment are unable to perform forced spirometry. It has been suggested that slow vital capacity (SVC) is easier to perform than forced vital capacity (FVC) because it requires less understanding and co-ordination. We conducted a study to determine whether that assertion is correct. METHODS: we studied 83 inpatients, mean age 83 years (range 67-95, 51 female). They had measurements made of FVC, SVC and the Mini-Mental State Examination (MMSE). The spirometry was conducted using the European Respiratory Society/American Thoracic Society standards. RESULTS: of the 83 subjects, 38 were able to do both FVC and SVC and 32 were unable to do either. The overall concordance was 84%. Twelve were able to do SVC but not FVC (eight due to excessive cough, two due to weakness and two had an MMSE < 24 with poor co-ordination). An inability to do neither FVC nor SVC was predicted by an MMSE < 24/30 (P < 0.0001) with a sensitivity of 88% and specificity of 67%. CONCLUSION: SVC is not a usable substitute for FVC for elderly patients with cognitive impairment but is of some utility for those who tend to cough. An MMSE < 24/30 is predictive of inability to perform FVC and SVC.

Modeling a sensitization stage and a precipitation stage for Parkinson's disease using prenatal and postnatal 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine administration.

Idiopathic Parkinson's disease (PD) is a neurodegenerative disorder of mature and older individuals. Since all aged individuals do not develop PD, predisposing conditions may exist that pair with the stress placed on the basal ganglia during aging to produce the symptoms of PD. In this project we used 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to test the hypothesis that a sensitization stage and a precipitating stage underlie idiopathic PD. To induce the sensitization stage, pregnant C57BL/6J mice were treated with MPTP (10 mg/kg/day) during gestation days 8-12 to target the emerging fetal nigrostriatal dopamine neurons. For the precipitating stage, the 3-months old offspring were administered MPTP for 7 days, to simulate the changes that occur during aging. The weights and motor activity of the offspring, high performance liquid chromatography (HPLC) striatal dopamine and its metabolites and Western blot for tyrosine hydroxylase (TH) were determined. Offspring exposed to prenatal MPTP showed lower birth weights that eventually recovered. Prenatal MPTP also reduced motor activity by 10-30%, striatal TH by 38%, dopamine by 14%, homovanillic acid by 16.5% and 3-methoxytyramine by 66%. The postnatal MPTP was more potent in the prenatal MPTP-exposed offspring. MPTP at 10, 20 and 30 mg/kg, dose-relatedly, reduced striatal TH by 9.4%, 48.6% and 82.4% in the prenatal-phosphate buffered saline (PBS) mice and by 48%, 78.7% and 92.7% in the prenatal-MPTP groups. More importantly, postnatal MPTP at 10 mg/kg that showed slight effects on DA, DOPAC, HVA and 3-MT in the prenatal-PBS offspring, showed 69.9%, 80.0%, 48.4% and 65.4% reductions in the prenatal-MPTP mice. The study may identify a new model for PD, and the outcome suggests that some cases of idiopathic PD may have a fetal basis in which early subtle nigrostriatal impairments occurred and PD symptoms are precipitated later by deteriorating changes in the nigrostriatum, that would not caused symptoms in individuals with normal nigrostriatal system.

4-Nonylphenol (NP) in food-contact materials: analytical methodology and occurrence.

Nonylphenol is a recognized environmental contaminant, but it is unclear whether its occurrence in food arises only through environmental pathways or also during the processing or packaging of food, as there are reports that indicate that materials in contact with food such as rubber products and polyvinylchloride wraps can contain nonylphenol. A review of the literature has highlighted the scarcity of robust analytical methodology or data on the occurrence of nonylphenol in packaging materials. This paper describes a methodology for the determination of nonylphenol in a variety of packaging materials, which includes plastics, paper and rubber. The method uses either Soxhlet extraction or dissolution followed by solvent extraction (depending on the material type), followed by purification using adsorption chromatography. Procedures were internally standardized using 13C-labelled nonylphenol and the analytes were measured by gas chromatography-mass spectrometry. The method is validated and data relating to quality parameters such as limits of detection, recovery, precision and linearity of measurement are provided. Analysis of a range of 25 food-contact materials found nonylphenol at concentrations of 64-287 microg g(-1) in some polystyrene and polyvinylchloride samples. Far lower concentrations (<0.03-1.4 microg g(-1)) were detected in the other materials. It is possible that occurrence at the higher levels has the potential for migration to food.

Prolonged peripheral nerve stimulation induces persistent changes in excitability of human motor cortex.

This study sought to determine whether prolonged peripheral nerve stimulation was effective in inducing persistent "plastic" changes in the excitability of the human motor cortex. The amplitude of the electromyographic response evoked in resting intrinsic hand muscles by focal transcranial magnetic stimulation (TMS) was taken as an index of motor cortical excitability. Twelve subjects were stimulated with each of three protocols, one of which was given on each of three separate occasions. The protocols consisted of various schedules of electrical stimulation of the radial and ulnar nerves or the motor point of the first dorsal interosseous muscle (FDI), or stimulation of FDI motor point paired with low-frequency TMS. Amplitudes of TMS-elicited motor evoked potentials (MEPs) were measured before peripheral stimulation and for 2 h after stimulation. The data from one subject were unusable. In every other subject, all three protocols induced a prolonged, significant facilitation of MEPs in at least some of the three intrinsic hand muscles used. In some instances, MEPs were not enlarged and occasionally were significantly depressed. Different protocols based on peripheral afferent stimulation can induce plastic changes in the organisation of the motor cortex that persist for at least 2 h.

Evidence for direct learning curve for technique on change of cervical sampling device.

The recommended sampling device within the NHS Cervical Screening Programme is the Aylesbury spatula. A local decision was taken to decrease brush usage (either alone or in combination) by 50% from an initial level of 41.8%, with an initial inadequate smear rate of 9.93%. This was managed by controlling smear taking equipment through provision of smear taking kits. The monthly inadequate rate unexpectedly rose to 17.8% before dropping back to previous levels. Brush usage overall fell to 35.2%, with a corresponding increase in spatula use. The possible reasons for this are discussed. The facts suggest that these changes were directly linked, and that there is a learning curve with change of sampling device and that a rise in the inadequate rate should be expected under these circumstances.

1-Methyl-4-phenyl-pyridinium increases S-adenosyl-L-methionine dependent phospholipid methylation.

1-Methyl-4-phenyl-pyridinium (MPP(+)) and S-adenosyl-L-methionine (SAM) cause Parkinson's disease (PD)-like changes. SAM and MPP(+) require their charged S-methyl and N-methyl groups, so the PD-like symptoms may be related to their ability to modulate the methylation process. The SAM-dependent methylation of phosphatidylethanolamine (PTE) to produce phosphatidylcholine (PTC), via phosphatidylethanolamine-N-methyltransferase (PEMT), and the hydrolysis of PTC to form lyso-PTC, a cytotoxic agent, are potential loci for the action of MPP(+). In this study, the effects of MPP(+) on the methylation of PTE to PTC and the production of lyso-PTC were determined. The results showed that SAM increased PTC and lyso-PTC. The rat striatum showed the highest PEMT activity and lyso-PTC formation, which substantiate with the fact that the striatum is the major structure that is affected in PD. MPP(+) significantly enhanced PEMT activity and the formation of lyso-PTC in the rat liver and brain. MPP(+) increased the affinity and the V(max) of PEMT for SAM. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) effect was lesser and inhibited by deprenyl (MAO-B inhibitor). The nor-methyl analogs of MPP(+) were inactive, but some of the charged analogs of MPP(+) showed comparable effects to those of MPP(+). Lyso-PTC that can be increased by SAM and MPP(+) caused severe impairments of locomotor activities in rats. These results indicate that SAM and MPP(+) have complementary effects on phospholipid methylation. Thus, SAM-induced hypermethylation could be involved in the etiology of PD and an increase of phospholipid methylation could be one of the mechanisms by which MPP(+) causes parkinsonism.

L-dopa upregulates the expression and activities of methionine adenosyl transferase and catechol-O-methyltransferase.

High nonphysiological doses of l-dopa are administered to Parkinson's disease (PD) patients, to replenish the depleted dopamine (DA). A large portion of the administered L-dopa and the newly formed DA undergoes methylation by reacting with S-adenosyl-L-methionine (SAM). In the process SAM, as well as L-dopa and DA, is utilized and great demands are placed on the transmethylation system. In this study we investigated whether L-dopa increases the transmethylation process by inducing methionine adenosyl transferase (MAT), the enzyme that produces SAM, and catechol-O-methyl transferase (COMT), the enzyme that transfers the methyl group from SAM to L-dopa and DA. Swiss Webster mice were injected with L-dopa, four times/day, for 1 to 16 days. Brain DA, 3-O-methyldopa (3-OMD), SAM, S-adenosylhomocysteine (SAH), MAT, and COMT were measured following a 24-h withdrawal period. An increase of 264% of brain DA occurred at days 2 and 3 after which it tapered to about 164% of control. The brain level of 3-OMD increased to 870% of the control. SAM was increased by 44% after the sixth day and SAH level was about double after the second day. After day 3, MAT activity was increased by about 35%. Western blot analysis showed that MAT is more clearly characterized in 10% mercaptoethanol reducing buffer in which 31.5-, 38- (beta), and 48-kDa (alpha1/alpha2) subunits were distinctly revealed. The induction of the 38-kDa and, more prominently, the 48-kDa subunits of MAT and the potential transactivator proteins of MAT, c-Jun/AP-1, was evident by day 6. The 31.5-kDa subunit was downregulated. COMT was detected as 24.7-, 30-, and 47.5-kDa bands in the brain, consistent with the membrane-bound COMT I (MB-COMT) and the dimeric COMT II. The 24.7- and the 30-kDa MB-COMT bands were induced in the brain by day 6 and peaked on day 9. The highlight of the study is the fact that L-dopa induces the enzymes MAT and COMT. In addition, the downturn in brain DA after the sixth day coincides with the increase in SAM and the 48-kDa MAT protein. Thus, during PD treatment with L-dopa the induction of MAT and COMT is likely to occur and in turn increase the methylation and reduction of L-dopa and DA that may help cause the tolerance or the wearing-off effect developed to L-dopa.