Safety and efficacy of filgotinib, lanraplenib and tirabrutinib in Sjögren's syndrome: a randomized, phase 2, double-blind, placebo-controlled study.

OBJECTIVE: The aim of this study was to characterize the safety and efficacy of filgotinib, lanraplenib and tirabrutinib in patients with active SS. METHODS: This multicentre, double-blind study randomized patients with active primary or secondary SS [EULAR SS disease activity index (ESSDAI) ≥5) to receive filgotinib 200 mg (Janus kinase-1 inhibitor), lanraplenib 30 mg (spleen tyrosine kinase inhibitor), tirabrutinib 40 mg (Bruton's tyrosine kinase inhibitor), or placebo. The composite primary end point was the week-12 proportion of patients fulfilling protocol-specified improvement criteria (based on CRP and SS-related symptoms). The EULAR SS patient-reported index (ESSPRI) and the ESSDAI change from baseline (CFB) were secondary end points. Exploratory end points included disease-related biomarkers. Treatment-emergent adverse events (AEs) represented safety outcomes. RESULTS: The mean of the baseline ESSDAI was 10.1, and of ESSPRI was 6.2 in the 150 patients who were treated; 125 completed the 24-week placebo-controlled treatment period. At week 12, 43.3% of the filgotinib group achieved the primary end point (P = 0.17 vs placebo) vs 42.3% (P = 0.16), 34.7% (P = 0.33), and 26.7% of lanraplenib, tirabrutinib, and placebo groups, respectively. Neither secondary end point was met. Biomarker reductions included immunoglobulins classically associated with SS disease activity. Filgotinib ESSDAI CFB appeared more pronounced in subgroups with baseline ESSDAI ≥14 or without DMARDs/CSs. Most AEs were Grade 1 or 2. CONCLUSION: Three drugs with disparate mechanisms were tested, but no significant differences vs placebo in primary or secondary end points were observed. These results may be considered hypothesis-generating, given the drug tolerability, subgroup analysis, and biomarker findings. TRIAL REGISTRATION: ClinicalTrials.gov, https://clinicaltrials.gov, NCT03100942.

Blood Harmane (1-Methyl-9H-Pyrido[3,4-b]indole) and Mercury in Essential Tremor: A Population-Based, Environmental Epidemiology Study in the Faroe Islands.

BACKGROUND: Essential tremor (ET) is among the most prevalent neurological diseases. Its environmental determinants are poorly understood. Harmane (1-methyl-9H-pyrido[3, 4-b]indole), a dietary tremor-producing neurotoxin, has been linked to ET in a few studies in New York and Madrid. Mercury, also a tremor-producing neurotoxin, has not been studied in ET. The Faroe Islands have been the focus of epidemiological investigations of numerous neurological disorders. OBJECTIVE: In this population-based, case-control study, we directly measured blood harmane concentrations (HA) and blood mercury concentrations (Hg) in ET cases and controls. METHODS: In total, 1,328 Faroese adults were screened; 26 ET cases were identified whose (HA) and (Hg) were compared to 197 controls. RESULTS: Although there were no statistically significant differences between diagnostic groups, median (HA) was 2.7× higher in definite ET (4.13 g-10/mL) and 1.5× higher in probable ET (2.28 g-10/mL) than controls (1.53 g-10/mL). Small sample size was a limitation. For definite ET versus controls, p = 0.126. (Hg) were similar between groups. CONCLUSIONS: We demonstrated marginally elevated (HA) in definite and probable ET. These data are similar to those previously published and possibly extend etiological links between this neurotoxin and ET to a third locale. The study did not support a link between mercury and ET.

Neurological and neuropsychiatric manifestations of porphyria.

Porphyrias are inherited disorders of the heme biosynthetic pathway, usually characterized by dermatological changes due to the accumulation of byproducts in the pathway. Select porphyrias also affect the nervous system, namely hereditary coproporphyria, acute intermittent porphyria and variegate porphyria. Complications include paralysis, hyponatremia which can risk central pontine myelinolysis, seizures and coma. Neurological complications usually result from severe episodes of acute attacks. Acute attacks may also elicit neuropsychiatric symptoms such as confusion, hallucinations, anxiety and psychosis. However, these manifestations are generally self-limiting. Due to the generally low incidence of porphyria and full knowledge the associated neurological and psychiatric manifestations, we review the relevant porphyrias along with their clinical manifestations, evaluation, and management to raise its awareness in the clinical picture and to prevent misdiagnosis. Porphyria should be considered within the differential diagnosis for unexplained neurological symptoms.

Blood harmane (1-methyl-9H-pyrido[3,4-b]indole) concentrations in essential tremor: repeat observation in cases and controls in New York.

Essential tremor (ET) is a widespread late-life neurological disease. Genetic and environmental factors are likely to play important etiological roles. Harmane (1-methyl-9H-pyrido[3,4-b]indole) is a potent tremor-producing neurotoxin. Previously, elevated blood harmane concentrations were demonstrated in ET cases compared to controls, but these observations have all been cross-sectional, assessing each subject at only one time point. Thus, no one has ever repeat-assayed blood harmane in the same subjects twice. Whether the observed case-control difference persists at a second time point, years later, is unknown. The current goal was to reassess a sample of our ET cases and controls to determine whether blood harmane concentration remained elevated in ET at a second time point. Blood harmane concentrations were quantified by a well-established high-performance liquid chromatography method in 63 ET cases and 70 controls. A mean of approximately 6 yr elapsed between the initial and this subsequent blood harmane determination. The mean log blood harmane concentration was significantly higher in cases than controls (0.30 ± 0.61 g(-10)/ml versus 0.08 ± 0.55 g(-10)/ml), and the median value in cases was double that of controls: 0.22 g(-10)/ml versus 0.11 g(-10)/ml. The log blood harmane concentration was highest in cases with a family history of ET. Blood harmane concentration was elevated in ET cases compared to controls when reassessed at a second time point several years later, indicating what seems to be a stable association between this environmental toxin and ET.

On the Shoulders of Giants: A Reckoning with Social Justice.

As a field, bioethics has failed to adequately change in a direction that pursues and addresses continually shifting contemporary social problems, in particular, anti-Black racism. In this essay, we draw from interviews with four senior Black scholars-Anita L. Allen, Claretta Y. Dupree, Patricia A. King, and Lawrence J. Prograis, Jr.-to learn from their experiences in this field dominated by White-majority thought and to consider thematically how best to recalibrate bioethics to imagine a braver, broader, and better bioethics, one that centers social justice and is equipped to work against anti-Black racism.

Brain regional pharmacokinetics of p-aminosalicylic acid and its N-acetylated metabolite: effectiveness in chelating brain manganese.

para-aminosalicylic acid (PAS; 4-amino-2-hydroxybenzoic acid), an antituberculosis drug in use since the 1950s, has recently been suggested to be an effective agent for treatment of manganese-induced parkinsonian disorders. However, the neuropharmacokinetics of PAS and its metabolite N-acetyl-para-aminosalicylic acid (AcPAS; N-acetyl-4-amino-2-hydroxybenzoic acid) are unknown. This study was designed to investigate the pharmacokinetics of PAS and its distribution in brain to help better design the dosing regimen for clinical trials. Male Sprague-Dawley rats received single femoral artery injections of PAS (200 mg/kg). Plasma, cerebrospinal fluid, and brain tissues were collected, and PAS and AcPAS concentrations were quantified by high-performance liquid chromatography. After administration, the concentrations of PAS declined rapidly in plasma with an elimination t(½) of 34 min; the metabolite AcPAS was detected in plasma and eliminated with a t(½) of 147 min. PAS and AcPAS were detected in brain tissues; AcPAS had a much higher tissue concentration and a longer t(½) than the parent PAS in most tissues examined. Although both were present in blood or tissues as free, unbound molecules, AcPAS appeared to have a higher tissue affinity than PAS. Taken together, our results suggest that a dosing regimen with continuous intravenous infusion of PAS is necessary to achieve therapeutic levels in targeted brain regions. Furthermore, PAS and AcPAS seem to be effective in reducing manganese levels in brain.

Antiracist Praxis in Public Health: A Call for Ethical Reflections.

The Covid-19 pandemic has revealed myriad social, economic, and health inequities that disproportionately burden populations that have been made medically or socially vulnerable. Inspired by state and local governments that declared racism a public health crisis or emergency, the Anti-Racism in Public Health Act of 2020 reflects a shifting paradigm in which racism is considered a social determinant of health. Indeed, health inequities fundamentally rooted in structural racism have been exacerbated by the Covid-19 pandemic, which calls for the integration of antiracist praxis to promote ethical public health research processes. This commentary describes ways in which antiracist praxis-which emphasizes empowerment of traditionally marginalized populations-offers strategies to explicitly address power imbalance, stigmatization, and other consequences of structural racism in public health research.

Meat Consumption and Meat Cooking Practices in Essential Tremor: A Population-Based Study in the Faroe Islands.

Background: Elevated tissue levels of the tremor-producing neurotoxin, harmane, have been detected in patients with essential tremor (ET) in the USA and Spain. Recently, a study in the Faroe Islands similarly noted an elevation in blood harmane concentrations in probable and definite ET cases. The underlying mechanism is not understood. Possible mechanisms include increased dietary consumption (esp. through cooked meats), impaired metabolism, or increased endogenous production of harmane. To investigate this issue further, we conducted a population-based study in the Faroe Islands to examine meat consumption and meat cooking practices in ET cases and controls. Methods: 1,328 Faroese adults were screened for tremor and 27 ET cases were identified. Meat consumption and meat cooking practices were compared to 200 controls. Detailed data were collected via questionnaires regarding current meat consumption for 14 meat types and meat cooking doneness for 8 meat types. Data were also available on blood harmane concentrations. Results: Current meat consumption was similar in ET cases and controls in 12 out of 14 meat types, with no differences observed after a Bonferroni correction in any meat type; no difference was observed when stratified by gender. No difference was observed in meat doneness between ET cases and controls. Blood harmane concentrations were not correlated with dietary data. Discussion: This is the first population-based study of harmane-linked dietary factors in ET. The study suggests the observed difference in blood harmane in ET is not driven by dietary differences and is likely due to other mechanisms (e.g., impaired metabolism).

Increased beta-amyloid levels in the choroid plexus following lead exposure and the involvement of low-density lipoprotein receptor protein-1.

The choroid plexus, a barrier between the blood and cerebrospinal fluid (CSF), is known to accumulate lead (Pb) and also possibly function to maintain brain's homeostasis of Abeta, an important peptide in the etiology of Alzheimer's disease. This study was designed to investigate if Pb exposure altered Abeta levels at the blood-CSF barrier in the choroid plexus. Rats received ip injection of 27 mg Pb/kg. Twenty-four hours later, a FAM-labeled Abeta (200 pmol) was infused into the lateral ventricle and the plexus tissues were removed to quantify Abeta accumulation. Results revealed a significant increase in intracellular Abeta accumulation in the Pb-exposed animals compared to controls (p<0.001). When choroidal epithelial Z310 cells were treated with 10 microM Pb for 24 h and 48 h, Abeta (2 microM in culture medium) accumulation was significantly increased by 1.5 fold (p<0.05) and 1.8 fold (p<0.05), respectively. To explore the mechanism, we examined the effect of Pb on low-density lipoprotein receptor protein-1 (LRP1), an intracellular Abeta transport protein. Following acute Pb exposure with the aforementioned dose regimen, levels of LRP1 mRNA and proteins in the choroid plexus were decreased by 35% (p<0.05) and 31.8% (p<0.05), respectively, in comparison to those of controls. In Z310 cells exposed to 10 microM Pb for 24 h and 48 h, a 33.1% and 33.4% decrease in the protein expression of LRP1 was observed (p<0.05), respectively. Knocking down LRP1 resulted in even more substantial increases of cellular accumulation of Abeta, from 31% in cells without knockdown to 72% in cells with LRP1 knockdown (p<0.05). Taken together, these results suggest that the acute exposure to Pb results in an increased accumulation of intracellular Abeta in the choroid plexus; the effect appears to be mediated, at least in part, via suppression of LRP1 production following Pb exposure.

Cancer and blood concentrations of the comutagen harmane in essential tremor.

Blood concentrations of harmane, a tremor-producing neurotoxin, are elevated in essential tremor (ET). Harmane is also a comutagen. Using a case-control design, we compared the prevalence of cancer in ET cases vs. controls, and determined whether blood harmane concentrations are elevated among ET cases with cancer. 66/267 (24.7%) ET cases vs. 55/331 (16.6%) controls had cancer (adjusted OR 1.52, 95% CI 1.01-2.30, P = 0.04). Among specific cancer types, colon cancer was more prevalent in ET cases than controls (2.6% vs. 0.6%, P = 0.04). Log blood harmane concentration was higher in ET cases vs. controls (P = 0.02) and in participants with vs. without cancer (P = 0.02). Log blood harmane concentration was highest in ET cases with cancer when compared with other groups (P = 0.009). These links between cancer and ET and between high blood harmane and cancer in ET deserve further study.

Manganese accumulates primarily in nuclei of cultured brain cells.

Manganese (Mn) is known to pass across the blood-brain barrier and interact with dopaminergic neurons. However, the knowledge on the subcellular distribution of Mn in these cell types upon exposure to Mn remained incomplete. This study was designed to investigate the subcellular distribution of Mn in blood-brain barrier endothelial RBE4 cells, blood-cerebrospinal fluid barrier choroidal epithelial Z310 cells, mesencephalic dopaminergic neuronal N27 cells, and pheochromocytoma dopaminergic PC12 cells. The cells were incubated with 100 microM MnCl(2) with radioactive tracer (54)Mn in the culture media for 24h. The subcellular organelles, i.e., nuclei, mitochondria, microsomes, and cytoplasm, were isolated by centrifugation and verified for their authenticity by determining the markers specific to cellular organelles. Data indicated that maximum Mn accumulation was observed in PC12 cells, which was 2.8, 5.2- and 5.9-fold higher than that in N27, Z310 and RBE4 cells, respectively. Within cells, about 92%, 72%, and 52% of intracellular (54)Mn were found to be present in nuclei of RBE4, Z310, and N27 cells, respectively. The recovery of (54)Mn in nuclei and cytoplasm of PC12 cells were 27% and 69%, respectively. Surprisingly, less than 0.5% and 2.5% of cellular (54)Mn was found in mitochondrial and microsomal fractions, respectively. This study suggests that the nuclei may serve as the primary pool for intracellular Mn; mitochondria and microsomes may play an insignificant role in Mn subcellular distribution.

Elevated blood harmane (1-methyl-9H-pyrido[3,4-b]indole) concentrations in essential tremor.

Essential tremor (ET) is a widespread late-life neurological disease. Genetic and environmental factors likely play an etiological role. Harmane (1-methyl-9H-pyrido[3,4-b]indole) is a potent tremor-producing neurotoxin. In 2002, we demonstrated elevated blood harmane concentrations in an initial sample of 100 ET cases compared to 100 controls. Between 2002 and 2007, we assembled a new and larger sample of ET cases and controls. We now attempt to replicate our previous findings. Cases and controls were frequency-matched on age, gender, and race. Blood harmane concentrations were quantified by high-performance liquid chromatography. Subjects comprised 150 ET cases and 135 controls (mean age 65.3+/-15.5 vs. 65.5+/-14.2 years, p=0.94). Mean log blood harmane concentration was approximately 50% higher in cases than controls (0.50+/-0.54g(-10)/ml vs. 0.35+/-0.62g(-10)/ml, p=0.038). In a logistic regression analysis, log blood harmane concentration was associated with ET (OR(adjusted) 1.56, 95% CI 1.01-2.42, p=0.04), and odds of ET was 1.90 (95% CI 1.07-3.39, p=0.029) in the highest versus lowest log blood harmane tertile. Log blood harmane was highest in ET cases with familial ET (0.53+/-0.57g(-10)/ml), intermediate in cases with sporadic ET (0.43+/-0.45g(-10)/ml) and lowest in controls (0.35+/-0.62g(-10)/ml) (test for trend, p=0.026). Blood harmane appears to be elevated in ET. The higher concentrations in familial ET suggests that the mechanism may involve genetic factors.

Higher blood harmane (1-methyl-9H-pyrido[3,4-b]indole) concentrations correlate with lower olfactory scores in essential tremor.

BACKGROUND: Harmane (1-methyl-9H-pyrido[3,4-b]indole), a neurotoxin, may be an environmental risk factor for essential tremor (ET). Harmane and related chemicals are toxic to the cerebellum. Whether it is through this mechanism (cerebellar toxicity) that harmane leads to ET is unknown. Impaired olfaction may be a feature of cerebellar disease. OBJECTIVE: To determine whether blood harmane concentrations correlate with olfactory test scores in patients with ET. METHODS: Blood harmane concentrations were quantified using high performance liquid chromatography. Odor identification testing was performed with the University of Pennsylvania Smell Identification Test (UPSIT). RESULTS: In 83 ET cases, higher log blood harmane concentration was correlated with lower UPSIT score (rho=-0.46, p<0.001). 25/40 (62.5%) cases with high log blood harmane concentration (based on a median split) had low UPSIT scores (based on a median split) vs. 12/43 (27.9%) ET cases with low log blood harmane concentration (adjusted odd ratios (OR) 4.04, 95% confidence intervals (CI) 1.42-11.50, p=0.009). When compared with the low log blood harmane tertile, the odds of olfactory dysfunction were 2.64 times higher in cases in the middle tertile and 10.95 times higher in cases in the high tertile. In 69 control subjects, higher log blood harmane concentration was not correlated with lower UPSIT score (rho=0.12, p=0.32). CONCLUSIONS: Blood harmane concentrations were correlated with UPSIT scores in ET cases but not controls. These analyses set the stage for postmortem studies to further explore the role of harmane as a cerebellar toxin in ET.

Ozone therapy for diabetic foot.

Diabetic foot ulcers (DFU) are a burden to the diabetic community. With increasing medical bills, to unsuccessful treatment, those suffering from DFUs can use alternative therapeutics. First seen in the mid-1800s, ozone (O3) is thought to be unstable, due to inherent molecular nature. With the help of pharmaceutical science, various O3 treatments have flourished in the medical community to help those suffering from DFUs. Promising results are seen through numerous studies. Usually, a mixture of both O2 and O3 is seen in pressurized machines as administered to the foot ulcer. Foot ulcers, specifically DFUs, need to be assessed, cleaned, and treated as fast as possible for the fastest results. Results such as amputation can be seen if the foot is not attended to as soon as possible. With fast growing clinical trials in O3 therapy and quick administration of the O3, O3 therapy may be on the rise to be at the forefront of treating DFUs. Compelling evidence is seen in clinical trials, but more must be done to fully understand the role of O3 in DFUs.

Clinical utility of ozone therapy for musculoskeletal disorders.

Oxygen-ozone (O3) therapy serves as an alternative medical technique that increases the oxygen in the body along with the introduction of O3. O3 therapy has finally reached a level where the biological mechanisms of action have been understood, showing that they are in the domain of physiology, biochemistry, and pharmacology. Few clinical applications have been reviewed here as well as exemplifying that O3 therapy is particularly useful in musculoskeletal disorders. In the therapeutic range, O3 can be used as a more effective and safe substitute of standard medications. O3 therapy has been used for many years for its ability to inactivate various viruses, cancer, and acquired immune deficiency syndrome but is now making strides in the treatment of musculoskeletal disorders such as rheumatoid arthritis, lumbar facet joint syndrome, subacromial bursitis, carpal tunnel syndrome, osteoarthritis, hip bursitis, shoulder adhesive capsulitis, herniated disc, and temporomandibular joint disorder.

Subchronic Manganese Exposure Impairs Neurogenesis in the Adult Rat Hippocampus.

Adult neurogenesis takes place in the brain subventricular zone (SVZ) in the lateral walls of lateral ventricles and subgranular zone (SGZ) in the hippocampal dentate gyrus (HDG), and functions to supply newborn neurons for normal brain functionality. Subchronic Mn exposure is known to disrupt adult neurogenesis in the SVZ. This study was designed to determine whether Mn exposure disturbed neurogenesis within the adult HDG. Adult rats (10 weeks old) received a single dose of bromodeoxyuridine (BrdU) at the end of 4-week Mn exposure to label the proliferating cells. Immunostaining and cell counting data showed that BrdU(+) cells in Mn-exposed HDG were about 37% lower than that in the control (p < .05). The majority of BrdU(+) cells were identified as Sox2(+) cells. Another set of adult rats received BrdU injections for 3 consecutive days followed by 2- or 4-week Mn exposure to trace the fate of BrdU-labeled cells in the HDG. The time course studies indicated that Mn exposure significantly reduced the survival rate (54% at 2 weeks and 33% at 4 weeks), as compared with that in the control (80% at 2 weeks and 51% at 4 weeks) (p < .01). A significant time-dependent migration of newborn cells from the SGZ toward the granule cell layer was also observed in both control and Mn-exposed HDG. Triple-stained neuroblasts and mature neurons further revealed that Mn exposure significantly inhibited the differentiation of immature neuroblasts into mature neurons in the HDG. Taken together, these observations suggest that subchronic Mn exposure results in a reduced cell proliferation, diminished survival of adult-born neurons, and inhibited overall neurogenesis in the adult HDG. Impaired adult neurogenesis is likely one of the mechanisms contribute to Mn-induced Parkinsonian disorder.

Quantification of the neurotoxic beta-carboline harmane in barbecued/grilled meat samples and correlation with level of doneness.

Harmane, one of the heterocyclic amines (HCAs), is a potent neurotoxin linked to human diseases. Dietary exposure, especially in cooked meats, is the major source of exogenous exposure for humans. However, knowledge of harmane concentrations in cooked meat samples is limited. Our goals were to (1) quantify the concentration of harmane in different types of cooked meat samples, (2) compare its concentration to that of other more well-understood HCAs, and (3) examine the relationship between harmane concentration and level of doneness. Thirty barbecued/grilled meat samples (8 beef steak, 12 hamburger, 10 chicken) were analyzed for harmane and four other HCAs (2-amino-1-methyl-6-phenylimidazo [4,5-b]pyridine [PhIP], amino-3,8-dimethylimidazo[4,5-f]quinoxaline [MeIQx], 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline [DiMeIQx], and 2-amino-1,6-dimethylfuro[3,2-e]imidazo[4,5-b]pyridine [IFP]). Mean (+/- SD) harmane concentration was 5.63 (+/- 6.63) ng/g; harmane concentration was highest in chicken (8.48 +/- 9.86 ng/g) and lowest in beef steak (3.80 +/- 3.6 ng/g). Harmane concentration was higher than that of the other HCAs and significantly correlated with PhIP concentration. Harmane concentration was associated with meat doneness in samples of cooked beef steak and hamburger, although the correlation between meat doneness and concentration was greater for PhIP than for harmane. Evidence indicates that harmane was detectable in nanograms per gram quantities in cooked meat (especially chicken) and, moreover, was more abundant than other HCAs. There was some correlation between meat doneness and harmane concentration, although this correlation was less robust than that observed for PhIP. Data such as these may be used to improve estimation of human dietary exposure to this neurotoxin.

Aberrant Adult Neurogenesis in the Subventricular Zone-Rostral Migratory Stream-Olfactory Bulb System Following Subchronic Manganese Exposure.

Adult neurogenesis occurs in brain subventricular zone (SVZ). Our recent data reveal an elevated proliferation of BrdU(+) cells in SVZ following subchronic manganese (Mn) exposure in rats. This study was designed to distinguish Mn effect on the critical stage of adult neurogenesis, ie, proliferation, migration, survival and differentiation from the SVZ via the rostral migratory stream to the olfactory bulb (OB). Adult rats received a single ip-dose of BrdU at the end of 4-week Mn exposure to label proliferating cells. Immunostaining and cell-counting showed a 48% increase of BrdU(+) cells in Mn-exposed SVZ than in controls (P< .05). These BrdU(+) cells were identified as a mixed population of mainly GFAP(+) type-B neural stem cells, Nestin(+) type-C transit progenitor cells, DCX(+) migratory neuroblasts and Iba1(+) microglial cells. Another group of adult rats received 3 daily ip-injections of BrdU followed by subchronic Mn exposure. By 4-week post BrdU labeling, most of the surviving BrdU(+) cells in the OB were differentiated into NeuN(+) matured neurons. However, survival rates of BrdU/NeuN/DAPI triple-labeled cells in OB were 33% and 64% in Mn-exposed and control animals, respectively (P< .01). Infusion of Cu directly into the lateral ventricle significantly decreased the cell proliferation in the SVZ. Taken together, these results suggest that Mn exposure initially enhances the cell proliferation in adult SVZ. In the OB, however, Mn exposure significantly reduces the surviving adult-born cells and markedly inhibits their differentiation into mature neurons, resulting in an overall decreased adult neurogenesis in the OB.

Age-dependent increase of brain copper levels and expressions of copper regulatory proteins in the subventricular zone and choroid plexus.

Our recent data suggest a high accumulation of copper (Cu) in the subventricular zone (SVZ) along the wall of brain ventricles. Anatomically, SVZ is in direct contact with cerebrospinal fluid (CSF), which is secreted by a neighboring tissue choroid plexus (CP). Changes in Cu regulatory gene expressions in the SVZ and CP as the function of aging may determine Cu levels in the CSF and SVZ. This study was designed to investigate the associations between age, Cu levels, and Cu regulatory genes in SVZ and plexus. The SVZ and CP were dissected from brains of 3-week, 10-week, or 9-month old male rats. Analyses by atomic absorption spectroscopy revealed that the SVZ of adult and old animals contained the highest Cu level compared with other tested brain regions. Significantly positive correlations between age and Cu levels in SVZ and plexus were observed; the SVZ Cu level of old animals was 7.5- and 5.8-fold higher than those of young and adult rats (p < 0.01), respectively. Quantitation by qPCR of the transcriptional expressions of Cu regulatory proteins showed that the SVZ expressed the highest level of Cu storage protein metallothioneins (MTs), while the CP expressed the high level of Cu transporter protein Ctr1. Noticeably, Cu levels in the SVZ were positively associated with type B slow proliferating cell marker Gfap (p < 0.05), but inversely associated with type A proliferating neuroblast marker Dcx (p < 0.05) and type C transit amplifying progenitor marker Nestin (p < 0.01). Dmt1 had significant positive correlations with age and Cu levels in the plexus (p < 0.01). These findings suggest that Cu levels in all tested brain regions are increased as the function of age. The SVZ shows a different expression pattern of Cu-regulatory genes from the CP. The age-related increase of MTs and decrease of Ctr1 may contribute to the high Cu level in this neurogenesis active brain region.

Elevated adult neurogenesis in brain subventricular zone following in vivo manganese exposure: roles of copper and DMT1.

The brain subventricular zone (SVZ) is a source of neural precursor cells; these cells travel along the rostral migratory stream (RMS) to destination areas in the process of adult neurogenesis. Recent x-ray fluorescence (XRF) studies reveal an extensive accumulation of copper (Cu) in the SVZ. Earlier human and animal studies also suggest an altered Cu homeostasis after manganese (Mn) exposure. This study was designed to test the hypothesis that Mn exposure by acting on the divalent metal transporter-1 (DMT1) altered Cu levels in SVZ and RMS, thereby affecting adult neurogenesis. Adult rats received intraperitoneal (i.p.) injections of 6 mg Mn/kg as MnCl2 once daily for 4 weeks with concomitant injections of bromodeoxyuridine (BrdU) for 5 days in the last week. In control rats, Cu levels were significantly higher in the SVZ than other brain regions examined. Mn exposure significantly reduced Cu concentrations in the SVZ (P < 0.01). Immunohistochemical data showed that in vivo Mn exposure significantly increased numbers of BrdU(+) cells, which were accompanied with increased GFAP(+) astrocytic stem cells and DCX(+) neuroblasts in SVZ and RMS. Quantitative RT-PCR and Western blot confirmed the increased expression of DMT1 in SVZ following in vivo Mn exposure, which contributed to Mn accumulation in the neurogenesis pathway. Taken together, these results indicate a clear disruptive effect of Mn on adult neurogenesis; the effect appears due partly to Mn induction of DMT1 and its interference with cellular Cu regulation in SVZ and RMS. The future research directions based on these observations are also discussed.