Study of the Serum Copper Levels in Patients with Major Depressive Disorder

Copper may be involved in the pathophysiology of depression. Clinical data on this issue are very limited and not conclusive. The purpose of the study was to determine the copper concentration in the serum of patients with major depressive disorder and to discuss its potential clinical usefulness as a biomarker of the disease. A case–control clinical study included 69 patients with current depressive episode, 45 patients in remission and 50 healthy volunteers. Cu concentration was measured by electrothermal atomic absorption spectrometry (ETAAS). The mean serum copper level in depressed patients was slightly lower (by 11 %; not statistically significant) than in the control group. Furthermore, there was no significant difference in Cu²⁺ concentration between depressive episode and remission, nor between remission and control group. In the remission group were observed significant correlations between copper levels and the average number of relapses over the past years or time of remission. There was no correlation between serum copper and severity of depression, as measured by HDRS and MADRS. The obtained results showed no significant differences between the copper concentration in the blood serum of patients (both with current depressive episode and in remission) and healthy volunteers, as well as the lack of correlations between the copper level in the active stage of the disease and clinical features of the population. Our study is the first conducted on such a large population of patients, so the results may be particularly important and reliable source of knowledge about the potential role of copper in depression.     

Metabolic changes in the visual cortex are linked to retinal nerve fiber layer thinning in multiple sclerosis

Objective

To investigate the damage to the retinal nerve fiber layer as part of the anterior visual pathway as well as an impairment of the neuronal and axonal integrity in the visual cortex as part of the posterior visual pathway with complementary neuroimaging techniques, and to correlate our results to patients'' clinical symptoms concerning the visual pathway.

Design, Subjects and Methods

Survey of 86 patients with relapsing-remitting multiple sclerosis that were subjected to retinal nerve fiber layer thickness (RNFLT) measurement by optical coherence tomography, to a routine MRI scan including the calculation of the brain parenchymal fraction (BPF), and to magnetic resonance spectroscopy at 3 tesla, quantifying N-acetyl aspartate (NAA) concentrations in the visual cortex and normal-appearing white matter.

Results

RNFLT correlated significantly with BPF and visual cortex NAA, but not with normal-appearing white matter NAA. This was connected with the patients'' history of a previous optic neuritis. In a combined model, both BPF and visual cortex NAA were independently associated with RNFLT.

Conclusions

Our data suggest the existence of functional pathway-specific damage patterns exceeding global neurodegeneration. They suggest a strong interrelationship between damage to the anterior and the posterior visual pathway.     

The nerve growth factor precursor proNGF exhibits neurotrophic activity but is less active than mature nerve growth factor

Nerve growth factor (NGF) promotes neuronal survival and differentiation and stimulates neurite outgrowth. NGF is synthesized as a precursor, proNGF, which undergoes post-translational processing to generate mature beta-NGF. It has been assumed that, in vivo, NGF is largely processed into the mature form and that mature NGF accounts for the biological activity. However, we recently showed that proNGF is abundant in CNS tissues whereas mature NGF is undetectable, suggesting that proNGF has biological functions beyond its role as a precursor. To determine whether proNGF exhibits biological activity, we mutagenized the precursor-processing site and expressed unprocessed, cleavage-resistant proNGF protein in insect cells. Survival and neurite outgrowth assays on murine superior cervical ganglion neurons and PC12 cells indicated that proNGF exhibits neurotrophic activity similar to mature 2.5S NGF, but is approximately fivefold less active. ProNGF binds to the high-affinity receptor, TrkA, as determined by cross-linking to PC12 cells, and is also slightly less active than mature NGF in promoting phosphorylation of TrkA and its downstream signaling effectors, Erk1/2, in PC12 and NIH3T3-TrkA cells. These data, coupled with our previous report that proNGF is the major form of NGF in the CNS, suggest that proNGF could be responsible for much of the biological activity normally attributed to mature NGF in vivo.     

Enterobacterial 38-kDa outer membrane protein is an age-dependent molecular marker of innate immunity and immunoglobulin deficiency as results from its reactivity with IgG and IgA antibody

In earlier studies on an animal model we observed protective properties of outer membrane proteins (OMPs) of Shigella, Hafnia, and Escherichia coli strains. In order to investigate human sera for reactivity with OMPs we subjected these proteins to immunoblotting with umbilical cord plasma and sera from children and adults. The IgG and IgA antibodies interacted primarily with a 38-kDa protein, in similar way for several enterobacterial strains, but different for Pseudomonas aeruginosa. This observation prompted us to determine the reactivity with the purified 38-kDa OMP in the sera of several groups of children. The reactivity of the protein from Shigella flexneri serotype 3a with sera in ELISA was age dependent, increasing from low reactivity in infants to the adult antibody level. The IgG and IgA antibody specific response thus revealed the normal pattern of immunity. The level of IgA and IgG antibody was significantly low in child patients with IgA and/or IgG immunoglobulin deficiencies, but was at the healthy control level in children with recurrent respiratory tract inflammation. These data correlated with total IgA and IgG levels in immunoglobulin-deficient children. The results indicate that this protein may serve as an immunodiagnostic marker, but also as an antigen carrier in vaccines.     

Dendritic Cells as an Alternate Approach for Treatment of Neurodegenerative Disorders

Despite years of research, Alzheimer’s disease (AD) remains incurable and thus poses a major health challenge in coming years. This neurodegenerative disease belongs to a heterogeneous group of human tauopathies, characterized by the extracellular deposition of beta amyloid-Aβ and intracellular accumulation of tau protein in neuronal and glial cells, whereby tau pathology best correlates with disease progression. For decades, several disease-modifying agents were brought to clinical studies with promising efficacy in preclinical trials; however, all of the subsequent clinical trials failed. Therefore, the pursuit for therapeutic agents for the treatment of AD and other tauopathies still continue. Recent evidences show previously unidentified role of peripheral immune system in regulating the inflammatory status of the brain, mainly the dendritic cells. A decrease in functionality and count of dendritic cells has been observed in Alzheimer’s disease. Here, we discuss a potential role of dendritic cell-based vaccines as therapeutic approach in ameliorating disease pathogenesis in AD and other tauopathies.     

Proton Gradient Regulation5-Like1-Mediated Cyclic Electron Flow Is Crucial for Acclimation to Anoxia and Complementary to Nonphotochemical Quenching in Stress Adaptation

To investigate the functional importance of Proton Gradient Regulation5-Like1 (PGRL1) for photosynthetic performances in the moss Physcomitrella patens, we generated a pgrl1 knockout mutant. Functional analysis revealed diminished nonphotochemical quenching (NPQ) as well as decreased capacity for cyclic electron flow (CEF) in pgrl1. Under anoxia, where CEF is induced, quantitative proteomics evidenced severe down-regulation of photosystems but up-regulation of the chloroplast NADH dehydrogenase complex, plastocyanin, and Ca²⁺ sensors in the mutant, indicating that the absence of PGRL1 triggered a mechanism compensatory for diminished CEF. On the other hand, proteins required for NPQ, such as light-harvesting complex stress-related protein1 (LHCSR1), violaxanthin de-epoxidase, and PSII subunit S, remained stable. To further investigate the interrelation between CEF and NPQ, we generated a pgrl1 npq4 double mutant in the green alga Chlamydomonas reinhardtii lacking both PGRL1 and LHCSR3 expression. Phenotypic comparative analyses of this double mutant, together with the single knockout strains and with the P. patens pgrl1, demonstrated that PGRL1 is crucial for acclimation to high light and anoxia in both organisms. Moreover, the data generated for the C. reinhardtii double mutant clearly showed a complementary role of PGRL1 and LHCSR3 in managing high light stress response. We conclude that both proteins are needed for photoprotection and for survival under low oxygen, underpinning a tight link between CEF and NPQ in oxygenic photosynthesis. Given the complementarity of the energy-dependent component of NPQ (qE) and PGRL1-mediated CEF, we suggest that PGRL1 is a capacitor linked to the evolution of the PSII subunit S-dependent qE in terrestrial plants.The conversion of solar energy into chemical energy and building material by oxygenic photosynthesis, as performed by plants, green algae, and cyanobacteria, supports much of the life on our planet. The production of oxygen and the assimilation of carbon dioxide into organic matter determines, to a large extent, the composition of our atmosphere. Plant photosynthesis is achieved thanks to a series of reactions that occur mainly in the chloroplast, resulting in light-dependent water oxidation, NADP⁺ reduction, and ATP formation (Whatley et al., 1963). Two separate photosystems (PSI and PSII) and an ATP synthase (ATPase) embedded in the thylakoid membrane catalyze these reactions. The ATPase produces ATP at the expense of the proton motive force that is generated by the light reactions (Mitchell, 1961). The cytochrome (cyt) b₆f complex assures the link between the two photosystems by transferring electrons from the membrane-bound plastoquinone to a soluble carrier, plastocyanin, or cyt c₆ and functions in the pumping of protons. NADPH and ATP that are produced by linear electron flow from PSII to PSI are fueled into the Calvin Benson Bassham cycle (Bassham et al., 1950) to fix CO₂. In parallel, cyclic electron flow (CEF) between the cyt b₆f complex and PSI may occur, which would solely lead to the production of ATP. CEF around PSI has been first recognized by Arnon (1959) and is involved in the reequilibration of the ATP poise and prevention of overreduction of the PSI acceptor side (Alric, 2010; Peltier et al., 2010; Leister and Shikanai, 2013; Shikanai, 2014). In microalgae and vascular plants, CEF operates via an NAD(P)H dehydrogenase-like complex (NDH)-dependent and/or PROTON GRADIENT REGULATION5 (PGR5)-related pathway (Alric, 2010; Peltier et al., 2010; Leister and Shikanai, 2013; Shikanai, 2014). The thylakoid protein Proton Gradient Regulation5-Like1 (PGRL1; DalCorso et al., 2008) has been first discovered as a novel component for the PGR5-dependent CEF pathway in Arabidopsis (Arabidopsis thaliana), as its knockout causes a PGR5-like photosynthetic phenotype and is suggested to operate as a ferredoxin-plastoquinone reductase (Hertle et al., 2013). PGRL1 is also important for efficient CEF in the green alga Chlamydomonas reinhardtii, which becomes particularly evident under settings where CEF is induced, such as in acclimation to iron deficiency, high light (HL), or anaerobic growth conditions (Petroutsos et al., 2009; Iwai et al., 2010; Tolleter et al., 2011; Terashima et al., 2012). Remarkably, a CEF protein supercomplex composed of PSI-light-harvesting complex I (LHCI), LHCII, the cyt b₆f complex, ferredoxin-NADPH oxidoreductase, and PGRL1 was isolated from state 2 conditions (Iwai et al., 2010). Under anaerobic conditions, the Ca²⁺ sensor CAS and Anaerobic response1 (ANR1) were shown to interact with PGRL1 in vivo (Terashima et al., 2012) and were found to be associated with the C. reinhardtii CEF supercomplex. Consistently, depletion of CAS and ANR1 by artificial microRNA expression in C. reinhardtii resulted in strong inhibition of CEF under anoxia, which could be partially rescued by an increase in the extracellular Ca²⁺ concentration, inferring that CEF is Ca²⁺ dependent (Terashima et al., 2012). Notably, the regulation of the proton motive force by a two-pore potassium channel in the thylakoid membrane of Arabidopsis (AtTPK3), is also Ca²⁺ dependent (Carraretto et al., 2013), suggesting that Ca²⁺-dependent activation of CEF and the channel may work hand in hand.qE, the energy-dependent component of nonphotochemical quenching (NPQ) that occurs due to thermal dissipation of excess absorbed light energy (Li et al., 2000; Peers et al., 2009), is dependent on rapid luminal acidification upon photosynthetic electron transfer (Wraight and Crofts, 1970; Li et al., 2000). Thus, processes such as CEF that contribute to the pH gradient across the thylakoid membrane are interrelated to NPQ, as an acidified lumen is required for efficient qE (Joliot and Finazzi, 2010). In vascular plants, PSII subunit S (PSBS) is essential for efficient qE (Li et al., 2000), whereas qE induction in the green alga C. reinhardtii is mediated by light-harvesting complex stress-related protein3 (LHCSR3), an ancient light-harvesting protein that is missing in vascular plants (Peers et al., 2009). The moss Physcomitrella patens, which possesses genes encoding for PSBS and LHCSR proteins, utilizes both types of regulatory proteins to operate qE (Alboresi et al., 2010), suggesting that land plants evolved a novel PSBS-dependent qE mechanism before losing the ancestral LHCSR-dependent qE found in algae. This makes mosses a very interesting subject for investigating the interrelation and evolution of the CEF and NPQ molecular effectors.Mosses diverged from vascular plants early after land colonization and are one of the oldest groups of land plants present on earth. This places the moss model system P. patens (Rensing et al., 2008) evolutionarily in the middle between algae and vascular plants and makes it an ideal model organism for the study of the evolution of photosynthetic organisms. Analysis of photosynthesis in P. patens can provide insights into the events leading to adaptation to the harsher physicochemical conditions of the terrestrial environment (Rensing et al., 2008), as evidenced by the presence and functional overlap of LHCSRs and PSBS (Alboresi et al., 2010).To obtain insights into the interrelation and evolution of CEF and NPQ, we knocked out the PGRL1 gene from P. patens and analyzed functional phenotypic consequences. Moreover, we compared these phenotypes with phenotypic analyses of C. reinhardtii pgrl1, npq4, and pgrl1 npq4 single and double mutants lacking PGRL1, LHCSR3, and both PGRL1 and LHCSR3, respectively. The data provided strong evidence that the green cut protein PGRL1 (Karpowicz et al., 2011) is required for acclimation to anoxia both in algae and mosses. Moreover, an involvement of PGRL1 in the evolution of PSBS-dependent qE in terrestrial plants is implied.     

Decrease in salivary lactoferrin output in chronically intoxicated alcohol-dependent patients

Salivary lactoferrin is a glycoprotein involved in the elimination of pathogens and the prevention of massive overgrowth of microorganisms that affect oral and general health. A high concentration of lactoferrin in saliva is often considered to be a marker of damage to the salivary glands, gingivitis, or leakage through inflamed or damaged oral mucosa, infiltrated particularly by neutrophils. We conducted a study to determine the effect of chronic alcohol intoxication on salivary lactoferrin concentration and output. The study included 30 volunteers consisting of ten non-smoking male patients after chronic alcohol intoxication (group A), and 20 control nonsmoking male social drinkers (group C) with no history of alcohol abuse. Resting whole saliva was collected 24 to 48 hours after a chronic alcohol intoxication period. Lactoferrin was assessed by enzyme-linked immunosorbent assay. For all participants, the DMFT index (decayed, missing, or filled teeth), gingival index (GI) and papilla bleeding index (PBI) were assessed. The differences between groups were evaluated using the Mann-Whitney U test. We noticed significantly decreased salivary flow (SF) in alcohol dependent patients after chronic alcohol intoxication (A), compared to the control group (C). Although there was no significant difference in salivary lactoferrin concentration between the alcohol dependent group A and the control group C, we found significantly decreased lactoferrin output in group A compared to group C. We found a significant correlation between the amount of daily alcohol use and a decrease in lactoferrin output. There was a significant increase in GI and a tendency of PBI to increase in group A compared to group C. We demonstrated that chronic alcohol intoxication decreases SF and lactoferrin output. The decreased lactoferrin output in persons chronically intoxicated by alcohol may be the result of lactoferrin exhaustion during drinking (due to its alcohol-related lower biosynthesis or higher catabolism) or to decreased function of neutrophils affected by the ethanol. The poorer periodontal state in alcohol dependent persons compared to controls may be a result of lower salivary flow and decreased protection of the oral cavity by lactoferrin.     

The influence of radiotherapy on ceruloplasmin and transferrin in whole blood of breast cancer patients

Ceruloplasmin and transferrin are proteins which play a potential role in the process of breast cancer development. These molecules contain Cu²⁺ (ceruloplasmin) or Fe³⁺ ions (transferrin) and thus constitute paramagnetic centers, which can be studied using electron paramagnetic resonance method. The aim of the study was to determine how paramagnetic centers in whole blood of breast cancer patients change under the influence of radiation therapy. Samples of whole blood were taken from 17 women with breast cancer treated with radiotherapy. The measurements were carried out at 170 K using X-band electron paramagnetic resonance (EPR) spectrometer Bruker EMX-10. Two distinct EPR lines, derived from high-spin Fe³⁺ in transferrin and Cu²⁺ from ceruloplasmin, were revealed in all frozen samples. The amplitude and integrated intensity of the EPR signal from Cu²⁺ in ceruloplasmin significantly decreased in all patients after the delivery of the radiation fraction. When comparing the integral intensity of the signal from Fe³⁺ in transferrin, three different situations were identified which are patient specific: a significant increase, an insignificant change, or a significant decrease after the irradiation. A decreased level of Cu²⁺ from ceruloplasmin in patients after radiotherapy means a low level of ceruloplasmin in the plasma or an increased content of reduced Cu⁺ ions. Differences in the integrated intensity of the EPR signal from transferrin translate directly into the amount of bound iron. The observed changes could indicate how well the organism fights against cancer and how easily it adapts to the situation of biochemical stress.     

Viral Hepatitis and Rapid Diagnostic Test Based Screening for HBsAg in HIV-infected Patients in Rural Tanzania

Background

Co-infection with hepatitis B virus (HBV) is highly prevalent in people living with HIV in Sub-Saharan Africa. Screening for HBV surface antigen (HBsAg) before initiation of combination antiretroviral therapy (cART) is recommended. However, it is not part of diagnostic routines in HIV programs in many resource-limited countries although patients could benefit from optimized antiretroviral therapy covering both infections. Screening could be facilitated by rapid diagnostic tests for HBsAg. Operating experience with these point of care devices in HIV-positive patients in Sub-Saharan Africa is largely lacking. We determined the prevalence of HBV and Hepatitis C virus (HCV) infection as well as the diagnostic accuracy of the rapid test device Determine HBsAg in an HIV cohort in rural Tanzania.

Methods

Prospectively collected blood samples from adult, HIV-1 positive and antiretroviral treatment-naïve patients in the Kilombero and Ulanga antiretroviral cohort (KIULARCO) in rural Tanzania were analyzed at the point of care with Determine HBsAg, a reference HBsAg EIA and an anti-HCV EIA.

Results

Samples of 272 patients were included. Median age was 38 years (interquartile range [IQR] 32–47), 169/272 (63%) subjects were females and median CD4+ count was 250 cells/µL (IQR 97–439). HBsAg was detected in 25/272 (9.2%, 95% confidence interval [CI] 6.2–13.0%) subjects. Of these, 7/25 (28%) were positive for HBeAg. Sensitivity of Determine HBsAg was rated at 96% (95% CI 82.8–99.6%) and specificity at 100% (95% CI, 98.9–100%). Antibodies to HCV (anti-HCV) were found in 10/272 (3.7%, 95% CI 2.0–6.4%) of patients.

Conclusion

This study reports a high prevalence of HBV in HIV-positive patients in a rural Tanzanian setting. The rapid diagnostic test Determine HBsAg is an accurate assay for screening for HBsAg in HIV-1 infected patients at the point of care and may further help to guide cART in Sub-Saharan Africa.