Differences in assembly or stability of complex I and other mitochondrial OXPHOS complexes in inherited complex I deficiency

NADH-ubiquinone oxidoreductase (complex I) deficiency is amongst the most encountered defects of the mitochondrial oxidative phosphorylation (OXPHOS) system and is associated with a wide variety of clinical signs and symptoms. Mutations in complex I nuclear structural genes are the most common cause of isolated complex I enzyme deficiencies. The cell biological consequences of such mutations are poorly understood. In this paper we have used blue native electrophoresis in order to study how different nuclear mutations affect the integrity of mitochondrial OXPHOS complexes in fibroblasts from 15 complex I-deficient patients. Our results show an important decrease in the levels of intact complex I in patients harboring mutations in nuclear-encoded complex I subunits, indicating that complex I assembly and/or stability is compromised. Different patterns of low molecular weight subcomplexes are present in these patients, suggesting that the formation of the peripheral arm is affected at an early assembly stage. Mutations in complex I genes can also affect the stability of other mitochondrial complexes, with a specific decrease of fully-assembled complex III in patients with mutations in NDUFS2 and NDUFS4. We have extended this analysis to patients with an isolated complex I deficiency in which no mutations in structural subunits have been found. In this group, we can discriminate between complex I assembly and catalytic defects attending to the fact whether there is a correlation between assembly/activity levels or not. This will help us to point more selectively to candidate genes for pathogenic mutations that could lead to an isolated complex I defect.     

The genetic toxicology of metal compounds: II. Enhancement of ultraviolet light-induced mutagenesis in Escherichia coli WP2

Salts of metals which are carcinogenic, noncarcinogenic, or of unknown carcinogenicity were assayed for their abilities to modulate ultraviolet (UV)-induced mutagenesis in Escherichia coli WP2. In addition to the previously reported comutagenic effect of arsenite, salts of three other compounds were found to enhance UV mutagenesis. CuCl2, MnCl2 (and a small effect by KMnO4), and NaMoO4 acted as comutagens in E coli WP2, which has wild-type DNA repair capability, but were much less comutagenic in the repair deficient strain WP2s (uvrA). The survival of irradiated or unirradiated cells was not affected by these compounds. No effects on UV mutagenesis were seen for 16 other metal compounds. We suggest that the comutagenic effects might occur either via metal-induced decreases in the fidelity of repair replication or (in the case of CuCl2) via metal-induced depurination.     

Phosphorylated KDR is expressed in the neoplastic and stromal elements of human renal tumours and shuttles from cell membrane to nucleus

Vascular endothelial growth factor (VEGF)-A is an important angiogenic factor in establishing the vasculature in renal cell carcinomas (RCCs). Since little is known about VEGF signalling in RCCs, the profile of phosphorylated KDR (pKDR) has been investigated and the intracellular location of the receptor has been examined in the present study. Using two monoclonal antibodies raised against the phosphorylated KDR epitopes (Y1059 and Y1214) known to mediate different VEGF functions, together with a commercial anti-KDR antibody and immunohistochemistry, the expression of pKDR was investigated in a series of normal (n = 25) and neoplastic kidneys (n = 54; clear cell n = 35; papillary n = 10; oncocytomas n = 8). pKDR was present in many tissue elements of both normal and neoplastic renal tissues, with strong expression in the cell membrane, cytoplasm, and nuclei of normal kidney and tumour cells, as well as endothelial cells in tumours of all histological types. Patterns and intensity were similar using both anti-pKDR antibodies. There was no significant correlation in clear cell carcinomas between pKDR expression and age (p = 0.57), tumour size (p = 0.2), gender (p = 0.59), grade (p = 0.2) or histological type (p = 0.36). To delineate further the intracellular processing that might account for the cellular distribution, confocal microscopy was also performed. Antibodies to the different phosphorylated epitopes demonstrated different intracellular staining patterns. This study shows that pKDR is present in a wide variety of renal tumours, suggesting that anti-VEGF therapy might have direct effects on tumour cells. It further suggests that cells traffic pKDR depending on the precise KDR tyrosines that are autophosphorylated in a manner that enables receptor activation to result in different functions.     

A novel isoform of the smooth muscle cell differentiation marker smoothelin

Studies on smooth muscle cell differentiation and those on vascular development in mouse and humans have long been hampered by the lack of suitable markers. Here we describe a novel, large isoform of smoothelin, a structural protein of differentiated, contractile smooth muscle cells. The protein, which is highly conserved in mouse and humans, shows homology with other cytoskeleton-associated smooth muscle cell proteins and contains an actinin-type actin-binding domain. Northern blot analysis from various mouse organs identified short and long smoothelin mRNA forms, which exhibit distinct tissue expression patterns. The short form is highly expressed in visceral muscle tissues such as intestine and stomach and is not detectable in brain, while the long mRNA form is expressed in all vascularized organs. These results may provide new tools and approaches to study both smooth muscle cell differentiation and proliferative vascular disease. Received: 25 August 1998 / Accepted: 19 October 1998     

Tumor necrosis factor-mediated inhibition of interleukin-18 in the brain: a clinical and experimental study in head-injured patients and in a murine model of closed head injury.

Tumor necrosis factor (TNF) and interleukin-(IL)-18 are important mediators of neuroinflammation after closed head injury (CHI). Both mediators have been previously found to be significantly elevated in the intracranial compartment after brain injury, both in patients as well as in experimental model systems. However, the interrelation and regulation of these crucial cytokines within the injured brain has not yet been investigated. The present study was designed to assess a potential regulation of intracranial IL-18 levels by TNF based on a clinical study in head-injured patients and an experimental model in mice. In the first part, we investigated the interrelationship between the daily TNF and IL-18 cerebrospinal fluid levels in 10 patients with severe CHI for up to 14 days after trauma. In the second part of the study, the potential TNF-dependent regulation of intracerebral IL-18 levels was further characterized in an experimental set-up in mice: (1) in a standardized model of CHI in TNF/lymphotoxin-alpha gene-deficient mice and wild-type (WT) littermates, and (2) by intracerebro-ventricular injection of mouse recombinant TNF in WT C57BL/6 mice. The results demonstrate an inverse correlation of intrathecal TNF and IL-18 levels in head-injured patients and a TNF-dependent inhibition of IL-18 after intracerebral injection in mice. These findings imply a potential new anti-inflammatory mechanism of TNF by attenuation of IL-18, thus confirming the proposed "dual" function of this cytokine in the pathophysiology of traumatic brain injury.     

Obsessive-compulsive bipolar comorbidity: focus on children and adolescents

BACKGROUND: Growing evidence documents the frequent co-morbidity between Obsessive Compulsive Disorder (OCD) and Bipolar Disorder (BP) in adults. The aim of the present study is to explore some clinical aspects of this interface in children and adolescents, as it appears in a setting of routine clinical practice. METHOD: The sample comprised 102 consecutively referred children and adolescents, both inpatients and outpatients, with BP, OCD or co-morbid BP-OCD during a 3-year period. The mean age was 14.2 (SD=3.2); 65 (63.7%) were males. Diagnoses and clinical features were collected by means of structured interview according to DSM-IV (DICA-R) and a rating scale for OCD (CY-BOCS). Clinical outcome was evaluated prospectively by means of clinical global impression (CGI) as part of routine clinical care, throughout the follow-up. RESULTS: Thirty-seven (36.3%) patients (21 males and 16 females) were diagnosed as BP, 35 (34.3%) patients (26 males and 9 females) were diagnosed as OCD and 30 (29.4%) patients (18 males and 12 females) were diagnosed as BP-OCD. BP II, was more frequent in the BP-OCD than in BP. When OCD was co-morbid with BP, age of onset was significantly earlier than in the 'pure' OCD patients. On the contrary, age of onset of BP was not affected by co-morbid OCD. According to CGI baseline scores, OCD patients were significantly less impaired than BP-OCD and BP patients, while the severity of the symptomatology was similar in the last two groups. Severity scores at the end of the follow-up were significantly higher in BP-OCD patients than in OCD patients. Patients with pure BP showed lower rates of panic disorder-agoraphobia than BP-OCD patients and higher rates of ADHD-conduct disorder. Pure OCD patients showed lower rates of ADHD and higher rates of Generalized Anxiety Disorder. The number of obsessions did not differentiate the two groups, whereas pure OCD patients showed significantly more compulsions. 'Other' obsessions-e.g., existential, philosophical, odd and/or superstitious-were significantly more frequent in BP-OCD than in pure OCD patients. Ordering compulsions were significantly more frequent in pure OCD patients. LIMITATIONS: Possible low reliability of children's and their parents' recall of past episodes of mental disorder. CONCLUSIONS: In a tertiary care center, co-morbidity between OCD and BP is a significant clinical problem affecting a large number of patients. The correct identification of OCD-bipolar co-morbidity has relevant clinical implications as far as other concomitant disorders, symptomatological features, course, complications, and treatment management and outcome are concerned.     

Effects of task difficulty and invested mental effort on peripheral vasoconstriction

We ran two experiments to investigate whether peripheral arterial tone reflects changes in mental effort. Finger pulse wave amplitude, interpulse interval, and pulse variability in the mid- and high-frequency bands were recorded by means of a newly developed finger plethysmograph during both rest and cognitive performance. Using a modified version of the Sternberg memory task, we selectively manipulated either the difficulty of the task (Experiment 1) or the subjects' level of engagement in the task (Experiment 2). We found a significant difference in finger pulse wave amplitude between rest and task periods, suggesting that the measure reflects changes in sympathetic activity due to task engagement. In addition, our results suggest that reduced pulse wave amplitude, signaling vasoconstriction, occurs when subjects are investing effort.