Imunofenotipagem e remodelamento da matriz extracelular na sarcoidose pulmonar e extrapulmonar

OBJETIVO: Investigar o significado de marcadores de imunidade celular e de componentes elásticos/colágeno da matriz extracelular em estruturas granulomatosas em biópsias de pacientes com sarcoidose pulmonar ou extrapulmonar. MÉTODOS: Determinações qualitativas e quantitativas de células inflamatórias, de fibras de colágeno e de fibras elásticas em estruturas granulomatosas em biópsias cirúrgicas de 40 pacientes com sarcoidose pulmonar e extrapulmonar foram realizadas por histomorfometria, imuno-histoquímica, e técnicas de coloração com picrosirius e resorcina-fucsina de Weigert. RESULTADOS: A densidade de linfócitos, macrófagos e neutrófilos nas biópsias extrapulmonares foi significativamente maior do que nas biópsias pulmonares. Os granulomas pulmonares apresentaram uma quantidade significativamente maior de fibras de colágeno e menor densidade de fibras elásticas que os granulomas extrapulmonares. A quantidade de macrófagos nos granulomas pulmonares correlacionou-se com CVF (p < 0,05), ao passo que as quantidades de linfócitos CD3+, CD4+ e CD8+ correlacionaram-se com a relação VEF1/CVF e com CV. Houve correlações negativas entre CPT e contagem de células CD1a+ (p < 0,05) e entre DLCO e densidade de fibras colágenas/elásticas (r = -0,90; p = 0,04). CONCLUSÕES: A imunofenotipagem e o remodelamento apresentaram características diferentes nas biópsias dos pacientes com sarcoidose pulmonar e extrapulmonar. Essas diferenças correlacionaram-se com os dados clínicos e espirométricos dos pacientes, sugerindo que há duas vias envolvidas no mecanismo de depuração de antígenos, que foi mais eficaz nos pulmões e linfonodos.

Biphasic positive airway pressure minimizes biological impact on lung tissue in mild acute lung injury independent of etiology

Abstract Introduction Biphasic positive airway pressure (BIVENT) is a partial support mode that employs pressure-controlled, time-cycled ventilation set at two levels of continuous positive airway pressure with unrestricted spontaneous breathing. BIVENT can modulate inspiratory effort by modifying the frequency of controlled breaths. Nevertheless, the optimal amount of inspiratory effort to improve respiratory function while minimizing ventilator-associated lung injury during partial ventilatory assistance has not been determined. Furthermore, it is unclear whether the effects of partial ventilatory support depend on acute lung injury (ALI) etiology. This study aimed to investigate the impact of spontaneous and time-cycled control breaths during BIVENT on the lung and diaphragm in experimental pulmonary (p) and extrapulmonary (exp) ALI. Methods This was a prospective, randomized, controlled experimental study of 60 adult male Wistar rats. Mild ALI was induced by Escherichia coli lipopolysaccharide either intratracheally (ALIp) or intraperitoneally (ALIexp). After 24 hours, animals were anesthetized and further randomized as follows: (1) pressure-controlled ventilation (PCV) with tidal volume (Vt) = 6 ml/kg, respiratory rate = 100 breaths/min, PEEP = 5 cmH2O, and inspiratory-to-expiratory ratio (I:E) = 1:2; or (2) BIVENT with three spontaneous and time-cycled control breath modes (100, 75, and 50 breaths/min). BIVENT was set with two levels of CPAP (Phigh = 10 cmH2O and Plow = 5 cmH2O). Inspiratory time was kept constant (Thigh = 0.3 s). Results BIVENT was associated with reduced markers of inflammation, apoptosis, fibrogenesis, and epithelial and endothelial cell damage in lung tissue in both ALI models when compared to PCV. The inspiratory effort during spontaneous breaths increased during BIVENT-50 in both ALI models. In ALIp, alveolar collapse was higher in BIVENT-100 than PCV, but decreased during BIVENT-50, and diaphragmatic injury was lower during BIVENT-50 compared to PCV and BIVENT-100. In ALIexp, alveolar collapse during BIVENT-100 and BIVENT-75 was comparable to PCV, while decreasing with BIVENT-50, and diaphragmatic injury increased during BIVENT-50. Conclusions In mild ALI, BIVENT had a lower biological impact on lung tissue compared to PCV. In contrast, the response of atelectasis and diaphragmatic injury to BIVENT differed according to the rate of spontaneous/controlled breaths and ALI etiology.

Influence of chest wall on lung mechanics during inspiration

RATIONALE: The interaction between lungs and chest wall influences lung volume, that determines lung history during respiration cycle. In this study, the influence of chest wall mechanics on respiratory system is assessed by the evaluation of inspiration pressure-volume curve (PV curve) under three different situations: closed-chest, open-chest and isolated lung. The PV curve parameters in each situation allow us to further understand the role played by different chest wall elements in the respiratory function. Methods: Twenty-four male Wistar rats (236 ± 29 g) were used. The animals were weighted and then anesthetized with xylazine 2% (O,SmL/kg) and ketamine 10% (0,9mL/kg), exsanguinated and later tracheostomies with a metallic cannula (14 gauge).The cannula was connected to an automatic small animal insufflator. This setup was connected to a pressure transducer (32 samples/s). The 24 animals were randomly separated in three groups:(i) closed chest,(ii) open chest and (iii) isolated lung. The rats were insufflated with 20mL quasi-statically (constant speed of 0,1mUs). lnsufflated volume and measured pressure data were kept and PV curves were obtained for all animals. The PV curves were fitted (non-linear least squares) against the sigmoid equation (1) to obtain the sigmoid equation parameters (a,b,c,d). Elastance measurements were obtained from linear regression of pressure/volume measurements in a 0,8s interval before and after the calculated point. Results: The parameters a,b and c showed no significant change, but the parameter d showed a significant variation among the three groups. The initial elastance also varied between open and closed chest, indicating the need of a higher pressure for the lung expansion, as can be seen in Table 1. Conclusion: A supporting effect of the chest wall was observed at the initial moments of inspiration, observed as a higher initial elastance in open chest situations than in closed chest situations (p=0,00001). The similar initial elastance for the isolated lung and closed chest may be explained by the specific method used for the isolated lung experiment. As the isolated lung is supported by the trachea vertically, the weight of the tissue may have a similar effect of the residual negative pressure in the thorax, responsible for maintaining the residual volume.

Influence of chest wall on lung mechanics during inspiration.

RATIONALE: The interaction between lungs and chest wall influences lung volume, that determines lung history during respiration cycle. In this study, the influence of chest wall mechanics on respiratory system is assessed by the evaluation of inspiration pressure-volume curve (PV curve) under three different situations: closed-chest, open-chest and isolated lung. The PV curve parameters in each situation allow us to further understand the role played by different chest wall elements in the respiratory function. Methods: Twenty-four male Wistar rats (236 ± 29 g) were used. The animals were weighted and then anesthetized with xylazine 2% (0,5mL/kg) and ketamine 10% (0,9mL/kg), exsanguinated and later tracheostomized with a metallic cannula (14 gauge). The cannula was connected to an automatic small animal insufflator. This setup was connected to a pressure transducer (32 samples/s). The 24 animals were randomly separated in three groups: (i) closed chest, (ii) open chest and (iii) isolated lung. The rats were insufflated with 20mL quasi-statically (constant speed of 0,1mL/s). Insufflated volume and measured pressure data were kept and PV curves were obtained for all animals. The PV curves were fitted (non-linear least squares) against the sigmoid equation (1) to obtain the sigmoid equation parameters (a,b,c,d). Elastance measurements were obtained from linear regression of pressure/volume measurements in a 0,8s interval before and after the calculated point. Results: The parameters a, b and c showed no significant change, but the parameter d showed a significant variation among the three groups. The initial elastance also varied between open and closed chest, indicating the need of a higher pressure for the lung expansion, as can be seen in Table 1. Table 1: Mean and Standard Deviation of parameters obtained for each protocol. Protocol: Closed Chest – a (mL) -0.35±0.33; b (mL) 13.93±0.89; c (cm H2O) 21.28±2.37; d (cm H2O) 6.17±0.84; r²** (%) 99.4±0.14; Initial Elastance* (cm H2)/mL) 12.72±6.66; Weight (g) 232.33±5.72. Open Chest - a (mL) 0.01±0.28; b (mL) 14.79±0.54; c (cm H2O) 19.47±1.41; d (cm H2O) 3.50±0.28; r²** (%) 98.8±0.34; Initial Elastance* (cm H2)/mL) 28.68±2.36; Weight (g) 217.33±7.97. Isolated Lung - a (mL) -0.09±0.46; b (mL) 14.22±0.75; c (cm H2O) 21.76±1.43; d (cm H2O) 4.24±0.50; r²** (%) 98.9±0.19; Initial Elastance* (cm H2)/mL) 7.13±8.85; Weight (g) 224.33±16.66. * Elastance measures in the 0-0,1 mL range. ** Goodness of sigmoid fit versus measured data Conclusion: A supporting effect of the chest wall was observed at the initial moments of inspiration, observed as a higher initial elastance in open chest situations than in closed chest situations (p=0,00001). The similar initial elastance for the isolated lung and closed chest may be explained by the specific method used for the isolated lung experiment. As the isolated lung is supported by the trachea vertically, the weight of the tissue may have a similar effect of the residual negative pressure in the thorax, responsible for maintaining the residual volume.

Outcome respiratorio a lungo termine nei soggetti affetti da cardiopatia congenita sottoposti ad intervento cardiochirurgico

Introduction: In the last years cardiac surgery for congenital heart disease (CHD) reduced dramatically mortality modifying prognosis, but, at the same time, increased morbidity in this patient population. Respiratory and cardiovascular systems are strictly anatomically and functionally connected, so that alterations of pulmonary hemodynamic conditions modify respiratory function. While very short-term alterations of respiratory mechanics after surgery were investigated by many authors, not as much works focused on long-term changes. In these subjects rest respiratory function may be limited by several factor: CHD itself (fetal pulmonary perfusion influences vascular and alveolar development), extracorporeal circulation (CEC), thoracotomy and/or sternotomy, rib and sternal contusions, pleural adhesions and pleural fibrosis, secondary to surgical injury. Moreover inflammatory cascade, triggered by CEC, can cause endothelial damage and compromise gas exchange. Aims: The project was conceived to 1) determine severity of respiratory functional impairement in different CHD undergone to surgical correction/palliation; 2) identify the most and the least CHD involved by pulmonary impairement; 3) find a correlation between a specific hemodynamic condition and functional anomaly, and 4) between rest respiratory function and cardiopulmonary exercise test. Materials and methods: We studied 113 subjects with CHD undergone to surgery, and distinguished by group in accord to pulmonary blood flow (group 0: 28 pts with normal pulmonary flow; group 1: 22 pts with increased flow; group 2: 43 pts with decreased flow; group 3: 20 pts with total cavo-pulmonary anastomosis-TCPC) followed by the Pediatric Cardiology and Cardiac Surgery Unit, and we compare them to 37 age- and sex-matched healthy subjects. In Pediatric Pulmonology Unit all pts performed respiratory function tests (static and dynamic volumes, flow/volume curve, airway resistances-raw- and conductance-gaw-, lung diffusion of CO-DLCO- and DLCO/alveolar volume), and CHD pts the same day had cardiopulmonary test. They all were examined and had allergological tests, and respiratory medical history. Results: restrictive pattern (measured on total lung capacity-TLC- and vital capacity-VC) was in all CHD groups, and up to 45% in group 2 and 3. Comparing all groups, we found a significant difference in TLC between healthy and group 2 (p=0.001) and 3 (p=0.004), and in VC between group 2 and healthy (p=0.001) and group 1(p=0.034). Inspiratory capacity (IC) was decreased in group 2 related to healthy (p<0.001) and group 1 (p=0.037). We showed a direct correlation between TLC and VC with age at surgery (p=0.01) and inverse with number of surgical interventions (p=0.03). Reduced FEV1/FVC ratio, Gaw and increased Raw were mostly present in group 3. DLCO was impaired in all groups, but up to 80% in group 3 and 50% in group 2; when corrected for alveolar volume (DLCO/VA) reduction persisted in group 3 (20%), 2 (6.2%) and 0 (7.1%). Exercise test was impaired in all groups: VO2max and VE markedly reduced in all but especially in group 3, and VE/VCO2 slope, marker of ventilatory response to exercise, is increased (<36) in 62.5% of group 3, where other pts had anyway value>32. Comparing group 3 and 2, the most involved categories, we found difference in VO2max and VE/VCO2 slope (respectively p=0.02 and p<0.0001). We evidenced correlation between rest and exercise tests, especially in group 0 (between VO2max and FVC, FEV1, VC, IC; inverse relation between VE/VCO2slope and FVC, FEV1 and VC), but also in group 1 (VO2max and IC), group 2 (VO2max and FVC and FEV1); never in group 3. Discussion: According with literature, we found a frequent impairment of rest pulmonary function in all groups, but especially in group 2 and 3. Restrictive pattern was the most frequent alteration probably due to compromised pulmonary (vascular and alveolar) development secondary to hypoperfusion in fetal and pre-surgery (and pre-TCPC)life. Parenchymal fibrosis, pleural adhesions and thoracic deformities can add further limitation, as showed by the correlation between group 3 and number of surgical intervention. Exercise tests were limited, particularly in group 3 (complex anatomy and lost of chronotropic response), and we found correlations between rest and exercise tests in all but group 3. We speculate that in this patients hemodynamic exceeds respiratory contribution, though markedly decreased.

Studio della funzionalità respiratoria in soggetti con sindrome di Turner

The aim of this study is to evaluate the pulmonary function in subjects with diagnosis of Turner Syndrome, in charge at the Syndromology Ward of the Paediatric Clinic of S.Orsola-Malpighi hospital. There are very few datas about lung function in patients with Turner syndrome’s genotype and phenotype in medical literature. Since the thorax of these subjects have peculiar anatomic shape (as “shield” or “overturned triangle”), we presupposed that these subjects could have also a peculiar respiratory function. Moreover we look for the possibility of correlation between pulmonary function and estroprogestinic replacement therapy and/or growth hormone (GH) replacement therapy. Material and methods: we studied 48 patients, with diagnosis of Turner Syndrome; they all made spirometry voluntarily and, when capable, also plethismografy. Results: - the parametres of pulmonary function are a little higher of the predicted values for age and sex but they are a little lower if they're corrected for each patient’s ideal high and weight: so we can conclude that in Turner Syndrme subjects pulmonary function is normal; -there’s not a statistically significant correlation between pulmonary function and GH therapy; -there’s not a statistically significant correlation between GH therapy’s length and pulmonary function except for Total Lung Capacity which increases with the number of years of GH therapy; - there’s not a statistically significant correlation between pulmonary function and estroprogestinic replacement herapy.

Assessment of bronchodilator responsiveness in preschool children using forced oscillations

BACKGROUND: The forced oscillation technique (FOT) requires minimal patient cooperation and is feasible in preschool children. Few data exist on respiratory function changes measured using FOT following inhaled bronchodilators (BD) in healthy young children, limiting the clinical applications of BD testing in this age group. A study was undertaken to determine the most appropriate method of quantifying BD responses using FOT in healthy young children and those with common respiratory conditions including cystic fibrosis, neonatal chronic lung disease and asthma and/or current wheeze. METHODS: A pseudorandom FOT signal (4-48 Hz) was used to examine respiratory resistance and reactance at 6, 8 and 10 Hz; 3-5 acceptable measurements were made before and 15 min after the administration of salbutamol. The post-BD response was expressed in absolute and relative (percentage of baseline) terms. RESULTS: Significant BD responses were seen in all groups. Absolute changes in BD responses were related to baseline lung function within each group. Relative changes in BD responses were less dependent on baseline lung function and were independent of height in healthy children. Those with neonatal chronic lung disease showed a strong baseline dependence in their responses. The BD response in children with cystic fibrosis, asthma or wheeze (based on both group mean data and number of responders) was not greater than in healthy children. CONCLUSIONS: The BD response assessed by the FOT in preschool children should be expressed as a relative change to account for the effect of baseline lung function. The limits for a positive BD response of -40% and 65% for respiratory resistance and reactance, respectively, are recommended.

Translational regulation of nuclear gene COX4 expression by mitochondrial content of phosphatidylglycerol and cardiolipin in Saccharomyces cerevisiae.

Previous results indicated that translation of four mitochondrion-encoded genes and one nucleus-encoded gene (COX4) is repressed in mutants (pgs1Delta) of Saccharomyces cerevisiae lacking phosphatidylglycerol and cardiolipin. COX4 translation was studied here using a mitochondrially targeted green fluorescence protein (mtGFP) fused to the COX4 promoter and its 5' and 3' untranslated regions (UTRs). Lack of mtGFP expression independent of carbon source and strain background was established to be at the translational level. The translational defect was not due to deficiency of mitochondrial respiratory function but was rather caused directly by the lack of phosphatidylglycerol and cardiolipin in mitochondrial membranes. Reintroduction of a functional PGS1 gene under control of the ADH1 promoter restored phosphatidylglycerol synthesis and expression of mtGFP. Deletion analysis of the 5' UTR(COX4) revealed the presence of a 50-nucleotide fragment with two stem-loops as a cis-element inhibiting COX4 translation. Binding of a protein factor(s) specifically to this sequence was observed with cytoplasm from pgs1Delta but not PGS1 cells. Using HIS3 and lacZ as reporters, extragenic spontaneous recessive mutations that allowed expression of His3p and beta-galactosidase were isolated, which appeared to be loss-of-function mutations, suggesting that the genes mutated may encode the trans factors that bind to the cis element in pgs1Delta cells.

The effectiveness of Alexander Technique on music performance and musicians’ health and well-being – a systematic review

Introduction Musicians often suffer injuries related to their music playing. Therefore, some use Alexander Technique (AT), a mental-physical method that facilitates to release unnecessary muscle tension and to re-educate non-beneficial movement patterns through enhanced kinaesthetic awareness. According to a recent review AT may be effective for chronic back pain [1]. This review aimed to evaluate the evidence for the effectiveness of AT lessons on music performance and musicians’ health and well-being. Methods The following electronic databases were searched up to July 2012 for relevant literature: PUBMED, Google Scholar, CINAHL and EMBASE. The search criteria were "Alexander technique" AND "music*" [all fields]. References were searched, and experts and societies of AT or musicians' medicine contacted for further publications. Results 100 studies were identified. 24 studies were included for further analysis, 5 of which were randomised controlled trials (RCTs), 5 controlled but not randomised (CTs), 5 without control group, 2 mixed methods (RCT and case studies), and 7 surveys. 13 to 72 musicians participated per RCT. In 5 RCTs AT groups received between 12 and 20 one-to-one lessons. In 4 RCTs control groups received no interventions. Primary outcomes were performance anxiety, music performance, "use" as well as respiratory function and pain. Performance anxiety decreased by AT in 3 of 4 RCTs and in 3 of 3 CTs. Music performance was improved by AT in 1 RCT, yet in 2 RCTs comparing neurofeedback (NF) to AT, only NF showed improvements. Discussion and Conclusion To investigate the effectiveness of AT in musicians a variety of study designs and outcome measures have been used. Evidence from RCTs suggests that AT may improve performance anxiety in musicians. Effects on music performance, body use and respiratory function yet remain inconsistent. Future trials with scientifically sound study designs are warranted to further and more reliably explore the potential of AT as a relatively low cost and low risk method in the interest of musicians. References [1] Woodman JP, Moore NR. Evidence for the effectiveness of Alexander Technique lessons in medical and health-related conditions: a systematic review. Int J Clin Pract 2012;66(1):98-112.

The Effectiveness of Alexander Technique on Music Performance and Musicians' Health and Well-being – a Systematic Review

Purpose Musicians often suffer injuries related to their music playing. Therefore, some use the Alexander Technique (AT), a psycho-physical method that helps to release unnecessary muscle tension and re-educates non-beneficial movement patterns through enhanced kinaesthetic awareness. According to a recent review AT may be effective for chronic back pain. This review aimed to evaluate the evidence for the effectiveness of AT lessons on music performance and musicians’ health and well-being. Methods The following electronic databases were searched up to July 2012 for relevant literature: PUBMED, Google Scholar, CINAHL and EMBASE. The search criteria were "Alexander technique" AND "music*" [all fields]. References were searched, and experts and societies of AT or musicians' medicine contacted for further publications. Results 100 studies were identified. 35 studies were included for further analysis, 5 of which were randomised controlled trials (RCTs), 5 controlled but not randomised, 5 not controlled, 5 qualitative case studies, 2 mixed-models (RCT and case studies), 7 surveys, 4 qualitative case reports and 2 unpublished pilot studies. 13 to 72 musicians participated per RCT. In 5 RCTs AT groups received between 12 and 20 one-to-one lessons. In 4 RCTs control groups received no interventions. Primary outcomes were performance anxiety, performance, "use" as well as respiratory function and pain. Performance anxiety decreased by AT in 3 of 4 RCTs. Music performance was improved by AT in 1 RCT, yet in 2 RCTs comparing neurofeedback (NF) to AT, only NF showed improvements. Conclusions To investigate the effectiveness of AT in musicians a variety of study designs and outcome measures have been used. Evidence from RCTs suggests that AT may improve performance anxiety in musicians. Effects on music performance, body use and respiratory function yet remain inconsistent. Future trials with well-established study designs are warranted to further and more reliably explore the potential of AT as a relatively low cost and low risk method in the interest of musicians.

The Alexander Technique and musicians: a systematic review of controlled trials

BACKGROUND: Musculoskeletal disorders, stress and performance anxiety are common in musicians. Therefore, some use the Alexander Technique (AT), a psycho-physical method that helps to release unnecessary muscle tension and re-educates non-beneficial movement patterns through intentional inhibition of unwanted habitual behaviours. According to a recent review AT sessions may be effective for chronic back pain. This review aimed to evaluate the evidence for the effectiveness of AT sessions on musicians' performance, anxiety, respiratory function and posture. METHODS: The following electronic databases were searched up to February 2014 for relevant publications: PUBMED, Google Scholar, CINAHL, EMBASE, AMED, PsycINFO and RILM. The search criteria were "Alexander Technique" AND "music*". References were searched, and experts and societies of AT or musicians' medicine contacted for further publications. RESULTS: 237 citations were assessed. 12 studies were included for further analysis, 5 of which were randomised controlled trials (RCTs), 5 controlled but not randomised (CTs), and 2 mixed methods studies. Main outcome measures in RCTs and CTs were music performance, respiratory function, performance anxiety, body use and posture. Music performance was judged by external experts and found to be improved by AT in 1 of 3 RCTs; in 1 RCT comparing neurofeedback (NF) to AT, only NF caused improvements. Respiratory function was investigated in 2 RCTs, but not improved by AT training. Performance anxiety was mostly assessed by questionnaires and decreased by AT in 2 of 2 RCTs and in 2 of 2 CTs. CONCLUSIONS: A variety of outcome measures have been used to investigate the effectiveness of AT sessions in musicians. Evidence from RCTs and CTs suggests that AT sessions may improve performance anxiety in musicians. Effects on music performance, respiratory function and posture yet remain inconclusive. Future trials with well-established study designs are warranted to further and more reliably explore the potential of AT in the interest of musicians.

Cessation and resuming of alglucosidase alfa in Pompe disease: a retrospective analysis.

Enzyme replacement therapy (ERT) with recombinant human alglucosidase alfa (rhGAA) in late-onset Pompe disease is moderately effective. Little is known about the clinical course after treatment termination and the resumption of ERT. In Switzerland, rhGAA therapy for Pompe disease was temporarily withdrawn after the federal court judged that the treatment costs were greatly out of proportion compared to the benefits. Re-treatment was initiated after the therapy was finally licensed. We retrospectively analysed seven Pompe patients, who underwent cessation and resumption of ERT (median age 43 years). The delay from first symptoms to final diagnosis ranged from 4 to 20 years. The demographics, clinical characteristics, assessments with the 6-min walking test (6-MWT), the predicted forced vital capacity (FVC) and muscle strength were analysed. Before initiation of ERT, all patients suffered from proximal muscle weakness of the lower limbs; one was wheelchair-bound and two patients received night-time non-invasive ventilation. Initial treatment stabilised respiratory function in most patients and improved their walking performance. After treatment cessation, upright FVC declined in most and the 6-MWT declined in all patients. Two patients needed additional non-invasive ventilatory support. Twelve months after resuming ERT, the respiratory and walking capacity improved again in most patients. However, aside for one patient, none of the patients reached the same level of respiratory function or distance walked in 6 min, as at the time of ERT withdrawal. We conclude that cessation of ERT in Pompe disease causes a decline in clinical function and should be avoided. Resuming treatment only partially recovers respiratory function and walking capacity.

Potential Role of Adenosine Signaling in Acetic Acid Activation of Murine Sensory Neurons

Chronic respiratory illnesses are a significant cause of morbidity and mortality, and acute changes in respiratory function often lead to hospitalization. Air pollution is known to exacerbate asthma, but the molecular mechanisms of this are poorly understood. The current studies were aimed at clarifying the roles of nerve subtypes and purinergic receptors in respiratory reflex responses following exposure to irritants. In C57Bl/6J female mice, inspired adenosine produced sensory irritation, shown to be mediated mostly by A-delta fibers. Secondly, the response to inhaled acetic acid was discovered to be dually influenced by C and A-delta fibers, as indicated by the observed effects of capsaicin pretreatment, which selectively destroys TRPV1-expressing fibers (mostly C fibers) and pretreatment with theophylline, a nonselective adenosine receptor antagonist. The responses to both adenosine and acetic acid were enhanced in the ovalbumin-allergic airway disease model, although the particular pathway altered is still unknown.

Mitochondrial dysfunction leads to NOX activation: A novel mechanism to maintain high glycolysis in cancer cells

Increased glycolysis and oxidative stress are common features of cancer cells. These metabolic alterations are associated with mitochondrial dysfunction and can be caused by mitochondrial DNA (mtDNA) mutations, oncogenic signals, loss of tumor suppressor, and tumor tissue hypoxia. It is well established that mitochondria play central roles in energy metabolism, maintenance of redox balance, and regulation of apoptosis. However, the biochemical and molecular mechanisms that maintain high glycolysis in cancer cells (the Warburg effect) with mitochondrial dysfunction and oxidative stress remain to be determined. The major goals of this study were to establish a unique experimental system in which the mitochondrial respiratory function can be regulated as desired, and to use this system to investigate the mechanistic link between mitochondrial dysfunction and the Warburg effect along with oxidative stress in cancer cells. To achieve these goals, I have established a tetracycline-inducible system in which a dominant negative form of mitochondrial DNA polymerase y (POLGdn) expression could be regulated by tetracycline; thus controlling mitochondrial respiratory function. Using this cell system, I demonstrated that POLGdn expression resulted in mitochondrial dysfunction through decreasing mtDNA content, depletion of mtDNA encoded mRNA and protein expression. This process was mediated by TFAM proteasome degradation. Mitochondrial dysfunction mediated by POLGdn expression led to a significant increase in cellular glycolysis and oxidative stress. Surprisingly, mitochondrial dysfunction also resulted in increased NAD(P)H oxidase (NOX) enzyme activity, which was shown to be essential for maintaining high glycolysis. Chemical Inhibition of NOX activity by diphenyliodonium (DPI) preferentially impacted the survival of mitochondrial defective cells. The colon cancer HCT116-/- cells that have lost transcriptional regulation of the mitochondrial assembling enzyme SCO2, leading to compromised mitochondrial respiratory function, were found to have increased NOX activity and were highly sensitive to DPI treatment. Ovarian epithelial cells with Ras transformation also exhibited an increase in NOX gene expression and NOX enzyme activity, rendering the cells sensitive to DPI inhibition especially under hypoxic condition. These data together suggest that NOX plays a novel role in maintaining high glycolysis in cancer cells with mitochondrial defects, and that NOX may be a potential target for cancer therapy. ^

Expanding the functional human mitochondrial DNA database by the establishment of primate xenomitochondrial cybrids

The nuclear and mitochondrial genomes coevolve to optimize approximately 100 different interactions necessary for an efficient ATP-generating system. This coevolution led to a species-specific compatibility between these genomes. We introduced mitochondrial DNA (mtDNA) from different primates into mtDNA-less human cells and selected for growth of cells with a functional oxidative phosphorylation system. mtDNA from common chimpanzee, pigmy chimpanzee, and gorilla were able to restore oxidative phosphorylation in the context of a human nuclear background, whereas mtDNA from orangutan, and species representative of Old-World monkeys, New-World monkeys, and lemurs were not. Oxygen consumption, a sensitive index of respiratory function, showed that mtDNA from chimpanzee, pigmy chimpanzee, and gorilla replaced the human mtDNA and restored respiration to essentially normal levels. Mitochondrial protein synthesis was also unaltered in successful “xenomitochondrial cybrids.” The abrupt failure of mtDNA from primate species that diverged from humans as recently as 8–18 million years ago to functionally replace human mtDNA suggests the presence of one or a few mutations affecting critical nuclear–mitochondrial genome interactions between these species. These cellular systems provide a demonstration of intergenus mtDNA transfer, expand more than 20-fold the number of mtDNA polymorphisms that can be analyzed in a human nuclear background, and provide a novel model for the study of nuclear–mitochondrial interactions.